Preface

As mentioned in the ancient textbooks of Ayurvedic medicine, inflammation was known to Ayurvedic physicians of the Indian Peninsula already back in 1 500 BCE and in 600 CE as a manifestation of a disease. They characterized inflammation as an elevation, swelling or edema, heaviness, and pain. After this, in 5 CE, a Greek physician introduced the term edema for inflammation. Later on, Aulus Cornelius Celsus (30 BCE–38 CE) described inflammation with four cardinal signs, that is, redness (*rubor*), elevated heat (*calor*), swelling (*tumor*), and pain (*dolor*), as occurring during acute inflammation in response to the localized acute infection or trauma. These four signs are known as Celsus tetrad of inflammation. Galen and later on Virchow (1871), with a more precise definition, introduced the fifth sign of inflammation called loss of function (*functio lasea*) of the affected organ or tissue. The modern term inflammation is derived from the Greek word *inflammare,*  meaning to set on fire.

The present book aims to discuss in detail the pathogenesis of inflammation, in a sterile context and infection-induced inflammation along with advances in its management. The 1st chapter "*Introductory Chapter: The Journey of Inflammation and Inflammatory Disease Research - Past, Present, and Future*" discusses the history, present studies and future directions of research focused on inflammation and inflammatory diseases. The 2nd chapter titled "*Prolouge: Initial Approach to Edema*" introduces the topic of edema. The 3rd chapter titled "*Peripheral Edema: Differential Diagnosis*" describes the pathogenesis of peripheral edema, including the lower limb edema in different conditions, differential diagnosis, and management. The 4th chapter "*Edema Induced by sPLA2 from* Crotalus durissus terrificus *Involves PLC and PKC Signaling, Activation of cPLA2, and Oxidative Stress*" discusses the pathogenesis of edema induced by *Crotalus durissus terrificus* involving phospholipase C (PLC) and protein kinase C (PKC) as signaling molecules. The 5th chapter "*Edema Management in Oral and Maxillofacial Surgery*" discusses the management of edema observed during oral and maxillofacial surgery. This chapter is crucial as edema is seen in patients who underwent surgery. The 6th chapter "*Neutrophil Counts and Rates in Otorhinolaryngology*" discusses the importance of neutrophils (potent innate immune cells playing a crucial role in the inflammatory process) in otorhinolaryngology. The 7th chapter "*Neutrophil Gelatinase-Associated Lipocalin as a Promising Biomarker in Acute Kidney Injury*" discusses the use of neutrophil gelatinase-associated lipocalin (NGAL) as a potential marker for acute kidney injury (AKI). The 8th chapter "*The Role of Neutrophil Extracellular Traps (NETs) in the Pathogenesis and Complications of Malignant Diseases*" discusses the importance of neutrophil extracellular traps (NETs) in the pathogenesis and complications of malignant diseases. NETs are large, extracellular web-like structures comprised of cytosolic and granule proteins assembled on the scaffold of decondensed chromatin, having both nuclear and mitochondrial DNA, which trap circulating pathogens during microbial infections. The 9th chapter "*The Role of Introns for the Development of Inflammation-Mediated Cancer Cell*" is of importance and discusses the role of introns in the transformation of normal cell into cancer cell during inflammation. This chapter is intended to discuss the genetic changes involved in the transformation of normal cell into cancer cell during chronic inflammation,

as chronic inflammation is one of the causal factors for cancer development. The 10th chapter "*Celiac Disease*" discusses celiac disease (a chronic inflammatory disease of the gut) which incidence is increasing due to modern lifestyle. The 11th chapter "*Sialoendoscopy in Juvenile Recurrent Parotitis That Could Be Primary Pediatric Sjogren's Syndrome*" discusses the use of sialoendoscopy in juvenile recurrent parotitis that may be a primary pediatric Sjögren's syndrome (an autoimmune disease). The 12th chapter "*Inflammation in the Pathogenesis of Rheumatoid Arthritis and in Experimental Arthritis: Evaluation of Combinations of Carnosic Acid and Extract of* Rhodiola rosea *L. with Methotrexate*" discusses the use of carnosic acid and extract of *Rhodiola rosea* L. with methotrexate in rheumatoid arthritis (RA), an autoimmune disease primarily affecting small joints, and the involvement of inflammation as the major disease mechanism. The 13th chapter "*Antiphospholipid Syndrome and Pregnancy-Diagnosis, Complications and Management: An Overview*" discusses the impact of antiphospholipid syndrome (another autoimmune disease) in the pregnancy outcome. The 14th chapter "*Non-Allergic Rhinitis*" discusses nonallergic rhinitis that occurs independently of allergen exposure and the diagnosis and treatment options. The 15th chapter "*Islet Inflammation: The Link between Type 2 Diabetes and Pancreatic Cancer*" discusses the islet inflammation (inflammation of pancreatic cells involved in the insulin secretion) that plays a crucial role in the pathogenesis of type 2 diabetes mellitus (T2DM). This chapter links T2DM and pancreatic cancer as results of chronic pancreatic inflammation. The 16th chapter "*Hypomelanosis Secondary to Cutaneous Inflammation*" discusses hypomelanosis and the disease links to the different cutaneous inflammatory conditions. The 17th chapter "*Helminth Induced Immunomodulation against Metainflammation and Insulin Resistance*" discusses the suppression of chronic inflammation associated with meta-inflammation and T2DM via helminthic infections through inducing immunomodulation. Hence, the current book will prove beneficial to audience interested in the wide aspect of inflammation and inflammatory diseases.

> **Vijay Kumar** Department of Pharmaceutical Sciences, College of Pharmacy, University of Tennessee Health Science Center (UTHSC), Memphis, TN, USA

> > **Alexandro Aguilera Salgado** National Institute of Pediatrics, Mexico

**Seyyed Shamsadin Athari** Zanjan University of Medical Sciences, Iran

Section 1 Introduction

#### **Chapter 1**

## Introductory Chapter: The Journey of Inflammation and Inflammatory Disease Research - Past, Present, and Future

*Vijay Kumar*

#### **1. Introduction**

**Inflammation** is known to ancient Indian Ayurvedic physicians dating back to 1500 BC that is well described in ancient Ayurvedic medicine textbooks (*Charaka Samhita*, *Susruta Samhita,* and *Astanga Samgraha)* [1]. Inflammation and the associated edema have gotten attention in Ayurveda as a pathological manifestation of the disease [2, 3]. Ayurvedic medicine mentioned inflammation in different ways, which include *Shotha and Shopha, and* many other terms *(Svayathu, Utsedha,* and *Samhata)* [2]*.* Ancient Ayurvedic medicine practitioners characterized inflammation in different ways, including elevation, edema, heaviness, and pain. The Greek physician, Hippocrates in the 5th century BC coined the term edema. Furthermore, Later on, Aulus Celsus (30 BC-38 AD) described the four major signs of inflammation, which include *Rubor*, *Calor*, *Tumor*, and *Dolor* [4]. Galen, the Roman physician introduced the fifth sign of inflammation as *loss of function*. However, *Virchow* in 1871 more precisely described the *function laesa* (loss of function) sign of inflammation [5, 6].

The modern terminology of inflammation has been derived from the Latin word *inflammare* (to set on fire) [6]. Now inflammation is considered as a host-generated protective host immune response in response to acute trauma or pathogens or their PAMPs (pathogen-derived molecular patterns) to contain the damage or to remove/ kill the pathogen [7]. The inflammation pathogenesis is a complex process involving the network of cellular and molecular signaling to restore homeostasis and induce tissue/organ repair and regeneration. However, any dysregulation of the inflammatory process may lead to the development of severe inflammatory conditions, including systemic inflammation and sepsis (during infection). Furthermore, any persistent inflammation (staying for months and years) may cause chronic inflammatory disorders (cancer and autoimmune diseases) [8–10]. The cellular components of the immune system (macrophages, neutrophils, mast cells, dendritic cells (DCs), T cells, and B cells etc.) play a central role in the process of both acute and chronic inflammation involve (**Figure 1**) [11–14].

#### **Figure 1.**

*Schematic representation of immune cell activation in response to different pathogens, PAMPs, MAMPs through interacting with different PRRs (mentioned in the text) to generate inflammatory immune response. This immune response is governed by immunometabolic reprograming, epigenetics, miRNAs, and the release of extracellular vesicles or exosomes, which directly or indirectly interact and affect each other.*

#### **2. Pattern recognition receptors (PRRs) in inflammation and inflammatory diseases**

The receptors expressed by these immune cells called **pattern recognition receptors** (PRRs) (**Figure 1**) are present on the outer surface of the cell membrane as well as in the cytosol, including different toll-like receptors (TLR1-TLR13 in mammals, including humans), intracellular PRRs (NOD [Nucleotide-binding and oligomerization domain])-like receptors (NLRs; NOD1 and NOD2), absent in melanoma-like receptors (ALRs, AIM2), and many more mentioned somewhere else recognize the potential pathogen and/or inflammogen to mount a protective inflammatory immune response [1, 15–17]. The intracellular proteins (NLRC1 or NLPR1, NLRP3, NLRC4, pyrin, and AIM2 or absent in melanoma 2 that recognizes cytosolic DNA) form an inflammatory complex called inflammasome upon recognition of intracellular threat (damage-associated molecular patterns or DAMPs). The inflammasome also becomes activated upon the recognition of external danger called pathogen or microbe-associated molecular patterns (PAMPs or MAMPs) and DAMPs by cell surface PRRs, which signal these inflammasome proteins to activate and stimulate another cascade of inflammation resulting in the release of pro-inflammatory cytokines (IL-1β, IL-18, and IL-33) and the cell death called pyroptosis that further aggravates the inflammatory process [18]. The details of inflammasomes in inflammation and inflammatory disease are

*Introductory Chapter: The Journey of Inflammation and Inflammatory Disease... DOI: http://dx.doi.org/10.5772/intechopen.101512*

mentioned somewhere else [18, 19]. The activation of cell surface PRRs, including TLRs (TLR2 and TLR4), may activate the inflammasome or TLRs and inflammasomes work in cooperation to control the inflammatory process [20, 21]. Another, cytosolic PRR system called cGAS (cyclic GMP-AMP synthase, recognizes cytosolic dsDNA)-stimulator of interferon genes (STING) pathway recognizes cytosolic dsDNA and induces the synthesis of type 1 interferons (IFNs) [22, 23]. The over-activation of this pathway is involved in different autoimmune and autoinflammatory diseases, along with other inflammatory conditions discussed in detail somewhere else [22–24]. The details of inflammatory pathways mediated by TLRs, inflammasomes, cGAS, and other PRRs have been discussed by the author somewhere else in detail [1, 18, 21, 22, 24–27].

#### **3. Immunometabolism in inflammation and inflammatory diseases**

Like, other non-immune cells, immune cells also have their energy demand that plays a crucial role in the maintenance of immune homeostasis and the mounting of the immune response to protect against invading foreign agents, including the pathogen and allergen. The metabolic changes occurring in immune cells from their normal/control stage (absence of inflammogen, pathogen, PAMPs, MAMPs, or DAMPs) to their activation or activated stage is called immunometabolic reprogramming [28]. Hence, the metabolic pathways governing or regulating the energy demand of immune cells to maintain immune homeostasis is called **immunometabolism** [28]. The metabolic demand of immune cells increases during the inflammatory process and reprogramming of different metabolic pathways governing the immune function takes place that depends on the immune cell type and the inflammatory conditions (**Figure 1**) [29]. The author has described immunometabolism of different immune cells and their immunometabolic reprogramming and its therapeutic targeting during inflammation and inflammatory diseases elsewhere [1, 30–35].

#### **4. Epigenetics in inflammation and inflammatory diseases**

**Epigenetics** (deals with the reversible impact of behavior and environmental factors on our genetic machinery without changing the DNA sequence. However, it may change the way of reading the genetic information coded by the DNA) also plays a crucial role in the inflammatory process and inflammatory diseases. The most frequent epigenetic changes involve aberrant DNA methylation and histone acetylation and deacetylation. The enzymes (arginine and lysine methyltransferases, DNA methyltransferase, histone acetyltransferases (HAT), and histone deacetylases or HDAC) involved in the process of epigenetics also control the inflammatory process, including airway inflammation, atopic dermatitis, and autoimmune diseases [36–41]. The histone modifications, DNA methylation, and noncoding RNAs (ncRNAs) have emerged as master regulators of gene expression, including the inflammatory genes [42]. The targeting of these enzymes, including HAT and HDAC, has shown great antiinflammatory potential in diverse inflammatory diseases. However, advances in the biological sciences have shown the interaction between epigenetics and immunometabolism converges at inflammation and regulates different inflammatory diseases, including cancer (**Figure 1**) [42–45]. Hence, epigenetics-based immunotherapies are emerging for targeting chronic inflammatory diseases, including cancer and autoimmunity [46].

#### **5. Extracellular vesicles in inflammation and inflammatory diseases**

**Extracellular vesicles (EVs)** are generated by different cell types, including the immune cells to remove cellular waste and communicate with adjacent as well as distant cells [47]. These EVs may contain protein, DNA, RNA, micro RNA (miRNA or miRs), and cytokines depending on the cell type and cell/tissue microenvironment. The microvesicles (MVs), a kind of EVs released from apoptotic cells are lessinflammatory than those released from viable cells [48]. These MVs have different miRs **(**miR-155, miR-34b, and miR-34a), which get dysregulated in autoimmune diseases, including systemic lupus erythematosus (SLE) in comparison to normal individuals. EVs play a crucial role in cell–cell communication in pulmonary inflammation upon exposure to toxicants [49]. EVs also affect immunometabolism during diverse inflammatory conditions, including autoimmunity (**Figure 1**) [50, 51]. The different types of EVs, their contents (miRs), and their role in inflammation and therapeutic potential, including in sepsis and coronavirus disease-2019 (COVID-19) have been discussed somewhere else [47, 52–56].

#### **6. MicroRNAs (miRNA) in inflammation and inflammatory diseases**

**MicroRNAs (miRNAs)** are small non-coding RNAs (typical length, 18–24 nucleotide long), which are crucial in regulating protein-coding genes via posttranscriptional repression [57]. They play an important role in regulating innate and adaptive immunity from their developmental stages to function during diverse inflammation conditions, including cancer and autoimmunity as fine-tuners of the system [57–60]. For example, miR-181a and miR-223 are crucial in the establishment and maintenance of immune cell fate [58]. The miR-146 also regulates innate immunity through controlling TLR signaling and ensuing cytokine response. They (miR-155 and miR-181a) regulate central elements of the adaptive immune response such as antigen presentation and T cell receptor (TCR) signaling [58]. Chronic inflammatory diseases exhibit altered miR (miR-203 and miR-146) levels, indicating their crucial role in immunological pathologies/diseases. The details of miRs in immunity and inflammation are discussed somewhere else [59, 61]. The emerging evidences have shown the regulation of immune cell metabolism or immunometabolism by miRs [62, 63]. The non-coding RNAs (ncRNAs) also regulate inflammasome activity controlling the inflammatory immune response [64]. More recently an atlas of miR expression in 63 different mouse immune cell populations has been generated and connected with an assay for transposaseaccessible chromatin using sequencing (ATAC–seq), chromatin immunoprecipitation followed by sequencing (ChIP–seq), and nascent RNA profiles to establish a map of miRNA promoter and enhancer usage in immune cells [65]. This will help to delineate the *cis*-regulatory elements controlling miRNA signatures of the immune system.

#### **7. Conclusion**

The story of inflammation had started from the Ancient Indian peninsula through the Ayurvedic medicine that further developed into its four peculiar signs (*rubor, tumor, calor, and dolor*) and fifth end-stage sign indicating the loss of function. The development in biomedical sciences, including immunology, cell signaling, pharmacology, epigenetics, and molecular biology or medicine has helped to understand the pathogenesis of inflammation (both, acute, and chronic) and

*Introductory Chapter: The Journey of Inflammation and Inflammatory Disease... DOI: http://dx.doi.org/10.5772/intechopen.101512*

associated inflammatory disease, varying from autoimmunity to cancer to severe infectious diseases, including the current COVID-19 pandemic. Thus, the long journey of inflammation that started dating back to 1500 BC and 600 AD has seen significant development in understanding its pathogenesis under diverse conditions and therapeutic advancement. Further studies in the 21st century will open new avenues to control and prevent inflammatory diseases responsible for human morbidity and mortality.

### **Author details**

Vijay Kumar

Department of Pharmaceutical Sciences, College of Pharmacy, University of Tennessee Health Science Center (UTHSC), Memphis, TN, USA

\*Address all correspondence to: vij\_tox@yahoo.com; vkumar7@uthsc.edu

© 2021 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

#### **References**

[1] Kumar V. Inflammation research sails through the sea of immunology to reach immunometabolism. International Immunopharmacology. 2019;**73**:128-145

[2] Ballakur V. Inflammation in ayurveda and modern medicine. International Ayurvedic Medical Journal. 2013;**1**(4):1-7

[3] Rao KVS. Immunology in India: An emerging story. Nature Immunology. 2008;**9**:1319

[4] Rocha e Silva, M. A brief survey of the history of inflammation. 1978. Agents and Actions. 1994;**43**:86-90

[5] Ley K. History of Inflammation Research. In: Ley K, editor. Physiology of Inflammation. New York, New York, NY: Springer; 2001. pp. 1-10

[6] Scott A, Khan KM, Cook JL, Duronio V. What is "inflammation"? Are we ready to move beyond Celsus? British Journal of Sports Medicine. 2004;**38**: 248-249

[7] Fullerton JN, Gilroy DW. Resolution of inflammation: a new therapeutic frontier. Nature Reviews Drug Discovery. 2016;**15**:551

[8] Shalapour S, Karin M. Immunity, inflammation, and cancer: an eternal fight between good and evil. The Journal of Clinical Investigation. 2015;**125**: 3347-3355

[9] Chen Z, Bozec A, Ramming A, Schett G. Anti-inflammatory and immune-regulatory cytokines in rheumatoid arthritis. Nature Reviews Rheumatology. 2019;**15**:9-17

[10] Shen HH, Yang YX, Meng X, Luo XY, Li XM, Shuai ZW, et al. NLRP3: A promising therapeutic target for autoimmune diseases. Autoimmunity Reviews. 2018;**17**:694-702

[11] Kumar V, Ahmad A. Role of MAIT cells in the immunopathogenesis of inflammatory diseases: New players in old game. International Reviews of Immunology. 2017:1-21

[12] Kumar V. Innate lymphoid cells: new paradigm in immunology of inflammation. Immunology Letters. 2014;**157**:23-37

[13] Kumar V, Sharma A. Neutrophils: Cinderella of innate immune system. International Immunopharmacology. 2010;**10**:1325-1334

[14] Kumar V, Sharma A. Mast cells: emerging sentinel innate immune cells with diverse role in immunity. Molecular Immunology. 2010;**48**:14-25

[15] Weiss E, Kretschmer D. Formylpeptide receptors in infection, inflammation, and cancer. Trends in Immunology. 2018;**39**:815-829

[16] Kawai T, Akira S. The roles of TLRs, RLRs and NLRs in pathogen recognitionInt Immunol. 21:313. International Immunology. 2009;**21**: 317-337

[17] Kanneganti T-D, Lamkanfi M, Núñez G. Intracellular NOD-like receptors in host defense and disease. Immunity. 2007;**27**:549-559

[18] Kumar V. Inflammasomes: Pandora's box for sepsis. Journal of Inflammation Research. 2018;**11**:477-502

[19] Guo H, Callaway JB, Ting JPY. Inflammasomes: mechanism of action, role in disease, and therapeutics. Nature Medicine. 2015;**21**:677-687

[20] Hanamsagar R, Hanke ML, Kielian T. Toll-like receptor (TLR) and inflammasome actions in the central nervous system. Trends in Immunology. 2012;**33**:333-342

*Introductory Chapter: The Journey of Inflammation and Inflammatory Disease... DOI: http://dx.doi.org/10.5772/intechopen.101512*

[21] Kumar V. The complement system, toll-like receptors and inflammasomes in host defense: Three musketeers' one target. International Reviews of Immunology. 2019;**38**:131-156

[22] Kumar V. A STING to inflammation and autoimmunity. Journal of Leukocyte Biology. 2019;**106**:171-185

[23] Decout A, Katz JD, Venkatraman S, Ablasser A. The cGAS–STING pathway as a therapeutic target in inflammatory diseases. Nature Reviews Immunology. 2021;**21**:548-569

[24] Kumar V. The Trinity of cGAS, TLR9, and ALRs Guardians of the Cellular Galaxy Against Host-Derived Self-DNA. Frontiers in Immunology. 2020;**11**:624597

[25] Kumar V. Toll-like receptors in immunity and inflammatory diseases: Past, present, and future. International Immunopharmacology. 2018;**59**:391-412

[26] Kumar V. Toll-like receptors in the pathogenesis of neuroinflammation. Journal of Neuroimmunology. 2019;**332**:16-30

[27] Kumar V. Toll-like receptors in sepsis-associated cytokine storm and their endogenous negative regulators as future immunomodulatory targets. International Immunopharmacology. 2020;**89**:107087

[28] O'Neill LAJ, Kishton RJ, Rathmell J. A guide to immunometabolism for immunologists. Nature Reviews Immunology. 2016;**16**:553-565

[29] Voss K, Hong HS, Bader JE, Sugiura A, Lyssiotis CA, Rathmell JC. A guide to interrogating immunometabolism. Nature Reviews Immunology. 2021;**21**:637-652

[30] Kumar V. T cells and their immunometabolism: A novel way to understanding sepsis immunopathogenesis and future

therapeutics. European Journal of Cell Biology. 2018;**97**:379-392

[31] Kumar V. Targeting macrophage immunometabolism: Dawn in the darkness of sepsis. International Immunopharmacology. 2018;**58**:173-185

[32] Kumar V. How could we forget immunometabolism in SARS-CoV2 infection or COVID-19? International Reviews of Immunology. 2021;**40**:72-107

[33] Kumar V. Immunometabolism: Another Road to Sepsis and Its Therapeutic Targeting. Inflammation. 2019;**42**:765-788

[34] Kumar V. Dendritic cells in sepsis: Potential immunoregulatory cells with therapeutic potential. Molecular Immunology. 2018;**101**:615-626

[35] Kumar V. Natural killer cells in sepsis: Underprivileged innate immune cells. European Journal of Cell Biology. 2019;**98**:81-93

[36] Shanmugam MK, Sethi G. Role of epigenetics in inflammation-associated diseases. Sub-Cellular Biochemistry. 2013;**61**:627-657

[37] Adcock IM, Tsaprouni L, Bhavsar P, Ito K. Epigenetic regulation of airway inflammation. Current Opinion in Immunology. 2007;**19**:694-700

[38] Schmidt AD, de Guzman Strong C. Current understanding of epigenetics in atopic dermatitis. Experimental Dermatology. 2021;**30**:1150-1155

[39] Nielsen HM, Tost J. Epigenetic changes in inflammatory and autoimmune diseases. Sub-Cellular Biochemistry. 2013;**61**:455-478

[40] Stylianou E. Epigenetics of chronic inflammatory diseases. Journal of Inflammation Research. 2018;**12**:1-14

[41] Surace AEA, Hedrich CM. The Role of Epigenetics in Autoimmune/ Inflammatory Disease. Frontiers in Immunology. 2019;**10**:1-16

[42] Karin M, Shalapour S. Regulation of antitumor immunity by inflammationinduced epigenetic alterations. Cellular & Molecular Immunology. 2021:1-8

[43] Guzik TJ, Cosentino F. Epigenetics and Immunometabolism in Diabetes and Aging. Antioxidants & Redox Signaling. 2018;**29**:257-274

[44] Raghuraman S, Donkin I, Versteyhe S, Barrès R, Simar D. The emerging role of epigenetics in inflammation and immunometabolism. Trends in Endocrinology and Metabolism. 2016;**27**:782-795

[45] Verberk SGS, de Goede KE, Van den Bossche J. Metabolic–epigenetic crosstalk in macrophage activation: an updated view. Epigenomics. 2019;**11**: 719-721

[46] Dunn J, Rao S. Epigenetics and immunotherapy: The current state of play. Molecular Immunology. 2017;**87**: 227-239

[47] Kumar V, Kiran S, Kumar S, Singh UP. Extracellular vesicles in obesity and its associated inflammation. International Reviews of Immunology. 2021:1-15

[48] Claßen L, Tykocinski LO, Wiedmann F, Birr C, Schiller P, Tucher C, et al. Extracellular vesicles mediate intercellular communication: Transfer of functionally active microRNAs by microvesicles into phagocytes. European Journal of Immunology. 2017;**47**:1535-1549

[49] Andres J, Smith LC, Murray A, Jin Y, Businaro R, Laskin JD, et al. Role of extracellular vesicles in cell-cell communication and inflammation following exposure to pulmonary

toxicants. Cytokine & Growth Factor Reviews. 2020;**51**:12-18

[50] de Candia P, De Rosa V, Gigantino V, Botti G, Ceriello A, Matarese G. Immunometabolism of human autoimmune diseases: from metabolites to extracellular vesicles. FEBS Letters. 2017;**591**:3119-3134

[51] Zhou Z, Tao Y, Zhao H, Wang Q. Adipose extracellular vesicles: Messengers from and to macrophages in regulating immunometabolic homeostasis or disorders. Frontiers in Immunology. 2021;**12**:666344-666344

[52] Buzas EI, György B, Nagy G, Falus A, Gay S. Emerging role of extracellular vesicles in inflammatory diseases. Nature Reviews Rheumatology. 2014;**10**:356-364

[53] Li Y, Tan J, Miao Y, Zhang Q. MicroRNA in extracellular vesicles regulates inflammation through macrophages under hypoxia. Cell Death Discovery. 2021;**7**:285

[54] Burgelman M, Vandendriessche C, Vandenbroucke RE. Extracellular vesicles: A double-edged sword in sepsis. Pharmaceuticals (Basel). 2021;**14**(8):1-40

[55] Kronstadt SM, Pottash AE, Levy D, Wang S, Chao W, Jay SM. Therapeutic potential of extracellular vesicles for sepsis treatment. Advanced Therapeutics (Weinh). 2021;**4**(7):1-29

[56] Xia X, Yuan P, Liu Y, Wang Y, Cao W, Zheng JC. Emerging roles of extracellular vesicles in COVID-19, a double-edged sword? Immunology. 2021;**163**:416-430

[57] Sonkoly E, Pivarcsi A. microRNAs in inflammation. International Reviews of Immunology. 2009;**28**:535-561

[58] Sonkoly E, Ståhle M, Pivarcsi A. MicroRNAs and immunity: Novel players *Introductory Chapter: The Journey of Inflammation and Inflammatory Disease... DOI: http://dx.doi.org/10.5772/intechopen.101512*

in the regulation of normal immune function and inflammation. Seminars in Cancer Biology. 2008;**18**:131-140

[59] Tahamtan A, Teymoori-Rad M, Nakstad B, Salimi V. Anti-inflammatory microRNAs and their potential for inflammatory diseases treatment. Frontiers in Immunology. 2018;**9**:1-4

[60] Mehta A, Baltimore D. MicroRNAs as regulatory elements in immune system logic. Nature Reviews Immunology. 2016;**16**:279-294

[61] Zhou X, Li X, Wu M. miRNAs reshape immunity and inflammatory responses in bacterial infection. Signal Transduction and Targeted Therapy. 2018;**3**:14

[62] Yao Q, Song Z, Wang B, Zhang JA. Emerging roles of microRNAs in the metabolic control of immune cells. Cancer Letters. 2018;**433**:10-17

[63] Nelson MC, O'Connell RM. MicroRNAs: At the Interface of Metabolic Pathways and Inflammatory Responses by Macrophages. Frontiers in Immunology. 2020;**11**:1797

[64] Wang W, Yang N, Yang YH, Wen R, Liu CF, Zhang TN. Non-coding RNAs: Master regulators of inflammasomes in inflammatory diseases. Journal of Inflammation Research. 2021;**14**: 5023-5050

[65] Rose SA, Wroblewska A, Dhainaut M, Yoshida H, Shaffer JM, Bektesevic A, et al. A microRNA expression and regulatory element activity atlas of the mouse immune system. Nature Immunology. 2021;**22**:914-927

Section 2 Edema

#### **Chapter 2**

## Prolouge: Initial Approach to Edema

*Alexandro Aguilera Salgado*

#### **1. Introduction**

Edema is one of the most underrated signs that can be found in many patients. The first step is to understand what edema is in order to give this sign the importance we should. It can be caused by many different situations, so once we find it, we must study our patient completely in order to reach an adequate diagnosis and start treating our patient correctly.

Edema is the swelling from fluid accumulation at the intercellular tissue originated from the abnormal expansion of the interstitial fluid volume. Fluid at the interstitial and intravascular space is regulated by the gradient between the hydrostatic and the oncotic capillary pressures, so when this balance is altered by local or systemic situations, this fluid begins to accumulate [1].

#### **2. Patient history**

The approach to edema must begin with a complete interrogation of the patient's background. The history must include the date of the first symptoms, if edema is altered by position, if it is unilateral or bilateral, history of previous chronic diseases, substance abuse, drugs used by the patient, and any other symptoms related to the appearance of edema. With these simple questions, we can get an initial idea of the diagnosis.

The acute onset of edema of less than 72 hours is more characteristic of deep venous thrombosis, cellulitis, popliteal cyst rupture, acute compartmental syndrome, or the use of calcium channel blockers. Also, stasis can play an important role in this acute setting as in venous insufficiency, venous obstruction, or lymphatic obstruction. On the other hand, the chronic onset of edema can be seen with the appearance or as a complication of chronic diseases like chronic cardiac insufficiency, pulmonary hypertension, and thyroid, renal, or hepatic disease.

#### **3. Physical exam**

We must include a complete physical exam in every patient, in order to investigate and rule out every possible cause of edema. For example, if we are thinking the cause of edema in our patient to be cardiac insufficiency, we must look for rales or crackles, dyspnea, cyanosis, or any other sign or symptom. Our efforts should always be focused on investigating each new sign we can find, so we can further investigate them until we have the correct diagnosis.

#### **4. Diagnosis**

Once we have an idea of the possible cause of edema, we can complete our investigation with some specific studies, like complete blood count, electrolytes, hepatic enzymes, albumin, creatinine, urine analysis, glucose, and thyroid stimulating hormone [2]. Other additional and specific tests should be indicated depending on the clinical presentation, for example, if we are thinking in a cardiac etiology, we should order an electrocardiogram, echocardiogram, and chest radiograph. Another common study in certain cases when we are thinking of a lymphatic origin is a lymphoscintigraphy which can be helpful to distinguish lymphedema from venous edema and determine the cause of lymphedema. We have to keep in mind every possible situation causing edema as we can see in **Figure 1** [3].

**Figure 1.**

*Algorithm for the diagnosis of edema.*

#### **5. Treatment**

The treatment plan is set once we have an accurate diagnosis [4].

#### **6. Conclusions**

As any other signs or symptoms we can think of, edema should be thoroughly investigated. In this book, we will find a comprehensive overview of the *Prolouge: Initial Approach to Edema DOI: http://dx.doi.org/10.5772/intechopen.83666*

mechanisms and pathophysiology of edema formation and the signs and symptoms which can be seen in the different types of edema so we can reach an accurate diagnosis in order to establish the adequate treatment of this specific situation.

#### **Author details**

Alexandro Aguilera Salgado Hand and Peripheral Nerve Department, National Institute of Pediatrics, Mexico City, Mexico

\*Address all correspondence to: alexandruss@hotmail.com

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

#### **References**

[1] Cho S, Atwood JE. Peripheral edema. The American Journal of Medicine. 2002;**113**(7):580-586

[2] Yale SH, Mazza JJ. Approach to diagnosing lower extremity edema. Comprehensive Therapy. 2001;**27**(3):242-252

[3] Trayes KP, Studdiford JS, Pickle S, Tully AS. Edema: Diagnosis and management. American Family Physician. 2013 Jul 15;**88**(2):102-110

[4] O'Brien JG, Chennubhotla SA, Chennubhotla RV. Treatment of edema. American Family Physician. 2005;**71**(11):2111-2117

#### **Chapter 3**

## Peripheral Edema: Differential Diagnosis

*Sandro Michelini, Alessandro Failla, Giovanni Moneta, Alessandro Fiorentino and Cardone Marco*

#### **Abstract**

Peripheral edemas can be generated by multiple causes, local and/or systemic. The difficulties in recognizing the exact nature of the edema and the cause that originates it often lead to erroneous considerations that determine an inappropriate therapeutic approach. In this chapter the various causes that generate peripheral edema are analyzed (systemic: cardiac diastolic dysfunction, kidney failure, liver failure, myxedema, from drugs, and idiopathic; and local: venous and/or lymphatic transport insufficiency). They are also described, according to the diagnosis made and the clinical and instrumental criteria to attain a correct and early diagnosis and to proceed to the most appropriate therapeutic measures (drugs, surgery, physical rehabilitative by means of manual and mechanical techniques) in individual cases.

**Keywords:** peripheral edema diagnosis, edema diagnosis, peripheral edema of limbs differential diagnosis

#### **1. Introduction**

Lower limb edema recognizes more etiological factors that are frequently confused during differential diagnosis. Sometimes there are more causes with preponderance of one over the other, either local or systemic.

The doubts in the diagnostic definition derive from an insufficient evaluation of clinical symptomatological aspects and of any instrumental and hemato-chemical tests performed in individual cases.

From a correct clinical and consequently ethiopathogenetic classification derives the most appropriate therapeutic option. Pharmacological, physical rehabilitative, or surgical therapies not inspired by edema correction principles based on its ethiopathogenesis may result in therapeutic failure or even in the worsening of local or systemic clinical status.

#### **1.1 Description**

The causes of edema of the lower limbs are various (local and/or systemic), sometimes multiple, and are to be found on the basis of a series of anamnestic and semeiological elements that, if properly considered and identified, allow better management of the clinical picture [1].

Too often, in fact, even today we are witnessing the diagnosis of lymphostatic edemas of lower limbs, ignoring that in many cases the loco-regional lymphatic system is normally developed and adequately functional.

#### *Inflammation in the 21st Century*

The same monolaterality of edema, by itself alone, allows to address the diagnostic suspicions towards a local and non-systemic cause. A systemic cause of edema of lower limbs, in fact, always determines bilateral edema (albeit with relative prevalence in one of the two limbs), never being unilateral [1].

Therefore, on the one hand, it is necessary to have a clear presence of the systemic causes of edema and of the loco-regional ones and, on the other hand, the clinical and instrumental criteria which, together, allow to formulate the correct diagnosis and to prepare the most indicated therapeutic measures.

#### **2. Causes of edema of the lower limbs**

#### **2.1 Local**


#### **2.2 Systemic**


Among the **loco-regional** causes, the most important and frequent is represented by lymphedema (primary and secondary).

#### **2.3 Lymphedema**

Lymphedema derives from an altered (qualitative or quantitative) development of the loco-regional system. In primary forms (which may occur at birth or, more frequently, in the second, third, fourth, fifth decade of life), an altered development

#### *Peripheral Edema: Differential Diagnosis DOI: http://dx.doi.org/10.5772/intechopen.82400*

of the lymphatic pathways, an altered lymph-node architectural code (lymphadenodysplasia), or an insufficient number of them (often on a genetic basis in the development of lymph glandular stations) causes a slowing of the lymphatic return that can go as far as the stop of the flow at the loco-regional level. In some cases the familiarity for the affection is documented, in others (the so-called sporadic forms, because we do not know their existence in other family members), the lymphedema appears in the only subject clinically affected without affecting other members of the same family nucleus; then there is a third type of primary lymphedema in which the edema constitutes only one (and not always the "determinant") of the various clinical aspects of a syndrome (Prader-Willi, Noonan, Proteus, Hennekam, Gorham-Stout).

The secondary forms are sometimes considered primary for the predisposition in some subjects to develop secondary edema following certain clinical conditions (one example is the "post-mastectomy lymphedema", which develops in one in four woman, while the others remain with the same limb, in volume and consistency, throughout their lives, even if they are of the same age and in the same physical condition, and undergo the same surgery by the same operator); the genesis and the ethiopathogenetic evolution, in these cases, are the same as in the primary forms; in these cases, as a result of inflammatory processes, traumas, or, more frequently, surgical lymphadenectomy for neoplasms or radiotherapy, the anatomical continuity of the local lymphatic circulation is lost in an acquired manner so that the clinical picture of the lymphostasis is observed. One of the clinical peculiarities of lymphedema of the lower limbs is the different progression of stasis along the limb: from the more distal portions towards the proximal ones in the primary forms and from the proximal ones to the distal ones in the secondary forms (**Figure 1a** and **b**).

Lymphedema is characterized by being the only edema with high interstitial protein concentration, distinguishing itself for this from all other types of edema [16, 17]. The presence of a high rate of proteins in the interstitium determines the activation of fibroblasts that increase their production of collagen fibers, inducing more or less early and more or less marked tissue sclerosis. Lymphatic edema, therefore, is characterized by an early increase in the consistency of the tissues in which it is located and that, in the most advanced clinical stages, can reach the wooden consistency. Under these conditions the compression of the skin generates a depression (or "pitting test") that can be "fleeting" or even absent [18–23].

**Figure 1.** *Lymphedema of the lower limb: primary (a) and secondary (b).*

#### *Inflammation in the 21st Century*

For the rarefaction of the arteriolar capillaries which, with the same volumetric unit, is carried out in the cutaneous and subcutaneous tissues, the skin color is not "rosy" as in the skin of normal limbs but pale. For the same ethiopathogenetic reason, the limb skin with lymphedema is colder than of a normal limb [24–28].

The lymphatic system presents some well-recognizable outward signs. The lymphatic system communicates us, but often the examiner does not recognize the messages sent. An example of this is the location of the primary or metastatic cancer in the lymphatic system itself. Sometimes a monolateral edema of the upper limb, sent for decongestive complex therapy with the prescription of a manual lymphatic drainage cycle for forearm edema and a recent onset, may be the expression of a symptomatic edema of a metastatic cancer (**Figure 2a** and **b**). In these cases, an aprioristic therapeutic approach, without the most opportune clinical considerations, can fatally delay the care that the patient really needs.

The commonly recognized four clinical stages of the disease are:


In primary forms of lower limbs, there is also a pathognomonic sign that takes its name from the person who first described it: the Stemmer sign. Its positivity consists in the impossibility to "pinch" with the fingers of the examiner the skin of the patient's toe; you cannot lift it from the underlying bone phalanx due to the

**Figure 2.**

*Forearm and hand edema, determined by the presence of metastases of the supraclavicular lymph nodes from unknown primitive pulmonary cancer (first clinical manifestation of neoplasia).*

#### *Peripheral Edema: Differential Diagnosis DOI: http://dx.doi.org/10.5772/intechopen.82400*

early fibrosis that is generated in the over-factory layers of the tissue itself. The diagnosis of lymphedema is clinical (fundamental is the anamnesis and a correct objective examination); however, there are some instrumental exams that complete the picture, allowing a better definition of the therapeutic approach and the prognostic one. In the primary forms, the lymphoscintigraphy is essential which consists of subcutaneous inoculation, at the root of the toes, of some drops of radioactive tracer (nanocolloids of albumin labeled with technetium-99) which has a particular tropism for the lymphatic system. After inoculation, the patient practices physical exercise to allow the tracer to "gain" more quickly the lymphatic pathways. After 30′, and after 90′, a gamma-chamber performs that uptake of the tracer which, in the meantime, is distributed in the lymphatic vessels and lymph nodes of the whole lower limb and in the iliac chains. The resulting image provides important morphological indications on the normal/pathological development of the lymphatic system allowing better orienting the therapeutic intervention and being able to conceive also a prognosis.

The high-resolution ultrasound examination also highlights the thickening of the epidermis on the affected side, the increase in thickness of the over-fascial layer, and the tissue compressibility that is a function of the more or less developed fibrosis.

The ultrasound also allows the monitoring of pharmacological, physical rehabilitative, and surgical treatment by comparing the pre- and posttreatment overfascial thicknesses.

Videofluoroscopy is more complex and difficult to access because it is not widely practiced at a territorial level. It consists of the study of the anatomy physio-lymphatic pathology by injection of a dye, the indocyanine green, which flows into the lymphatic vessels and allows to visualize the flow in real time (on videoscope) both basal and during manual or mechanical stimulation; the lymphangio-MR, even less widespread as practiced in very few centers at the international level, allows, in more detail, to highlight the entire local lymphatic system and its possible anatomical defects.

#### **2.4 Venous edema**

It is very rare, in contrast to the lymphatic stasis edema, to observe a venous edema deriving from the supra-fascial venous compartment. In this sense the lymphatic system and the venous system behave in a completely opposite way from the clinical point of view. Lymphedema never develops in deep tissues, unless congenital dysplasia is located in the deep tissues; it is always located at the suprafascial level.

Venous edema, on the other hand, never develops in the supra-fascial compartment (in the clinical practice, it is common to find patients who have large varicose veins of the lower limbs, but their feet are "dry", with no signs of edema.). Venous edema is an edema that is located at a deep level (for this reason, it is difficult to manage, from the therapeutic point of view, with conventional manual or mechanical drainage techniques). It is located at the sub-fascial level and, in the overwhelming majority of cases, represents the most striking aspect of the permanent clinical picture of a so-called "post-thrombotic syndrome" (**Figure 3**).

In deep venous thrombosis of the lower limbs, in fact, after thrombotic occlusion of the deep veins, follows more or less precociously a "compensatory" dilatation of the deep collateral and often superficial circle in correspondence of the same anatomical district (secondary or symptomatic varices). This muscular imbibition (therefore deep) assumes the characters of chronicity and corresponds to the permanent edema in which it is not possible to observe alterations of skin color or

**Figure 3.** *Post-thrombotic oedema of the lower limb.*

skin temperature in its correspondence, and the pitting sign is absent and does not appreciate changes in tissue texture. In doubtful cases (previous venous thrombosis passed "unobserved" from a clinical point of view due to lack of characteristic signs and symptoms), a key examination is represented by the computerized tomography.

According to the "CT cuts" of the two limbs in comparison with the lymphatic edema it is possible to observe an increase in supra-fascial thickness, over-folded with the sub-fascial compartment coinciding in the two limbs, in venous edema (from deep vein thrombosis); on the contrary, the supra-fascial thicknesses appears coincident, while the sub-fascial thickness is considerably increased in the affected side.

Symptoms in the post-thrombotic syndrome of the lower limb (in the one-sided form) are generally non specific; patients often show paresthesia, rarely pain, mostly vague, hardly epicritic, and often associated with protopathic sensitivity.

The so-called venous claudication which consists in pain during walking but with a variable free-range of motion (unlike "arterial claudication"), is rare and appears in the worsening of the deep venous circulation due to incomplete recanalization of the deep venous axis when the acute phase is past.

Obviously, the high-resolution ultrasound examination in these cases shows a relative increase of the sub-fascial layers, and the echo color Doppler of the examined districts confirms the outcomes of deep vein thrombosis with frequent evidence of parietal sclerosis and partial or total disappeared endoluminal valvular structures.

#### **2.5 Phlebolymphedema**

Phlebolymphedema represents a particular type of peripheral edema that is determined by the contemporary ethiopathogenetic association of venous and

#### *Peripheral Edema: Differential Diagnosis DOI: http://dx.doi.org/10.5772/intechopen.82400*

lymphatic insufficiency. It is generally present in cancer, due to the simultaneous macroscopic and microscopic anatomical involvement of the two venous and lymphatic drainage systems of a certain anatomical district8 .

The most striking manifestation is a secondary lymphedema of the lower limb determined by inguinal or pelvic lymphadenectomy (necessary for compliance with surgical criteria of "radical cancer therapy") associated with a partial or complete occlusion of a deep venous vessel of the limb same. In these cases the increase in interstitial oncotic pressure determined by the mechanical lymphatic stasis (removal of lymphoglandular stations) is associated with a venous hypertension (increase of the intravascular hydrostatic pressure in the venous side of the microvascular tissue unit) caused by the occlusion of the main venous axis of outflow from the lower limb itself (**Figure 4**).

#### **2.6 Lipedema**

Lipedema is a very common disease in the female population consisting predominantly of lipid cells that is established in certain specific anatomical districts. It has a familiar character with the male that results (as found in several pedigrees) in "healthy carrier." In the lower limbs, the localization can be variable but always bilateral; it can be limited to the thighs, but it can also affect the gluteal regions, affecting only the legs, affecting the thighs and legs, or involving, in addition to these, the buttocks (**Figure 5**). The feet are always spared. Sometimes the arms and forearms are also affected, and hands are always spared. The edema is associated with constant pain, which becomes acute with the passing of the hours of the day and during the summer, and ease of spontaneous bruising [10–12, 29].

Edema, generally, appears at puberty and is exacerbated at some particular moments in a woman's life (breastfeeding more than pregnancy and menopause). The edema appears simultaneously in all the regions of the affected limbs, and there is never a progression along the limbs (neither in the distal-proximal or proximal-distal sense) but only a possible simultaneous increase of the edematous zones. It is an edema that does not respond to hypocaloric dietary treatments or physical exercise.

#### **Figure 4.**

*(a and b) Phlebolymphedema: a case of prostate cancer with pelvic lymphadenectomy associated with venous left iliac thrombosis. (a) Clinical case and (b) phlebography of the lower limb.*

**Figure 5.** *Lipedema of the lower limbs.*

We recognize four clinical stages of the disease:


The lymphoscintigraphy of lower limbs, in the early stages, shows a normal development and draining lymphatic circulation. In the advanced clinical stages of the disease, it is possible to underline bilateral sub-rotuleous stagnation of the tracer (dermal back flow) and lymph node stop that corresponds, from the clinical point of view, to the so-called lipolymphedema of the lower limbs.

High-resolution ultrasound allows to highlight a constant echogenic pattern of the supra-fascial compartment (skin-fascia). It is an extremely useful test for the differential diagnosis with lymphedema of the lower limbs. In lipedema, in fact, the compression of tissues with a linear probe shows a reduction in the sub-fascial thicknesses with the supra-fascial which remained unchanged; in the case of lymphedema, on the contrary, the compression with linear probe shows a decrease

#### *Peripheral Edema: Differential Diagnosis DOI: http://dx.doi.org/10.5772/intechopen.82400*

in the over-fascial thicknesses, while the sub-fascial remain unchanged. This testifies that in lipedema the volumetric increase is given by an increased cellular component (hypertrophic and hyperplastic lipid cells) while in the lymphedema the volumetric increase is given by more or less copious presence of extracellular interstitial fluids that the pressure of the probe can move.

The BMI is variable in the two pathologies. In lipedema it is generally within the limits of the norm and is not minimally influenced by physical treatments nor by the overall weight loss. In lymphedema it can be equally variable even if it is higher on average than in patients with lipedema.

The differential diagnosis with obesity is quite simple. In the obese patients, the collection of fatty deposits is widespread in all the body regions with a particular preference for the anatomical areas typical of each sex (gynoid type and android type).

Obesity responds positively to physical exercises and diet, in all body districts, while lipedema is not affected at all by these factors.

#### **2.7 Inflammatory or infectious states**

The edema of the lower limb can be determined by inflammatory and/or infectious diseases. In these cases the localization is generally monolateral and is secondary to an inflammatory/infectious process of the soft tissues (cellulitis, myositis, myofascitis, necrotizing fasciitis). The localization may involve only one area of the limb (thigh, leg, foot) assuming the topographic configuration of the "suspended edema." Besides the edema, all the other characters of inflammation are generally present (increase in skin temperature, hyperemia, pain, and reduction of functional capacity (*functio laesa*). The resolution of edema is due, in these cases, to anti-inflammatory and pharmacological treatment and, if necessary, to antibiotics. A special case of edema that can induce doubts of differential diagnosis with deep vein thrombosis and that is determined by the presence of serous cyst of the popliteal cable is known as Baker's cyst. Particularly developed, it can compress the surrounding venous and lymphatic vessels, inducing a distal edema (generally subpatellar) also extended to the whole leg and to the foot. With the treatment of cystic formation (puncture with evacuation, surgical excision, simple anti-inflammatory therapy), we are witnessing the resolution of the edema.

#### **2.8 Benign or malignant neoplasms**

Benign or malignant tumoral formations may develop in the lower limbs, especially the soft tissues (mainly muscles). Their localization, particularly in the leg muscles, occupying space, also due to the compressive phenomena exerted on the surrounding vessels, can determine circumscribed or diffused edema which, for the differential diagnosis, must make use of more discriminating investigations, such as CT or MR. Obviously, the edema, in these cases, recedes only after surgical removal of the mass.

#### **2.9 Systemic causes**

#### *2.9.1 Edema in heart failure (from diastolic dysfunction)*

In the case of heart failure, the echocardiographic examination may be apparently normal (the ventricular ejection fraction, in these cases, provides normal indications, and no particular problems are highlighted). In these conditions, however, a careful clinical examination is required which highlights a bilateral and symmetrical lower limb edema (often confused with lymphedema); the sign of the fovea is particularly

evocable and persistent over time. The Stemmer sign is negative (**Figure 6**). The edema is established by an important increase in venous pressure in the microvascular tissue units, whereby the normal pressure gradient at the micro-tissutal level which, from the hydrostatic point of view, under normal conditions, would help the return of fluids from the interstitium towards the venous capillaries is gone and many water molecules remain in the interstitium. The thoracic auscultation demonstrates the presence of bilateral basal crackles. The liver may appear increased in volume and with rounded margins13. The subject refers to dyspnea for slight efforts or even at rest. During nocturnal rest the patient needs to observe a decubitus which raises the thorax and head with respect to the other bodily districts. In these cases, the dosage of the inactive form of the Brain Natriuretic Peptide (BNP, substance produced by the cardiac endothelium, whose serum concentration increases in the case of inability of filling by the cardiac chambers) results elevated, suggesting that the heart has difficulty in receiving fluids returning from the periphery (second- or third-degree diastolic dysfunction); this disfunctions are relatively frequent in women above 65 years. In these cases conventional draining physical therapies risk aggravating subjective and objective symptoms, and the same elastic garment to be worn in the morning should be prescribed only after the clinical compensation obtained with the appropriate dosage of diuretics and positive inotropic drugs (e.g., digoxin) indicated for each individual case.

#### *2.9.2 Hepatic insufficiency*

The edema of hepatic insufficiency can be defined as "gravitational"; it is collected mainly in the sloping body areas. It is essentially localized in the toes and feet when the subject is standing or sitting with the legs "dangling." When the subject is lying on the bed, the most gravitational area is the presacral region14. It is not uncommon in these cases, if the patient holds a higher limb outside the bed, observe the edema at the level of the elbow of the arm itself. The conventional physical therapies, even in this case, do not solve the problem that benefits only from the administration of intravenous albumin. It is hypo-albumin, which in fact generates edema: since each protein molecule behaves like a kind of magnet to water molecules (it attracts them); in the case of hypo-albuminemia, liquids are no longer held within the intra-vascular compartment and tend to flee to intertial space, following the gravity.

**Figure 6.**

*Stemmer sign: positive in lymphedema (a), negative in cardiac failure (b) before compression and (c) after compression.*

#### *Peripheral Edema: Differential Diagnosis DOI: http://dx.doi.org/10.5772/intechopen.82400*

#### *2.9.3 Acute and chronic renal failure*

In chronic renal failure at third or fourth stage, the relative inability of the nephron to produce "pre-urine" inevitably leads to a generalized "water retention" affecting all body districts. The subject, frequently, in the morning wakes up with the edematous eyelids and only an adequate dosage of diuretics, respecting the values of the electrolytes and, above all, of the renal function (azotemia, creatinine). Decongestive physical treatments cannot find an elective indication even in this form.

#### *2.9.4 Myxedema*

Myxedema is a particular form of edema affecting the lower limbs (generally bilaterally, symmetrically, and localized to the pretibial surfaces) determined by accumulation of mucopolysaccharides in the derma. In these cases, the pretibial edema is also accompanied by ocular edema (exophthalmos), with generalized dryness of the skin and, sometimes, psychic hypo-evolution [15]. It is determined by serious conditions of hypothyroidism (congenital or acquired), in which TSH (which, for various reasons, does not respond to the thyroid parenchyma) would stimulate tissue fibroblasts and adipocytes to replicate and produce mucopolysaccharide complexes, with local deposition inside the dermis, or, on the contrary, in cases of hyperthyroidism (as in Flaiani-Basedow's disease). This type of edema is reversible and recognizes as a fundamental therapeutic treatment the correction of thyroid defect (in defect or excess of glandular function). It is presented as a "suspended edema," not painful, and without local typical signs of inflammatory processes.

#### *2.9.5 Drug edema*

Drug edema occurs, in most cases, by a particular idiosyncrasy of the subject to certain molecules. The drugs most commonly called into question in these forms are some molecules with antihypertensive effect (especially amlodipine and other calcium antagonists, in which the edema is localized mainly in the two ankles bilateral edema—and in the back of the feet, and the sartanics that can induce generalized edemas, up to anasarca) and the corticosteroids which cause a generalized water retention. In all these forms, physical treatment is not conclusive, and the therapy consists in the simple suspension of the drug (with substitution with different molecules). In almost all the cases, the complete resolution of the clinical picture is achieved within a maximum of 7 days from the suspension of the drug.

#### *2.9.6 Idiopathic edema*

Many authors are reminded of the possible presence of "idiopathic edema," or an edema (generally distal and bilateral) that arises at certain times of the day (especially after prolonged standing) or in the summer season. It regresses with the wearing of the definitive elastic garment for a variable period of time.

#### *2.9.7 Differential diagnosis of edema of the lower limbs*

The differential diagnosis of edema of the lower limbs can be easily formulated through simple observations concerning skin color, skin temperature, mono- or bilaterality localization, the presence of the sign of pitting, the presence of the Stemmer sign, the sense of progression of the edema along the limb, and the date

#### *Inflammation in the 21st Century*

of onset of edema, compared to the time of observation. From a combined analysis of these elements, it is possible to easily reach the diagnosis that can be further confirmed by other exams, already described in the individual-treated paragraphs [4, 12, 30–36].

In particular, in relation to:


itself. In lipedema, in the early clinical stages, the pitting sign is absent (the volumetric increase is determined by the exclusive presence of hyperplastic and hypertrophic adipose cells and not to fluids in the interstitium). It can appear in the most advanced clinical stages (lipolymphedema) [39].


#### **3. Conclusions**

The opinion that an edema of the lower limbs, regardless of the patient's age, of the general clinical conditions and symptoms and signs that accompany the picture, is of a lymphostatic nature is still widespread today. So it happens that many cardiologists send to the angiologist or to the vascular surgeon patients over 70 years, with a fairly delineated symptom complex, albeit unidentified, with the diagnosis of "recent-onset limb lymphedema"; clinical cases that, if properly considered, are of strict cardiological relevance and not of physical rehabilitative medicine. Just as Lipedema is still unknown, as a pathology, by over 50% of the same vascular surgeons and of the angiology and by the overwhelming majority of family doctors [46].

The hemato-chemical and instrumental examinations are undoubtedly useful for a better definition of individual cases, both for the purposes of the therapeutic approach and the prognosis and monitoring.

However, the diagnosis must be essentially clinical and is based on the considerations described, simply by analyzing the individual objective and subjective parameters, between them, and crossing the information. Clinical experience can accelerate the diagnosis and the accuracy of the subsequent therapeutic approach, but it is fundamental, in any case, that before a definitive diagnosis, we consider the "semeiological picture" which, combined with an accurate clinical history, allows to reach the certainty of differential diagnosis.

Even today there are diagnostic mistakes in evaluation of many edema of the lower limbs. The lack of specific clinical experience and the underestimation of important anamnestic elements, supported by clinical evidence that often are not sought in the various details or, the misinterpretation of instrumental investigations can lead to inaccurate ethiopathogenetic diagnoses with negative consequences from the point of view of treatment that is undertaken in the individual clinical case.

The proposed analysis aims at avoiding reckless or "discounted" clinical judgments, but not responding to the real needs of the individual patient, and helping the medical doctor, the physiotherapist, and the nurse to follow the most appropriate diagnostic and therapeutic procedures in line with the current principle prevention, early diagnosis, and treatment.

#### **Conflict of interest**

The authors declare no conflict of interest.

#### **Author details**

Sandro Michelini\*, Alessandro Failla, Giovanni Moneta, Alessandro Fiorentino and Cardone Marco Department of Rehabilitation, S. Giovanni Battista Hospital, Rome, Italy

\*Address all correspondence to: s.michelini@acismom.it

© 2020 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

*Peripheral Edema: Differential Diagnosis DOI: http://dx.doi.org/10.5772/intechopen.82400*

#### **References**

[1] Tiwari A, Cheng KS, Button M, Myint F, Hamilton G. Differential diagnosis, investigation, and current treatment of lower limb lymphedema. Archives of Surgery. 2003;**138**(2):152-161

[2] Michelini S. Phlebolymphoedema. From Diagnosis to Therapy. Bologna: Edizioni P.R.; 1998

[3] Badini A, Fulcheri E, Campisi C, Boccardo F. A new approach in histopathological diagnosis of lymphedema: Pathophysiological and therapeutic implications. Lymphology. 1996;**29**(S):190-198

[4] Gasbarro V, Michelini S, Antignani PL, Tsolaki E, Ricci M, Allegra C. The CEAP-L classification for lymphedemas of the limbs: The Italian experience. International Angiology. 2009;**28**(4):315-324

[5] Lee B, Andrade M, Bergan J, Boccardo F, Campisi C, Damstra R, Flour M, Gloviczki P, Laredo J, Piller N, Michelini S, Mortimer P, Villavicencio JL. Diagnosis and treatment of primary lymphedema. Consensus Document of the International Union of Phlebology (IUP)-2009. International Angiology. 2010;29(5):454-470

[6] International Lymph Framework. Best Practice for the Management of Lymphoedema. 2nd ed. 2012. Available at: www.lympho.org

[7] The diagnosis and treatment of peripheral lymphedema: 2013 consensus document of the international society of lymphology. Lymphology;**46**(2013):1-11

[8] Michelini S, Cardone M. Venolymphatic vascular malformations: Medical therapy. In: Mattassi R, Loose DA, Vaghi M, editors. Hemangiomas and Vascular

Malformations. Italia: Springer; 2015. pp. 445-450

[9] Michelini S, Pissas A, Olszewski W, Dimakakos E, Cordero IF, Caldirola R, et al. Linforoll: A new device for lymphoedema treatment: Preliminary experience. Lymphology. 2015;**47**(Suppl):218-221

[10] Mariani G, Campisi C, Taddei G, Boccardo F, Martini F, Rahimi Mansour A, et al. The current role of lymphoscintigraphy in the diagnostic evaluation of patients with peripheral lymphedema. Lymphology. 1998;**31**(Suppl):316-319

[11] Cavezzi A, Michelini S. PHlebolymphoedema. Bologna: Edizioni P.R; 1998

[12] Nicolaides AN. Therapeutic outcome and quality of life in patients with chronic venous and lymphatic disorders. Phlebolymphology. 2008;**20**:2-3

[13] Schmeller W, Hueppe M, Meier-Vollrath I. Tumescent liposuction in lipoedema yields good long-term results. The British Journal of Dermatology. 2012;**166**:161-168

[14] Schmeller W, Meier-Vollrath. Lipödem: Ein update (Lipedema: an update). Lymphol Forsch Prax. 2005;**9**(1):10-20

[15] Forner-Cordero I, Szolnoky G, Forner-Cordero A, Kemény L. Lipedema: An overview of its clinical manifestations, diagnosis and treatment of the disproportional fatty deposition syndrome—Systematic review. Clinical Obesity. 2012;**2**:86-95

[16] Pascual-Figal DA, Domingo M, Casas T, Gich I, Ordoñez-Llanos J, Martínez P, et al. Usefulness of clinical and NT-proBNP monitoring for

prognostic guidance in destabilized heart failure outpatients. European Heart Journal. 2008;**29**(8):1011-1018

[17] Younossi ZM, Guyatt G, Kiwi M, Boparai N, King D. Development of a disease specific questionnaire to measure health related quality of life in patients with chronic liver disease. Gut. 1999;**45**:295-300

[18] Schwartz KM, Fatourechi V, Ahmed DDF, Pond GR. Dermopathy of Graves' disease (pretibial myxedema): Long-term outcome. The Journal of Clinical Endocrinology & Metabolism. 2002;**87**(2):438-446

[19] Bellini C, Arioni C, Mazzella M, Campisi C, Taddei G, Boccardo F, et al. Lymphoscintigraphic evaluation of congenital lymphedema of the newborn. Clinical Nuclear Medicine. 2002;**27**(5):383-384

[20] Boccardo F, Michelini S, Zilli A, Campisi C. Epidemiology of lymphedema. Phlebolymphology. 1999;**26**:24-28

[21] Werngren-Elgstrom M, Lidman D. Lymphoedema of the lower extremities after surgery and radiotherapy for cancer of the cervix. Scandinavian Journal of Plastic and Reconstructive Surgery and Hand Surgery. 1994;**28**(4):289-293

[22] Campisi C, Michelini S, Boccardo F, Zilli A. Lymphedema epidemiology in Italy. Lymphology. 1998;**31**(Suppl):243-244

[23] Casley-Smith J. Modern Treatment for Lymphoedema. Adelaide: The Lymphoedema Association of Australia, Inc.; 1994

[24] Dellachà A, Fulcheri E, Boccardo F, Campisi C. Post-surgical lymphedema: Iatrogenic or pre-existing disease? Lymphology. 1998;**31**:562-565

[25] Földi E, Földi M. Physiothérapie Complete Décongestive. Paris: Editions Frison-Roche; 1993

[26] Leduc A. Le drainage lymphatique. Paris, Masson: Théorie et pratique; 1980

[27] Michelini S, Failla A, Moneta G, Campisi C, Boccardo F. Clinical staging of lymphedema and therapeutical implications. Lymphology. 2002;**35**:168-176

[28] Michelini S, Failla A. Linfedemi: Inquadramento diagnostico clinico e strumentale. Minerva Cardioangiologica. 1997;**45**(Suppl I): 11-15

[29] Tosatti E. Lymphatique profonds et lymphoedèmes chroniques des membres. Paris: Masson; 1974

[30] Vodder E. La méthode Vodder— Le drainage lymphatique manuel. DK-2880, Bagsvaer: Inst. For Lymph Drainage; 1969

[31] Olszewski W. Recurrent bacterial dermatolymphangioadenitis (DLA) is responsible for progression of lymphoedema. Lymphology. 1996;**29**(Suppl):331

[32] Michelini S, Campisi C, Failla A, Boccardo F. Proposal for stadiation of phlebolymphoedema. European Journal of Lymphology and Related Problems. 1995;**6**(20):I-14

[33] Campisi C. Lymphoedema: Modern diagnostic and therapeutic aspects. International Angiology. 1999;**18**(1):14-24

[34] Case TC, Witte CL, Witte MH, Unger EC, Williams WH. Magnetic resonance imaging in human lymphedema: Comparison with Lymphangioscintigraphy. Journal of Magnetic Resonance Imaging. 1992;**10**:549-558

[35] Ferrel RE, Levinson KL, Esman JH, Komak MA, Lawrence EC, Barmada MM, et al. Hereditary lymphedema evidence for linkage and

#### *Peripheral Edema: Differential Diagnosis DOI: http://dx.doi.org/10.5772/intechopen.82400*

genetic heterogeneity. Human Molecular Genetics. 1998 Dec. 7;**13**:2073-2078

[36] Michelini S, Failla A, Moneta G. Lymphedema: Epidemiology, disability and social costs. Lymphology. 2002;**35**:169-171

[37] Michelini S, Failla A, Moneta G, Zinicola V, Romaldini PD. International classification of lymphedema functioning and disability evaluation. European Journal of Lymphology. 2007;**17**(51):16-19

[38] Michelini S, De Giorgio D, Cestari M, Corda D, Ricci M, Cardone M, et al. Clinical and genetic study of 46 Italian patients with primary lymphoedema. Lymphology. 2012;**45**:3-12

[39] Michelini S, Failla A, Moneta G, Cardone M, Michelotti L, Zinicola V, et al. Linee guida e protocolli diagnostico-terapeutici nel linfedema. Eur. Med. Phys. 2008;**44**(Suppl. 1-3)

[40] Michelini S, Failla A, Moneta G, Zinicola V, Macaluso B, Cardone M, et al. Treatment of lymphedema with shockwave therapy: Preliminary study. The European Journal of Lymphology and Related Problems. 2007;**17**(51):29

[41] Michelini S, Failla A, Moneta G, Cardone M, Fiorentino A. Immunestimulation and reduction of infective complications in patients with lymphoedema. European Journal of Lymphology and Related Problems. 2009;**20**(56):17-18

[42] Partsch H. Indirect lymphography in different kinds of leg oedema. In: Lymphology: Advances in Europe. Ecig: Genova; 1989. pp. 95-99

[43] Pecking AP, Cluzan RV. Assessment of lymphatic function: 15 years experience using radionuclide methods. Lymphology. 1994;**27**(Suppl):301-304

[44] Schingale FJ. Lipoedema. In: Schingale FJ, editor. Lymphoedema, Lipoedema: A Guide for those Effected. Hannover: Schlǖtersche; 2003. pp. 64-71

[45] Trévidic P, Marzelle J, Cormier JM. Apport de la microchirurgie au traitement des lymphoedèmes. In: Editions Techniques -Encycl. Méd. Chir. Paris, France: Techniques chirurgicales-Chirurgie vasculaire; 1994 F.a. 43-225

[46] Földi M. The therapy of lymphedema. European Society of Lymphology. 1993-1994;**14**:43-49

#### **Chapter 4**

## Edema Induced by sPLA2 from *Crotalus durissus terrificus* Involves PLC and PKC Signaling, Activation of cPLA2, and Oxidative Stress

*Marcos H. Toyama, Caroline R.C. Costa, Mariana N. Belchor, Danielle P. Novaes, Marcos A. de Oliveira, Rolando Ie, Henrique Hessel Gaeta and Daniela de O. Toyama*

#### **Abstract**

sPLA2 from *Crotalus durissus terrificus* venom, free of crotapotin (Cdt sPLA2), purified and isolated sPLA2, was able to significantly increase lipid peroxidation, which occurred simultaneously with increased arachidonic acid (AA) metabolism. In addition, MDA and AA levels were elevated at 15 min after Cdt sPLA2 injection and after peak edema (negative control). Thus, oxidative stress and ROS play important roles in the inflammation induced by Cdt sPLA2. On the other hand, edema induced by sPLA2 involves the direct and indirect mobilization of arachidonic acid by the involvement of phosphokinase C (PKC) and phospholipase C (PLC), which indirectly stimulates cytosolic PLA2 (cPLA2). We also observed that the specific antivenin against Cdt venom had no significant effect on the neutralization of induced edema compared to the natural products 5-caffeine-linoleic acid (5CQA) and dexamethasone (AACOCF3). Our results also indicate that there was improvement in the inhibition of edema of natural polyphenolic compounds compared to antivenin or inhibition of the enzymatic activity of sPLA2 due to the fact that 5CQA is a potent antioxidant compound. Thus, our results show a clear correlation between increased arachidonic acid metabolism and oxidative stress.

**Keywords:** *Crotalus durissus terrificus* (Cdt), secretory snake venom phospholipase A2, edema, PKC, PLC, inflammation, oxidative stress

#### **1. Introduction**

#### **1.1 Arachidonic acid "dogma"**

Arachidonic acid (ARA) is a 20-carbon chain fatty acid with four methyleneinterrupted *cis* double bonds; the first, with respect to the methyl end (omega, ω or n), is located between carbons 6 and 7. Arachidonic acid (AA) has three possible

#### *Inflammation in the 21st Century*

destinations: participating in the remodeling process of the cell membrane, release into the extracellular medium by diffusion, or its intracellular metabolism [1, 2]. In addition to AA, lysophosphatidic acid (lyso-platelet aggregation factor (PAF)) is another product of the enzymatic hydrolysis of membrane phospholipids, which, in the presence of lyso-PAF acyl transferase, is converted in PAF [3]. PAF is an extracellular lipid signaling molecule involved in a range of cellular activities, including survival, differentiation, cellular proliferation, morphological changes, and migration, among others [4]. Besides, its biological action is mediated by the presence of a cellular receptor (PAF-receptor (PAF-R)) (**Figure 1**). These physiological and pharmacological activities of PAF depend on the presence of its receptors, designated as PAF-R1 to PAF-R6. These receptors are G protein-coupled transmembrane receptors, and recent studies revealed that the PAF-R signaling pathway clearly affects different aspects of tumor progression [5, 6]. In the literature, it is well established that phospholipases A2 (PLA2s) are key enzymes involved in AA generation by hydrolytic digestion of membrane phospholipids. PLA2 is a superfamily

#### **Figure 1.**

*Central dogma of arachidonic acid metabolism. AA cascade and its destination following three major oxidative pathways: (1) cyclooxygenase (COX), producing prostaglandins and related eicosanoids; (2) lipoxygenase (LOX), forming leukotrienes and related compounds; and (3) CYP450, forming arachidonic acid epoxides.*

*Edema Induced by sPLA2 from* Crotalus durissus terrificus *Involves PLC and PKC Signaling… DOI: http://dx.doi.org/10.5772/intechopen.80848*

of enzymes distributed throughout six major classes: secretory PLA2 (sPLA2), calcium-dependent cytosolic PLA2 (cPLA2), calcium-independent cytosolic PLA2 (iPLA2), lysosomal PLA2 (lPLA2), mitochondrial PLA2 (mPLA2), and, more recently, PAF-acetyl hydrolases (PAF-AHs). PAF-AHs are a small family of phospholipases A2 with a high specificity for the hydrolysis of the unsaturated fatty acid residue located at the sn-2 position [7, 8]. sPLA2 is considered a simple and primitive enzyme, acting as an inducer of the inflammatory process, besides being able to act as a pseudohormone. In addition to generating AA directly, this enzyme can also increase the activity of cPLA2 [9]. Furthermore, the produced AA usually follows one of three distinct enzymatic pathways involving cyclooxygenase, lipoxygenase, and cytochrome P450. Several products of these routes can modulate the functions of ion channels, protein kinases, and ion pumps. In addition, newly formed eicosanoids are excreted and mediate various physiological functions, including insulin secretion and muscle contraction, and most of these actions involve protein G. Ultimately, the products of AA metabolism are rapidly degraded [1, 10]. Briefly, AA, as well as other polyunsaturated fatty acids (PUFAs) generated at the cellular level, can be mobilized through the hydrolytic activity of various enzymes. It is possible to highlight the action of PLA2 through a single reaction pathway that produces AA and lysophospholipid (LysP), which is considered the classical pathway of AA generation—it is the most widely known and studied. In addition, AA is metabolized by cyclooxygenase (COX) and 5-lipoxygenase, resulting in the synthesis of prostaglandins and leukotrienes, respectively. These intracellular messengers play an important role in the regulation of signal transduction, leading to pain and inflammatory responses. Recently, the literature has shown that AA can follow a third pathway, resulting in its metabolism by cytochrome P450 enzymes—Cyt450 epoxygenase and Cyt450 omega hydroxylase. P450s are typical monooxygenases, which enzymatically cleave molecular oxygen, followed by the insertion of a single atom of oxygen into the substrate, while the remainder is released as water [11–14]. Cytochrome P450s metabolize AA to produce the collectively designated hydroxyeicosatetraenoic acids and epoxyeicosatrienoic acids; these bioactive compounds are generated in a tissue- and cell-specific manner, and numerous biological functions have been revealed (**Figure 1**).

#### **2. Secretory phospholipase A2**

Phospholipase A2 (EC 3.1.1.4, PLA2) belongs to the group of enzymes, which catalyze the hydrolysis of the ester bond at the sn-2 position of glycerophospholipids and, consequently, are capable of generating free fatty acids, including arachidonic acid (AA). Under physiological conditions, PLA2s are crucial for membrane phospholipid homeostasis, ensuring membrane stability, fluidity, and permeability, and they are involved in the regulation of transport processes through the cell membrane. Phospholipases A2 are enzymes widely diffused in bacteria, plants, venom (of various animals), and mammal cells. Several studies suggest that these enzymes can be classified into 19 groups, which have been identified in mammalian tissues. Besides, many of these groups exhibit significant A2 phospholipase enzymatic activity. At a high level, PLA2s can be classified into two groups: cytosolic PLA2 (cPLA2), and a large and diverse group of secretory PLA2s (sPLA2). Cytosolic PLA2 comprises calcium-dependent cPLA2 (cPLA2), calciumindependent cytosolic PLA2 (iPLA2), lysosomal PLA2 (lPLA2), mitochondrial PLA2 (mPLA2), and, more recently, PAF-acetyl hydrolases (PAF-AHs), which display a small family of phospholipases A2 with high specificity for hydrolysis of the unsaturated fatty acid residue located at the sn-2 position [7, 8, 10]. Several

studies suggest that the proinflammatory action induced by mammalian sPLA2 and even snake venom sPLA2 involve a significant increase of both oxidative activity and reactive oxygen species (ROS) in the cell. ROS are involved in processes such as lipid peroxidation and protein carbonylation, which, at certain levels, can lead to pathological events [15]. Studies conducted by Chiricozzi et al. (2010) [16] reveal that there is a relationship between the increased enzymatic activity of sPLA2, which belongs to the IIA family, and a significant cellular production of free radicals, which contribute strongly to the development of neurodegenerative diseases. Snake venom sPLA2 shares similar mechanisms of action and the same pathways of action with mammalian sPLA2. Experimental evidence in the literature demonstrates that both sPLA2 isoforms are able to induce inflammation and other similar biological activities [10, 17–19]. It is noteworthy that literature data demonstrate sPLA2 can activate signaling events that cannot be explained simply by its catalytic activity, and this fact emphasizes that sPLA2 could act essentially as a ligand of a receptor, rather than as an enzyme [20]. In contrast, studies suggest that products generated by sPLA2 may act as second intracellular messengers, and its enzymatic activity provides a crucial point in the biosynthesis pathways of several classes of inflammatory mediators [21]. In addition, studies performed with other sPLA2s suggest that, during the inflammatory process, leukocytes are recruited to the damaged site (via chemotaxis), where there are conditions necessary to produce a "respiratory explosion." This condition is characterized by high oxygen consumption and the production of reactive oxygen species (ROS), such as the superoxide anion radical (O2 −• ) and hydrogen peroxide (H2O2), which can generate the hydroxyl radical (•OH) directly or indirectly through chemical reactions, such as Fenton and Harber Weiss [22].

Nucleic acids, proteins, and lipids are important targets of ROS, and their attack may lead to an increased risk of mutagenesis due to the modification of these molecules. Moreover, during the inflammatory process, they synthesize soluble mediators, such as arachidonic acid metabolites, cytokines, and chemokines, which lead to the recruitment of more cells that are involved in the inflammatory process to the injured site, thus increasing ROS production. These key mediators may activate signal transduction cascades and induce changes in transcription factors, such as nuclear transcription factor κ-β (NFκ-β) and signal transducer/transcriptional activator 3 (STAT 3), which mediate the response to cellular stress. In addition, induction of cyclooxygenase-2 (COX2) was reported to contribute to nitric oxide synthesis by the enzyme inducible nitric oxide synthetase (iNOS), besides the increased expression of tumor necrosis factor (TNF-α), interleukin-1 (IL-1), interleukin-6 (IL-6), and alterations in the expression of specific microRNAs [23, 24]. It should be noted that nitric oxide can form reactive nitrogen species (RNS) that are highly damaging to cells [25, 26]. Signaling of inflammation is recognized globally by IL-1, IL-6, and TNF-α through Toll-like receptors (TLRs), which belong to the IL-1R family. IL-1 and TNF-α represent the proinflammatory cytokine archetypes that are readily released in response to tissue injury or infection, and they represent a programmed recognition system to trigger inflammation [27–29]. It is important to note that although nitric oxide (NO<sup>−</sup>• ), generated by iNOS, has been revealed to have an essential role as a cellular marker, in an environment with oxidative stress, it can react with O2 −• to generate peroxynitrite (NOO<sup>−</sup>) and other harmful RNS species [26, 30]. Some authors suggest that preventing the formation of NOO- or inducing its efficient decomposition in inflammatory processes may result in a new therapeutic strategy for the treatment of inflammatory processes [30]. In this context, enzymes such as glutathione peroxidase (Gpx) and peroxiredoxin (Prx) appear to have great importance, since they respond to NOO<sup>−</sup> decomposition with high efficiency [30–33].

*Edema Induced by sPLA2 from* Crotalus durissus terrificus *Involves PLC and PKC Signaling… DOI: http://dx.doi.org/10.5772/intechopen.80848*

#### **3. Edema induced by sPLA2 from** *Crotalus durissus terrificus* **involves oxidative stress signaling**

There is no significant evidence that enzymatic toxins from snake venom are able to increase cellular oxidative stress during inflammation [34]; there has been neither a molecular nor a physiological connection shown between edema and other pharmacological activities induced by secretory phospholipase A2 from *Crotalus* 

**Figure 2.**

*Edema values induced by Cdt sPLA2 at the adjusted concentration of 10 μg/site (n = 5). Blood and tissue samples were collected from the animals in two phases: at 30 min (B–D) and 90 min (E–F). Measurements of COX2, PGE2, and MDA levels are representative of the analysis of five animals.*

*durissus terrificus* (Cdt sPLA2). However, our results show there is a biochemical, physiological, and temporal connection between the AA metabolism induced by sPLA2, culminating in edema, and the increase of cellular oxidative stress, which was evaluated by measuring malondialdehyde (MDA) content. MDA is a highly reactive three-carbon dialdehyde produced as a byproduct of polyunsaturated fatty acid peroxidation and AA metabolism. This compound produced by oxidative stress can interact with several molecules, including proteins, lipoproteins, and DNA. The main source of MDA in biological samples is the peroxidation of polyunsaturated fatty acids with two or more methylene-interrupted double bonds [35, 36]. H2O2 represents a messenger capable of altering redox homeostasis, contributing, at various levels, to related inflammatory diseases. Although H2O2 is not an inherently reactive compound, it can be converted into highly reactive and deleterious products that kill cells. In this context, several studies have shown that plant phenolic compounds have great neutralization capacity toward hydrogen peroxide, because these compounds can donate electrons to hydrogen peroxide and neutralize it as water [37, 38].

The edema values plotted in **Figure 2A** were obtained by subtracting the edema values induced by saline (negative control). In this work, we evaluated the activity of COX2 and quantified PEG2 and MDA in blood and tissue samples collected at two different time points—30 and 90 min after sPLA2 administration. **Figure 2A** shows that the amount of COX2 present in swollen tissue after a 5 μg/site Cdt sPLA2 injection was 18.7 ± 1.23 ng/mL (n = 5), compared to values resulting from saline injection that were close to zero. In **Figure 2B**, quantification of PGE2 in the blood of animals collected after Cdt sPLA2 injection (5 μg/site) reveals a concentration of 783 ± 32.4 pg./mL (n = 5), while the saline treatment resulted in 65 ± 18.6 pg./mL (n = 5). Thus, the amount of PGE2 was 12-fold higher than the control values. MDA, produced during lipid peroxidation, is widely used for determining oxidative stress, and the results (shown in **Figure 2C**) indicate that the amount of MDA in plasma was 17.82 ± 8.65 nmol, whereas the amount of MDA released after the saline injection was 0.58 ± 0.22 nmol (n = 5). The results presented in **Figure 2A**–**C** were obtained before the edema peak, and they show that COX2, PGE2, and MDA levels were extremely high in comparison with the control. However, the samples from the material collected at 90 min or after the peak of edema showed that the COX2, PGE2, and MDA levels did not significantly vary from the control (saline), as shown in **Figure 2D**–**F**.

#### **4. Edema induced by sPLA2 from** *Crotalus durissus terrificus* **involves PLC and PKC signaling**

The metabolism of AA is a crucial point in the course of proinflammatory secretory phospholipase A2 (sPLA2). These enzymes basically have two distinct molecular domains, one involved in catalysis and the other responsible for receptor interaction, which allows sPLA2 to mobilize other enzymes involved in the production of proinflammatory mediators. In addition, studies indicate that sPLA2 receptors can mediate their activity through G-protein, and therefore, they can trigger the activation of phospholipase C (PLC), activating the phosphokinase C (PKC) signaling pathway and leading to potentialization of cytoplasmic PLA2 (cPLA2) and COX2. In **Figure 2A**, we show the effect of the different treatments on edema induced by sPLA2 of *Crotalus durissus terrificus* (Cdt sPLA2). In **Figure 2A**, the results clearly show that the edema peak induced by sPLA2 produces an increase of 0.278 ± 0.016 mL (5 μg/site; n = 5). About 20 μL of PKC inhibitor (GF109203X; Tocris 30 mg/kg, dissolved in 0.5% DMSO) was injected by endovenous route 30 min (n = 5) before administering sPLA2. The PKC inhibitor was able to significantly reduce edema induced by sPLA2, which was 0.123 ± 0.018 mL (n = 5).

*Edema Induced by sPLA2 from* Crotalus durissus terrificus *Involves PLC and PKC Signaling… DOI: http://dx.doi.org/10.5772/intechopen.80848*

About 20 μL of PLC inhibitor (U73122; Tocris; 30 mg/kg, dissolved in 0.5% DMSO) was injected intravenously 30 min prior to application of sPLA2, revealing that the peak of edema was 0.167 ± 0.021 mL (n = 5), which was significantly lower than the edema peak induced by sPLA2. In **Figure 2B**, we show the effect of the specific inhibitor against cPLA2 and COX2. To assess the effect of arachidonyl trifluoromethyl ketone (AACOCF3) (Sigma-Aldrich, 30 mg/kg, dissolved in 0.5% DMSO), each animal received 20 μL of the compound by endovenous route 30 min (n = 5) before injecting sPLA2; there was a significant decrease in the edema, revealing a maximum edema of 0.218 ± 0.018 mL (n = 5). About 20 μL of *N*-[2- (cyclohexyloxy)-4-nitrophenyl]methanesulfonamide (NS-398) (Cayman Chemical, 30 mg/kg, dissolved in 0.5% DMSO) was injected intravenously 30 min prior to application of sPLA2, and the peak of the resulting edema was 0.146 ± 0.021 mL (n = 5), which is also significantly lower than the edema peak induced by sPLA2.

The **Figures 1** and **2** show that sPLA2 triggers proinflammatory activity by a signaling pathway involving PKC and PLC. In the case of PLC, two products are generated, diacylglycerol (DAG) and inositol triphosphate (IP3), which can induce

#### **Figure 3.**

*Values of edema induced by sPLA2 of Cdt at the adjusted concentration of 10 μg/site (n = 5). (A) The effect of the inhibitor of PKC (PKC inhibitor 30′) and inhibitor of PLC (PLC inhibitor 30′). In (B), we evaluated the edema induced by sPLA2 in the presence of a specific inhibitor of cPLA2 (AACOCF3) and inhibitor of COX2 (NS-398).*

the phosphorylation of several proteins [14, 39–43]. Thus, the sPLA2 of *Crotalus durissus terrificus* venom may induce an increase in AA metabolism through the interaction of Cdt sPLA2 with G-protein coupled cellular receptors, which activate PLC, generating PUFAs and AA. **Figures 2** and **3** present evidence of interconnections and pathways that generate AA, PLC, and PKC, with cPLA2 and COX2 revealing a possible route of signaling and mobilization of AA, and which could include PUFA release from membrane phospholipids. **Figure 2** also shows that the edema induced by sPLA2 involves the presence of ROS and lipid peroxidation, and that the AA produced can be oxidized to generate MDA as one of the byproducts [39, 44–46]. The results shown in **Figures 2** and **3** suggest that increased cellular oxidative stress and AA mobilization happen intensely and quickly. In this work, we have shown a possible mechanism of edema action induced by sPLA2 from *Crotalus durissus terrificus*, suggesting that the enzymatic activity of Cdt sPLA2 may participate in the inflammatory process, but this activity could also involve the presence of cellular receptors. sPLA2 induces two mechanisms. One mechanism increases oxidative stress, especially in the form of hydrogen peroxide, which leads to increased MDA concentrations; thus, increased oxidative stress has a relevant role in the course of edema. On the other hand, edema induced by sPLA2 also involves a PLC signaling pathway, which mobilizes IP3 (and intracellular calcium) and DAG. These two compounds potentiate the PKC signaling pathway and can lead to a significant increase of cPLA2 through cPLA2 phosphorylation, and this results in enhanced AA metabolism via COX2, an enzyme that could be a second important point in the control of induced inflammation by sPLA2 from *Crotalus durissus terrificus*.

#### **5. "To be or not to be" enzymatically active important for Cdt sPLA2 inflammation**

A great question that arises for characterizing the pharmacological and biological activity of Cdt sPLA2 is the importance of the enzymatic activity of sPLA2. For many years, several studies concluded that all biological, physiological, pharmacological, and pathological activity depended on the enzymatic activity of sPLA2, and this remained unanimous until the 1990s. In 1984, the structure and function of the basic sPLA2 of *Agkistrodon piscivorus* were elucidated, leading to the first structural characterization of basic Lys49 sPLA2 [47]. This enzyme also exhibits a moderate enzymatic activity on membrane phospholipids [47]. Subsequently, several works with purified Lys49 basic sPLA2 from snake (*Bothrops sp.*) were able to induce several pharmacological activities, such as pronounced edema, myonecrosis, oxidative stress, nephrotoxicity, insulin degranulation, and anticoagulant activity [17, 48–52]. In the case of the sPLA2 from Cdt, it was observed that its enzymatic activity can be practically abolished through treatment with certain compounds. Numerous natural compounds have the potential to downregulate or modulate the PLA2 activities, as well as other enzymes involved in AA metabolism, including cPLA2 or enzymes involved in prostaglandin metabolization [52–56]. One of the most abundant polyphenols in the human diet, 5-caffeoylquinic acid (5CQA), exerts potent anti-inflammatory, antibacterial, and antioxidant activities. The anti-inflammatory activity of 5CQA may involve multiple mechanisms of action, including the inhibition of the production and secretion of chemical mediators involved in the inflammatory process.

In **Figure 3A**, we show the effect of 5CQA on edema induced by purified sPLA2 from Cdt. When incubated with sPLA2, 5CQA forms a stable molecular complex and may interact with the catalytic site of the protein and strongly decrease its enzymatic activity, changing the secondary structure and leading to the virtual abolishment of sPLA2 enzymatic activity. The edematogenic assay performed with

#### *Edema Induced by sPLA2 from* Crotalus durissus terrificus *Involves PLC and PKC Signaling… DOI: http://dx.doi.org/10.5772/intechopen.80848*

native sPLA2 and 5CQA incubated with sPLA2 clearly showed that edema induced by sPLA2:5CQA was not abolished, but significantly diminished (**Figure 4A**). Thus, in part, the anti-inflammatory effect of 5CQA probably involves the downregulation of pharmacological and enzymatic activity of sPLA2 [57, 58]. In **Figure 3B**, we show the effect of p-bromophenacyl bromide (p-BPB) and umbelliferone (7-HOC) on edema induced by sPLA2. These data reveal that previous treatment with sPLA2/7-HOC highly decreased the proinflammatory effect induced by sPLA2 purified from Cdt, whereas previous treatment with p-BPB abolished this effect.

Unlike flavonoids, both compounds 7-HOC and p-BPB chemically react with the structure of sPLA2 and form highly stable molecular complexes, both inducing large structural modifications that lead to the virtual abolishment of the enzymatic activity of sPLA2. However, the edematogenic experiments conducted with both compounds incubated with sPLA2 did not abolish the proinflammatory effect induced by the protein, as shown in **Figure 3B**. Thus, in this case, comparison between the results from pharmacological assays suggests that the abolishment of enzymatic activity did not suppress or inhibit the pharmacological effect of sPLA2. This paradox between enzymatic activity and pharmacological effect suggests that at least one more complex pharmacological mechanism is involved in the enzymatic activity, which is independent of the enzymatic activity only. These facts suggest the existence of a distinct pharmacological site, as already proposed by [10, 20].

#### **Figure 4.**

*In (A), we show paw edema induced after the injection of sPLA2 and sPLA2:5CQA (10μg/paw) into the right paw of Swiss mice. Measurements were performed after 30, 60, 120, 180, and 240min, and statistical differences were observed with sPLA2 incubated with 5CQA. In (B), we show enzymatic activity analyzed using 4N3OBA as a substrate, then monitored at a wavelength of 425nm. In this condition, we examined the effect of the substrate on the enzymatic activity of the native and 5CQA-pretreated sPLA2 (sPLA2:5CQA). Chemical treatment of sPLA2 with 5CQA shifts both the Km and Vmax of the native sPLA2. In (C), we show the mouse paw edema induced by untreated sPLA2 and sPLA2 treated with umbelliferone (sPLA2:7-HOC) or with p-bromophenacyl bromide (sPLA2:p-BPB). Doses of 10 μg/paw were used. Observations were conducted at intervals of 30, 60, 90, 120, and 180 min. (D) Results of enzymatic kinetic analysis of untreated (sPLA2) and 7-HOC- or p-BPB-treated sPLA2 (sPLAs:7-HOC) using 4N3OBA as substrate. sPLA2 Vmax; sPLA2:7- HOC Vmax. For the enzymatic assay results in (B) and (D), each point represents the mean ± SEM of n = 12 and \*p < 0.05, and in (A) and (C), each point represents the mean ± SEM of five experiments and \*p < 0.05.*

The authors performed several mutagenesis experiments besides those analyzing its catalytic site; there is another pharmacological site located in the calcium binding loop, and the presence of a second pharmacological site has also been considered by [8, 59, 60]. Thus, the enzymatic activity of sPLA2 from Cdt is not crucial for its pharmacological effect and involves other molecular regions, which are collectively designated as pharmacological sites [51, 61]. Some studies performed with sPLA2 from *Crotalus durissus ssp.* showed that the calcium binding loop is involved in the pharmacological activity [57], and others performed by [52] showed that regions close to the active site of sPLA2 could also be involved. According to [54], the C-terminal region could also participate in the interaction with pharmacological receptors. Even so, the crucial and commonly raised point is that the decreased enzymatic activity of Cdt sPLA2 is not accompanied by a proportional decrease in the proinflammatory activity of this enzymatic toxin, as shown by treatment of Cdt sPLA2 with p-BPB (**Figure 4**).

#### **6. Analysis of peroxiredoxins during edema induced by sPLA2 from**  *Crotalus durissus terrificus*

Oxidative stress is implicated in numerous proinflammatory responses in mammalian cells. H2O2 is known to trigger the release and metabolism of AA in various cell types, but the mechanisms involved appear to diverge profoundly from one cell to another. Thus, mobilization of AA in response to oxidative stress appears to be a very complex process involving potentially multiple enzymes and pathways. Studies reveal that the pathological actions induced by sPLA2 from snake venom involve the induction of significant increases in proinflammatory mediators that may also induce a significant rise in reactive oxygen species levels, which can effectively lead to the establishment of numerous events. Thus, the decrease or control of the concentration of these reactive oxygen species may contribute to the decrease of several pathological actions induced by the A2 secretory phospholipase venom. This is evidenced in some studies, such as those that used plant extracts with antioxidant action. The increase in the cellular oxidative process resulting from the mobilization of AA is, in short, associated with the mobilization of H2O2 [62–64]; however, this event is not known to be the case for the sPLA2 found in several snake venoms. Some studies show that there is a direct cause and effect relationship between the increased expression of several calcium-dependent PLA2 isoforms and the increased concentration of hydrogen peroxide. Besides, this mechanism involves the presence of G-protein-bound cellular receptors and the consequent protein kinase activation. In addition, much data support the possible existence of cross talk between cPLA2 and sPLA2 while eliciting a full AA release response [63, 65, 66]. During the action of secretory and cytosolic A2 phospholipases, a large amount of AA is produced, which can be considered one of the major components that may be reduced via enzymatic peroxidation to prostaglandins, leukotrienes, thromboxanes, and other cyclooxygenase-, lipoxygenase-, or cytochrome P-450-derived products. Thus, during the process of oxidative stress, AA and other bioactive lipids can be converted into lipid hydroperoxide (LOOH). LOOHs are the primary products of lipid peroxidation, which are relatively stable and long lasting compared to other ROS. Among the many different aldehydes, which can be formed as secondary products during lipid peroxidation, MDA appears to be the most mutagenic [36, 56, 67].

The most accepted paradigm is that oxidative stress initiates a chain reaction of lipid peroxidation, which can be reduced by the presence of tocopherol (e.g., vitamin E) or some other chain-breaking antioxidant. However, several

#### *Edema Induced by sPLA2 from* Crotalus durissus terrificus *Involves PLC and PKC Signaling… DOI: http://dx.doi.org/10.5772/intechopen.80848*

studies have shown that these antioxidants do not neutralize the oxidized phospholipids that were formed prior to the application of these compounds. Thus, lipid peroxidation is not spontaneously reversible, and enzymatic pathways that return lipids to their reduced states have been described. On the other hand, several authors showed that peroxiredoxins (Prxs), particularly Prx 6, play an essential role in the reduction of H2O2 and short hydroperoxides; besides, they can directly reduce phospholipid hydroperoxides. Prxs are thiol-dependent peroxidases that catalyze the reduction of a wide variety of hydroperoxides, and the catalytic activity is provided by the presence of a highly conserved catalytic cysteine residue, whose oxidation by hydroperoxide generates sulfenic acid (Cys-SOH). The Prx reduction mechanism involving Cys-SOH is a matter of debate, with glutaredoxin 2 (GRX2), thioredoxin 3 (Trx3), thioredoxin reductase 2 (Trr2), and ascorbate being proposed as possible reducers [68–70]. Several other studies revealed that, during oxidative stress, several Prxs are overexpressed, which can be used as a sensor of oxidative stress in several cells [71–73]. Thus, Prxs represent a group of antioxidant proteins able to decompose several types of hydroperoxides at rates of 105–8 M/second. These enzymes utilize a cysteine residue, which, after the peroxide decomposition, oxidizes (CP-SOH), forming a disulfide bond with a second cysteine, which is reduced by the enzymes thioredoxin (Trx) and thioredoxin reductase (TrxR). In addition, several drugs have been characterized as peroxiredoxin inhibitors, and their use has been helpful in unraveling the physiological and biological roles of certain peroxiredoxins. Among these Prx inhibitors, the best known is adenanthin (Adn), which inhibits Prxs I, Prx II, and other thiol-dependent antioxidant enzymes [74, 75]. Another commercial drug is MJ33, which is described as a potent inhibitor of Prx 6, an extremely essential enzyme for regulating oxidative stress, inflammation, and NADPH oxidase (NOX)2 activation [76]. In addition, conoidin A (ConA) is characterized as a potent inhibitor of peroxiredoxin II, an antioxidant enzyme that acts in the intracellular signaling and defense against oxidative stress [77]. Enzyme inhibition is one of the ways in which enzyme activity is regulated experimentally and naturally.

In the case of the pharmacological tests, inoculation of 5 μg sPLA2 purified from the total venom of *Crotalus durissus terrificus* induced an inflammatory reaction, revealing a typical acute edema with a peak at 60 min (**Figure 5**). To assess the effects of inhibitors, MJ33, ConA, and Adn were injected intraperitoneally (2 μg/g mice) 30 min prior to administration of PLA2 isolated from Cdt venom. As shown in **Figure 5A**, MJ33 showed insignificant anti-inflammatory activity that was only observed along with the edema peak. **Figure 5B** shows the effect of ConA administrated before sPLA2, revealing insignificant inhibition of edema. Although both MJ33 and ConA are essential Prx inhibitors, they display some limitations, as found with MJ33, which is a specific inhibitor of Prx 6. Prx 6 is a complex Prx, exhibiting its maximal antioxidant activity only at acidic pH values [78].

Prx 6 shows calcium-independent phospholipase A2 enzyme activity that is also maximal at acidic pH [79]. The determination of its functional and enzymatic properties was recently elucidated. The low MJ33 inhibitory effect observed in our study could have been due to the presence of a calcium-independent PLA2 domain. Some studies showed that Prx 2 appear to be an essential negative regulator of LPS-induced inflammatory signaling through modulation of ROS synthesis via NADPH oxidase activities; therefore, Prx 2 is crucial for the prevention of excessive host responses to microbial products [80]. Although ConA shows the ability to covalently inhibit Prx 2 activity, the results presented in **Figure 5B** suggest that Prx 2 does not play a relevant role in reducing edema induced by Cdt sPLA2. On the other hand, LPS stimulates monocytes/macrophages through Toll-like receptor

#### **Figure 5.**

*In (A), we show paw edema induced after the injection of sPLA2 and sPLA2:MJ33 (5 μg/paw) into the right paw of Swiss mice. Measurements were performed after 30, 60, 120, 180, 240 and 480 min, and statistical differences were observed with sPLA2 applied after MJ33 injection 30 minutes before sPLA2 injection. In (B), we show paw edema induced after the injection of sPLA2 and sPLA2:ConA (5 μg/paw) into the right paw of Swiss mice. Measurements were performed after 30, 60, 120, 180, 240 and 480 min, and statistical differences were observed with sPLA2 incubated with ConA (conoidin A) applied 30 minutes before sPLA2. In (C), we evaluate the effect of sPLA2 in comparison with adenanthin (Adn) previously applied 30 min before sPLA2. Each point represents the mean ± SEM of five experiments and \*p < 0.05.*

4 (TLR4), resulting in a series of signaling activation events, which potentiate the production of inflammatory mediators, such as IL-6 and TNF-α [81, 82]. The results presented in **Figure 5C** clearly show that thiol-dependent antioxidant enzymes play an essential role in edema control and recovery induced by sPLA2 purified from

Cdt, and, similar to ConA and MJ33, these enzymes did not exhibit an inhibition or decrease of the edema peaks that occur at 60 min. **Figure 5C** also reveals that the edematogenic effect induced by sPLA2 diminished after 60–90 min, and the hind paw volume returned to its normal volume after 240 min. However, in animals treated with Adn 30 min before the sPLA2 injection, the edematogenic effect persisted for even 8 h after the experiment.

### **7. Conclusion**

During inflammation (edema), induced by purified sPLA2, arachidonic acid generation and its metabolization by COX2 during the edema play crucial roles during this pharmacological event. Arachidonic acid can be mobilized by the catalytic activity of sPLA2 from *Crotalus durissus terrificus* (or other sources) or by activation of cytosolic PLA2. The enzymatic activity of secretory PLA2 (sPLA2) was not crucial for this initial mobilization, and the presence of sPLA2 receptors plays a crucial role in the mobilization of high amounts of arachidonic acid (AA). The classic AA production pathway, which basically involves cPLA2 modulation, also involves the interaction of a more complex pathway that includes the activation of PLC, producing IP3 and DAG. In turn, IP3 and DAG activate PKC, stimulating a strong increase of AA by cPLA2 [1, 83, 84]. However, AA is also mobilized by two other distinct pathways. One involves PLC activation, which has an essential role in AA production by DAG lipase and MAG lipase. In this pathway, catalysis leads to diacylglycerol hydrolysis, releasing a free fatty acid and monoacylglycerol as 2-acyl glycerol, which is converted to AA by MAG lipase action [85–87].

Another pathway that is initiated during AA mobilization involves the release of platelet aggregation factor (PAF)—another subproduct of the enzymatic hydrolysis of membrane phospholipids that cross through the cell membrane—and its specific receptor (PAF receptor or PAF-R) leads to the stimulation of PLC by G-protein [83, 88]. Thus, it is possible that sPLA2 from snake venom, such as venom from *Crotalus durissus terrificus*, mobilizes AA by three different pathways, and AA oxidative metabolism is a key factor that induces increased ROS and oxidative stress during edema. In addition, there are several studies that show AA production is an important way to increase the generation of hydrogen peroxide during inflammation. Thus, it is possible that the action of sPLA2 also increases cell oxidative stress and AA metabolism, culminating in the production of PGE2 and MDA [36]. All this occurs through the interaction of sPLA2 with its receptors to modulate the activity and function of cPLA2 and iPLA2, inducing a significant increase in AA metabolism and COX2 expression, a fact that contributes to the production of free radicals (**Figure 6**) [45, 89–95].

Several studies have shown that arachidonic acid produced by the action of sPLA2 and cPLA2 can activate NADPH oxidase (NOX) enzymes and induce a significant increase in hydrogen peroxide, which gains entry to the intracellular environment through aquaporins and has a predominant role in increasing cellular oxidative stress [91–96]. This would explain the importance of thioldependent antioxidant enzymes playing key roles in the control of edema induced by *Crotalus durissus terrificus* sPLA2. On other side, the inflammation (edema) induced by sPLA2 involves the mobilization of arachidonic acid and hydrogen peroxide, and both are the main elements involved in the inflammatory process. The data compiled in this work suggest that oxidative stress is integral in the progression and maintenance of inflammation (edema) induced by sPLA2 from

*Crotalus durissus terrificus*. Furthermore, our results show that *Crotalus durissus terrificus* sPLA2-induced edema is strongly regulated by thiol-dependent enzymes, and that adenanthin (Adn) was able to neutralize this control and the inflammatory process (edema).

On the other hand, several articles have reported that natural antioxidant compounds, such as flavonoids and related substances, when given prior to sPLA2 injection, have significant anti-inflammatory activities. This probably stems from the ability of many of these compounds to partially inhibit the enzymatic and pharmacological activities of sPLA2 from *Crotalus durissus terrificus*, as well as from their strong antioxidant activities [53–56, 97]. Thus, the search for new natural compounds with anti-inflammatory properties remains an important area of research.

**Figure 6.** *Summary of possible inflammation mechanism of Cdt sPLA2 action during the inflammatory process.*

*Edema Induced by sPLA2 from* Crotalus durissus terrificus *Involves PLC and PKC Signaling… DOI: http://dx.doi.org/10.5772/intechopen.80848*

#### **Acknowledgements**

The present project had the financial support of the FAPESP process no: 2017/20291-0 on the responsibility of Professor M.H. Toyama and the resources coming from the FAPESP process no: 2017/19942-7, to CNPq and UNESP.

#### **Conflict of interest**

The authors have no conflict of interests to declare.

### **Author details**

Marcos H. Toyama1 \*, Caroline R.C. Costa1 , Mariana N. Belchor1 , Danielle P. Novaes1 , Marcos A. de Oliveira<sup>2</sup> , Rolando Ie2 , Henrique Hessel Gaeta1 and Daniela de O. Toyama1

1 UNESP, Institute of Biosciences, Campus do Litoral Paulistac (CLP), BIOMOLPEP, São Vicente, São Paulo, Brasil

2 UNESP, Institute of Biosciences, Campus do Litoral Paulista (CLP), LABIMES, São Vicente, São Paulo, Brasil

\*Address all correspondence to: marcoshikaritoyama@gmail.com

© 2019 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

#### **References**

[1] Hanna VS, Hafez EAA. Synopsis of arachidonic acid metabolism: A review. Journal of Advanced Research. 2018;**11**:23-32

[2] Liu C et al. Arachidonic acid metabolism pathway is not only dominant in metabolic modulation but associated with phenotypic variation after acute hypoxia exposure. Frontiers in Physiology. 2018;**9**:236

[3] Hosford D, Braquet P. Plateletactivating factor (PAF), P. J. B. T.-E. of I. E. Delves, 2nd ed. Oxford: Elsevier, 1998, pp. 1971-1973.

[4] da Silva IA Jr, Chammas R, Lepique AP, Jancar S. Platelet-activating factor (PAF) receptor as a promising target for cancer cell repopulation after radiotherapy. Oncogene. 2017;**6**:e296

[5] Kasperska-Zajac A, Brzoza Z, Rogala B. Platelet-activating factor (PAF): A review of its role in asthma and clinical efficacy of PAF antagonists in the disease therapy. Recent Patents on Inflammation & Allergy Drug Discovery. 2008;**2**(1):72-76

[6] Xu B et al. Effects of plateletactivating factor and its differential regulation by androgens and steroid hormones in prostate cancers. British Journal of Cancer. 2013;**109**(5):1279-1286

[7] Murakami M, Taketomi Y, Miki Y, Sato H, Hirabayashi T, Yamamoto K. Recent progress in phospholipase A2 research: From cells to animals to humans. Progress in Lipid Research. 2011;**50**(2):152-192

[8] Murakami M, Sato H, Miki Y, Yamamoto K, Taketomi Y. A new era of secreted phospholipase A 2. Journal of Lipid Research. 2015;**56**(7):1248-1261

[9] Yarla NS, Bishayee A, Vadlakonda L, Chintala R, Duddukuri GR, Reddanna P, et al. Phospholipase A2 isoforms as novel targets for prevention and treatment of inflammatory and oncologic diseases. Current Drug Targets. 2016;**17**(16):1940-1962

[10] Murakami M, Lambeau G. Emerging roles of secreted phospholipase A2 enzymes: An update. Biochimie. 2013;**95**(1):43-50

[11] Pompeia C, Cury-Boaventura MF, Curi R. Arachidonic acid triggers an oxidative burst in leukocytes. Brazilian Journal of Medical and Biological Research. 2003;**36**(scielo):1549-1560

[12] Covey TM, Edes K, Fitzpatrick FA. Akt activation by arachidonic acid metabolism occurs via oxidation and inactivation of PTEN tumor suppressor. Oncogene. 2007;**26**:5784

[13] Chen J-K, Capdevila J, Harris RC. Cytochrome P450 Epoxygenase metabolism of arachidonic acid inhibits apoptosis. Molecular and Cellular Biology. 2001;**21**(18):6322-6331

[14] Balboa MA, Balsinde J. Oxidative stress and arachidonic acid mobilization. Biochimica et Biophysica Acta—Molecular and Cell Biology of Lipids. 2006;**1761**(4):385-391

[15] Yagami T et al., Human group IIA secretory phospholipase A2 induces neuronal cell death via apoptosis. Molecular Pharmacology. 2002;**61**(1):114-126

[16] Elena C, Seila F-F, Vincenza N, Angeles A, Pedro BJ, Gianfrancesco G. Group IIA secretory phospholipase A2 (GIIA) mediates apoptotic death during NMDA receptor activation in rat primary cortical neurons. Journal of Neurochemistry. 2010;**112**(6):1574-1583

[17] Fagundes FHR, Aparicio R, dos Santos ML, Filho EBSD, Oliveira SCB, *Edema Induced by sPLA2 from* Crotalus durissus terrificus *Involves PLC and PKC Signaling… DOI: http://dx.doi.org/10.5772/intechopen.80848*

Toyama DO, et al. A catalytically inactive Lys49 PLA2 isoform from *Bothrops jararacussu* venom that stimulates insulin secretion in pancreatic beta cells. Protein & Peptide Letters. 2011;**18**(11):1133-1139

[18] Ximenes RM et al. Harpalycin 2 inhibits the enzymatic and platelet aggregation activities of PrTX-III, a D49 phospholipase A (2) from *Bothrops pirajai* venom. BMC Complementary and Alternative Medicine. 2012;**12**:139

[19] Murakami M, Taketomi Y. Secreted phospholipase A2 and mast cells. Allergology International. 2015;**64**(1):4-10

[20] Lambeau G, Gelb MH. Biochemistry and physiology of mammalian secreted phospholipases A2. Annual Review of Biochemistry. 2008;**77**(1):495-520

[21] Farooqui AA, Horrocks LA. Phospholipase A₂-generated lipid mediators in the brain: The good, the bad, and the ugly. Neuroscience. 2006;**12**(3):245-260

[22] Halliwell B, Gutteridge JMC. Reactive species in disease: Friends or foes? In: Free Radicals in Biology and Medicine. 5th ed. Oxford: Oxford University Press; 2015

[23] Hoesel B, Schmid JA. The complexity of NF-κB signaling in inflammation and cancer. Molecular Cancer. 2013;**12**:86

[24] Kalisperati P et al. Inflammation, DNA damage, helicobacter pylori and gastric tumorigenesis. Frontiers in Genetics. 2017;**8**:20

[25] Szabó C. Hydrogen sulphide and its therapeutic potential. Nature Reviews. Drug Discovery. 2007;**6**:917

[26] Orient A, Donkó Á, Szabó A, Leto TL, Geiszt M. Novel sources of reactive oxygen species in the human body.

Nephrology, Dialysis, Transplantation. 2007;**22**(5):1281-1288

[27] Lawrence T. The nuclear factor NF-κB pathway in inflammation. Cold Spring Harbor Perspectives in Biology. 2009;**1**(6):a001651

[28] Reuter S, Gupta SC, Chaturvedi MM, Aggarwal BB. Oxidative stress, inflammation, and cancer: How are they linked? Free Radical Biology & Medicine. 2010;**49**(11):1603-1616

[29] Ong ZY et al. Pro-inflammatory cytokines play a key role in the development of radiotherapy-induced gastrointestinal mucositis. Radiation Oncology. 2010;**5**(1):22

[30] Knoops B, Argyropoulou V, Becker S, Ferté L, Kuznetsova O. Multiple roles of peroxiredoxins in inflammation. Molecules and Cells. 2016;**39**(1):60-64

[31] Barros LO, Silva SV, Almeida FC, Silva ECB, Carneiro GF, Guerra MMP. Efeito da adição de glutationa peroxidase e cisteína ao diluidor de congelação do sêmen equino. Arquivo Brasileiro de Medicina Veterinária e Zootecnia. 2013;**65**(scielo):430-438

[32] Ribas V, García-Ruiz C, Fernández-Checa JC. Glutathione and mitochondria. Frontiers in Pharmacology. 2014;**5**:151

[33] Brown DI, Griendling KK. Regulation of signal transduction by reactive oxygen species in the cardiovascular system. Circulation Research. 2015;**116**(3):531-549

[34] Meléndez-Martínez D et al. Rattlesnake *Crotalus molossus nigrescens* venom induces oxidative stress on human erythrocytes. Journal of Venomous Animals and Toxins including Tropical Diseases. 2017;**23**:24

[35] Del Rio D, Stewart AJ, Pellegrini N. A review of recent studies on

malondialdehyde as toxic molecule and biological marker of oxidative stress. Nutrition, Metabolism, and Cardiovascular Diseases. 2005;**15**(4):316-328

[36] Ayala A, Muñoz MF, Argüelles S. Lipid peroxidation: Production, metabolism, and signaling mechanisms of malondialdehyde and 4-hydroxy-2-nonenal. Oxidative Medicine and Cellular Longevity. 2014;**2014**:360438

[37] Özyürek M, Bektaşoğlu B, Güçlü K, Güngör N, Apak R. A novel hydrogen peroxide scavenging assay of phenolics and flavonoids using cupric reducing antioxidant capacity (CUPRAC) methodology. Journal of Food Composition and Analysis. 2010;**23**(7):689-698

[38] Lee I-T, Lin C-C, Lin W-N, Wu W-L, Hsiao L-D, Yang C-M. Lung inflammation caused by adenosine-5′ triphosphate is mediated via Ca2+/PKCsdependent COX-2/PGE2 induction. The International Journal of Biochemistry & Cell Biology. 2013;**45**(8):1657-1668

[39] Fonteh AN, Atsumi G, LaPorte T, Chilton FH. Secretory phospholipase A2 receptor-mediated activation of cytosolic phospholipase A2 in murine bone marrow-derived mast cells. Journal of Immunology*.* 2000;**165**(5):2773-2782

[40] Reséndiz JC, Kroll MH, Lassila R. Protease-activated receptor-induced Akt activation—Regulation and possible function. Journal of Thrombosis and Haemostasis. 2007;**5**(12):2484-2493

[41] Holinstat M et al. Proteaseactivated receptor Signaling in platelets activates cytosolic phospholipase a(2) (α) differently for cyclooxygenase-1 and 12-lipoxygenase catalysis. Arteriosclerosis, Thrombosis, and Vascular Biology. 2011;**31**(2):435-442

[42] Carrim N et al. Thrombin-induced reactive oxygen species generation in

platelets: A novel role for proteaseactivated receptor 4 and GPIbα. Redox Biology. 2015;**6**:640-647

[43] Duvernay MT, Matafonov A, Lindsley CW, Hamm HE. Platelet lipidomic profiling: Novel insight into cytosolic phospholipase A2α activity and its role in human platelet activation. Biochemistry. 2015;**54**(36):5578-5588

[44] Sun GY et al. Role of cytosolic phospholipase A(2) in oxidative and inflammatory signaling pathways in different cell types in the central nervous system. Molecular Neurobiology. 2014;**50**(1):6-14

[45] Quach ND, Arnold RD, Cummings BS. Secretory phospholipase A(2) enzymes as pharmacological targets for treatment of disease. Biochemical Pharmacology. 2014;**90**(4):338-348

[46] Soichiro T et al. C-type lectin-like domain and fibronectin-like type II domain of phospholipase A2 receptor 1 modulate binding and migratory responses to collagen. FEBS Letters. 2015;**589**(7):829-835

[47] Chen Y-C, Maraganore JM, Reardon I, Heinrikson RL. Characterization of the structure and function of three phospholipases A2 from the venom of Agkistrodon halys pallas. Toxicon. 1987;**25**(4):401-409

[48] Beghini DG, Toyama MH, Hyslop S, Sodek LC, Novello JC, Marangoni S. Enzymatic characterization of a novel phospholipase A2 from *Crotalus durissus cascavella* rattlesnake (maracambóia) venom. Protein Journal. 2000;**19**(8):679-684

[49] Lee W-H et al. Crystallization and preliminary X-ray diffraction studies of piratoxin II, a phospholipase A2 isolated from the venom of *Bothrops pirajai*. Acta Crystallographica. Section D, Biological Crystallography. 1998;**54** (6 II):1229-1230

*Edema Induced by sPLA2 from* Crotalus durissus terrificus *Involves PLC and PKC Signaling… DOI: http://dx.doi.org/10.5772/intechopen.80848*

[50] Hernandez-Oliveira S, Toyama MH, Toyama DO, Marangoni S, Hyslop S, Rodrigues-Simioni L. Biochemical, pharmacological and structural characterization of a new PLA2 from *Crotalus durissus terrificus* (South American Rattlesnake) venom. Protein Journal. 2005;**24**(4):233-242

[51] Dos Santos ML, Fagundes FHR, Teixeira BRF, Toyama MH, Aparicio R. Purification and preliminary crystallographic analysis of a new Lys49-PLA2 from *B. jararacussu*. International Journal of Molecular Sciences. 2008;**9**(5):736-750

[52] Ximenes RM et al. Inhibition of neurotoxic secretory phospholipases A2 enzymatic, edematogenic, and myotoxic activities by harpalycin 2, an isoflavone isolated from Harpalyce brasiliana Benth. Evidence-Based Complementary and Alternative Medicine. 2012;**12**:139

[53] Iglesias CV et al. Effects of morin on snake venom phospholipase A2 (PLA2). Toxicon. 2005;**46**(7):751-758

[54] Fonseca FV et al. Effect of the synthetic coumarin, ethyl 2-oxo-2Hchromene-3-carboxylate, on activity of *Crotalus durissus ruruima* sPLA2 as well as on edema and platelet aggregation induced by this factor. Toxicon. 2010;**55**(8):1527-1530

[55] Belchor MN et al. Evaluation of rhamnetin as an inhibitor of the pharmacological effect of secretory phospholipase A2. Molecules. 2017;**22**(9):1441

[56] Tamayose CI et al. Non-clinical studies for evaluation of 8-C-rhamnosyl apigenin purified from *Peperomia obtusifolia* against acute edema. International Journal of Molecular Sciences. 2017;**18**(9):1972

[57] Toyama DO, Ferreira MJP, Romoff P, Fávero OA, Gaeta HH, Toyama MH. Effect of chlorogenic acid

(5-caffeoylquinic acid) isolated from *Baccharis oxyodonta* on the structure and pharmacological activities of secretory phospholipase A2 from *Crotalus durissus terrificus*. BioMed Research International. 2014;**2014**:1-10

[58] Toyama DDO et al. An evaluation of 3-rhamnosylquercetin, a glycosylated form of quercetin, against the myotoxic and edematogenic effects of sPLA2 from *Crotalus durissus terrificus*. BioMed Research International. 2014;**2014**(341270):11

[59] Chioato L, Ward RJ. Mapping structural determinants of biological activities in snake venom phospholipases A2 by sequence analysis and site directed mutagenesis. Toxicon. 2003;**42**(8):869-883

[60] Dennis EA, Cao J, Hsu Y-H, Magrioti V, Kokotos G. Phospholipase A2 enzymes: Physical structure, biological function, disease implication, chemical inhibition, and therapeutic intervention. Chemical Reviews. 2011;**111**(10):6130-6185

[61] Toyama DO, Marangoni S, Diz-Filho EBS, Oliveira SCB, Toyama MH. Effect of umbelliferone (7-hydroxycoumarin, 7-HOC) on the enzymatic, edematogenic and necrotic activities of secretory phospholipase A2 (sPLA2) isolated from *Crotalus durissus collilineatus* venom. Toxicon. 2009;**53**(4):417-426

[62] Balboa MA, Balsinde J. Involvement of calcium-independent phospholipase A2 in hydrogen peroxide-induced accumulation of free fatty acids in human U937 cells. The Journal of Biological Chemistry. 2002;**277**(43):40384-40389

[63] Han WK, Sapirstein A, Hung CC, Alessandrini A, Bonventre JV. Cross-talk between cytosolic phospholipase A2α (cPLA2α) and secretory phospholipase A2 (sPLA2) in hydrogen peroxide-induced arachidonic acid release in murine mesangial cells: sPLA2 regulates cPLA2α activity that is responsible for arachidonic acid release. The Journal of Biological Chemistry. 2003;**278**(26):24153-24163

[64] Sun GY, Shelat PB, Jensen MB, He Y, Sun AY, Simonyi A. Phospholipases A2 and inflammatory responses in the central nervous system. Neuromolecular Medicine. 2010;**12**(2):133-148

[65] Martínez J, Moreno JJ. Role of Ca2+-independent phospholipase A2 on arachidonic acid release induced by reactive oxygen species. Archives of Biochemistry and Biophysics. 2001;**392**(2):257-262

[66] Adibhatla RM, Hatcher JF. Phospholipase a(2), reactive oxygen species, and lipid peroxidation In CNS pathologies. BMB Reports. 2008;**41**(8):560-567

[67] Catalá A. Lipid peroxidation of membrane phospholipids generates hydroxy-alkenals and oxidized phospholipids active in physiological and/or pathological conditions. Chemistry and Physics of Lipids. 2009;**157**(1):1-11

[68] Wood ZA, Schröder E, Robin Harris J, Poole LB. Structure, mechanism and regulation of peroxiredoxins. Trends in Biochemical Sciences. 2003;**28**(1):32-40

[69] Cordray P, Doyle K, Edes K, Moos PJ, Fitzpatrick FA. Oxidation of 2-Cys-peroxiredoxins by arachidonic acid peroxide metabolites of lipoxygenases and Cyclooxygenase-2. The Journal of Biological Chemistry. 2007;**282**(45):32623-32629

[70] Rhee SG, Woo HA. Multiple functions of peroxiredoxins: Peroxidases, sensors and regulators of the intracellular messenger H2O2, and protein chaperones. Antioxidants & Redox Signaling. 2010;**15**(3):781-794

[71] Lee D, Moawad AR, Morielli T, Fernandez MC, O'Flaherty C. Peroxiredoxins prevent oxidative stress during human sperm capacitation. Molecular Human Reproduction. 2017;**23**(2):106-115

[72] Fisher AB. Peroxiredoxin 6 in the repair of peroxidized cell membranes and cell signaling. Archives of Biochemistry and Biophysics. 2017;**617**(Supplement C):68-83

[73] Fisher AB, Vasquez-Medina JP, Dodia C, Sorokina EM, Tao J-Q , Feinstein SI. Peroxiredoxin 6 phospholipid hydroperoxidase activity in the repair of peroxidized cell membranes. Redox Biology. 2018;**14**:41-46

[74] Soethoudt M, Peskin AV, Dickerhof N, Paton LN, Pace PE, Winterbourn CC. Interaction of adenanthin with glutathione and thiol enzymes: Selectivity for thioredoxin reductase and inhibition of peroxiredoxin recycling. Free Radical Biology & Medicine. 2014;**77**:331-339

[75] Siernicka M et al. Adenanthin, a new inhibitor of thiol-dependent antioxidant enzymes, impairs the effector functions of human natural killer cells. Immunology. 2015;**146**(1):173-183

[76] Lee I, Dodia C, Chatterjee S, Feinstein SI, Fisher AB. Therapeutic efficacy of MJ33, a novel inhibitor of phospholipase A2 (PLA2) of peroxiredoxin 6 (Prdx6), in LPSinduced acute lung injury (ALI). FASEB Journal. 2013;**27**(1\_supplement):1107.11

[77] Haraldsen JD et al. Identification of conoidin A as a covalent inhibitor of peroxiredoxin II. Organic & Biomolecular Chemistry. 2009;**7**:3040-3048

[78] Chang C-H, Lo W-Y, Lee T-H. The antioxidant peroxiredoxin 6 (Prdx6)

*Edema Induced by sPLA2 from* Crotalus durissus terrificus *Involves PLC and PKC Signaling… DOI: http://dx.doi.org/10.5772/intechopen.80848*

exhibits different profiles in the livers of seawater- and fresh water-acclimated milkfish, Chanos chanos, upon hypothermal challenge. Frontiers in Physiology. 2016;**7**:580

[79] Manevich Y, Reddy KS, Shuvaeva T, Feinstein SI, Fisher AB. Structure and phospholipase function of peroxiredoxin 6: Identification of the catalytic triad and its role in phospholipid substrate binding. Journal of Lipid Research. 2007;**48**(10):2306-2318

[80] Yang C-S et al. Roles of peroxiredoxin II in the regulation of proinflammatory responses to LPS and protection against endotoxin-induced lethal shock. The Journal of Experimental Medicine. 2007;**204**(3):583-594

[81] Fang H et al. Lipopolysaccharideinduced macrophage inflammatory response is regulated by SHIP. Journal of Immunology. 2004;**173**(1):360-366

[82] Hoareau L et al. Signaling pathways involved in LPS induced TNFalpha production in human adipocytes. Journal of Inflammation (London). 2010;**7**:1

[83] Tang X, Edwards EM, Holmes BB, Falck JR, Campbell WB. Role of phospholipase C and diacylglyceride lipase pathway in arachidonic acid release and acetylcholine-induced vascular relaxation in rabbit aorta. American Journal of Physiology-Heart and Circulatory Physiology. 2006;**290**(1):H37-H45

[84] Reisenberg M, Singh PK, Williams G, Doherty P. The diacylglycerol lipases: Structure, regulation and roles in and beyond endocannabinoid signalling. Philosophical Tansactions of the Royal Society of London. Series B, Biological Sciences. 2012;**367**(1607):3264-3275

[85] Labar G, Wouters J, Lambert DM. A review on the monoacylglycerol

lipase: At the Interface between fat and endocannabinoid signalling. Current Medicinal Chemistry. 2010;**17**(24):2588-2607

[86] Chang JW et al. Remarkably selective inhibitors of monoacylglycerol lipase bearing a reactive group that is bioisosteric with endocannabinoid substrates. Chemistry & Biology. 2012;**19**(5):579-588

[87] Grabner GF, Zimmermann R, Schicho R, Taschler U. Monoglyceride lipase as a drug target: At the crossroads of arachidonic acid metabolism and endocannabinoid signaling. Pharmacology & Therapeutics. 2017;**175**:35-46

[88] Soliman ML, Ohm JE, Rosenberger TA. Acetate reduces PGE(2) release and modulates phospholipase and cyclooxygenase levels in neuroglia stimulated with lipopolysaccharide. Lipids. 2013;**48**(7):651-662

[89] Rupprecht G, Scholz K, Beck K-F, Geiger H, Pfeilschifter J, Kaszkin M. Cross-talk between group IIAphospholipase A(2) and inducible NO-synthase in rat renal mesangial cells. British Journal of Pharmacology. 1999;**127**(1):51-56

[90] Patel MI et al. Cytosolic phospholipase a(2)-α: A potential therapeutic target for prostate cancer. Clinical Cancer Research. 2008;**14**(24):8070-8079

[91] Drummond GR, Selemidis S, Griendling KK, Sobey CG. Combating oxidative stress in vascular disease: NADPH oxidases as therapeutic targets. Nature Reviews. Drug Discovery. 2011;**10**(6):453-471

[92] Nisimoto Y, Diebold BA, Constentino-Gomes D, Lambeth JD. Nox4: A hydrogen peroxidegenerating oxygen sensor. Biochemistry. 2014;**53**(31):5111-5120

#### *Inflammation in the 21st Century*

[93] Vieceli Dalla Sega F et al. Specific aquaporins facilitate Nox-produced hydrogen peroxide transport through plasma membrane in leukaemia cells. Biochimica et Biophysica Acta—Molecular and Cell Research. 2014;**1843**(4):806-814

[94] Montezano AC, Touyz RM. Reactive oxygen species, vascular Noxs, and hypertension: Focus on translational and clinical research. Antioxidants & Redox Signaling. 2014;**20**(1):164-182

[95] Lennicke C, Rahn J, Lichtenfels R, Wessjohann LA, Seliger B. Hydrogen peroxide—Production, fate and role in redox signaling of tumor cells. Cell Communication and Signaling: CCS. 2015;**13**:39

[96] Sies H. Hydrogen peroxide as a central redox signaling molecule in physiological oxidative stress: Oxidative eustress. Redox Biology. 2017;**11**:613-619

[97] Ximenes RM et al. Harpalycin 2 Inhibits the Enzymatic and Platelet Aggregation Activities of PrTX-III, a D49 Phospholipase A. BMC Complementary and Alternative Medicine. 2012;**12**:139

#### **Chapter 5**

## Edema Management in Oral and Maxillofacial Surgery

*Renato Yassutaka Faria Yaedu, Marina de Almeida Barbosa Mello, Juliana Specian Zabotini da Silveira and Ana Carolina Bonetti Valente*

#### **Abstract**

This chapter will discuss the expected edema and intercurrences in maxillofacial surgery, which involves important anatomical structures, such as the upper airways. It will also discuss important issues such as intrinsic and extrinsic enhancers of edema and the main consequences of a severe edema setting according to physiological, functional, and psychosocial points of view. Edema assessment and measurement is still performed subjectively in the clinical routine. However, for the accomplishment of studies, more objective forms are being tested, but still not very successful for clinical applicability. It is known that the best way to deal with edema is prevention; so in elective surgeries, much is discussed about the best management forms. This way, besides edema prevention, it is important not to cause unwanted reactions for the patient or in the performed procedure. Therefore, it will also be debated about preoperative medications and their consequences. Another point discussed involves main treatments for the underdeveloping edema and the one already installed, such as manual lymphatic drainage therapy, a treatment that is well known and used in other specialties, but is still very little widespread among maxillofacial surgeons.

**Keywords:** edema, oral surgery, maxillofacial surgery, postoperative period, postoperative care

#### **1. Introduction**

Every surgical procedure presents pain and edema in a variable degree, and many pharmacological and alternative methods have been used in an attempt to control and reduce them.

Maxillofacial surgery acts on the patient's face. The maxillofacial surgical procedures include outpatient surgeries using local anesthesia and also more extensive and invasive procedures under general anesthesia. The most used procedures are exodontia, biopsies, surgical cysts and tumors treatment, bone grafts, rehabilitations with osseous integrable implants, orthognathic surgery, face trauma treatment, and infections treatments.

An inflammatory response is expected after any injury or surgical procedure, in an attempt to defend and repair damage tissues. Inflammatory mediators (prostaglandins, leukotrienes, bradykinin, and others) are released, and consequently,

there is an increase in vascular dilatation and permeability, resulting in an edema. However, when it comes to facial edema, the major concerns are related to airway permeability, making the care with this edema a fundamental step for the treatment. It is known that in outpatient surgeries, the extent and the consequence related to edema are smaller and more predictable than in hospital surgeries, but not less important, as we will discuss further on the topic of complications.

Many studies discuss the importance of edema for such surgeries, especially outpatient procedures, which not always presents significant amounts of edema. Besides that, the discussions about the treatment are not conclusive.

Edema is characterized by the excess of plasma proteins in the interstitial space. Its formation occurs when the lymphatic flow exceeds the transport capacity of the lymphatic system or when this system becomes inefficient in absorbing and transporting these proteins [1]. Although the primary edema is a condition usually developed by vascular and/or congenital diseases, the secondary edema occurs due to a lymphatic system injury, whether by infection, cancer, or surgery [2, 3].

#### **2. Edema: risk factors**

Despite the fact that the edema is part of the inflammatory process, and therefore, a consequence of the surgical process, the severity and localization of it can be related to some factors, intrinsic to the patient or related to the surgery.

The increase in the surgical procedure difficulty due to one or more of these factors directly influences on the severity and extension of the postoperative morbidities [4].

#### **2.1 Preexisting conditions**

Any condition that affects postsurgical inflammatory response directly interferes with the postoperative quality, recovery, and also with the edema formation. Therefore, all efforts are made to maintain airway permeability and prevent its obstruction.

An worrying condition is the angioedema, which results from changes in the immunoglobulins involved in the inflammatory response. Due to the fact that is a severe, acute, and rapidly evolving edema that mainly affects the larynx, pharynx, and face, there is a great risk of airways obstruction, and therefore, it is associated with reintubation and risk of death [5–8].

Unfortunately, the occurrence of angioedema is a difficult prediction factor, mainly if the patient never presented its manifestation. For this reason, a rapid and accurate diagnosis is essential, as well as the establishment of artificial airways and adequate drug treatment [6–8].

#### **2.2 Body mass index (BMI)**

The BMI consists in the division of ratio of body weight per height of the individual. Despite there is no consensus in the literature, some studies have related BMI with the severity of postoperative edema [4, 9–11].

Although is expected that individuals with higher BMI (overweight) develop greater edema, this correlation is not always found. Therefore, on those studies, other variables such as age and gender were considered more influential than BMI in the postoperative edema formation [4, 9].

The relation between BMI and the facial edema occurs because adipose tissue is responsible for most of the pro-inflammatory cytokines. So people with a higher BMI have more adipose tissue, more inflammatory biomarkers and, consequently, greater inflammation and greater edema [11, 12].

In the literature, a positive correlation between BMI values and developed edema is observed. Thus, individuals with higher BMI develop greater edema, but their rate of reduction is faster in the first postoperative days [10, 11]. However, individuals with lower BMI develop smaller edema, and although the rate of reduction in the first postoperative days is slower, the total resolution of edema occurs before than in people with a high BMI [10].

#### **2.3 Operative time**

The duration of surgery is appointed as one of the predictive factors for a greater or smaller postoperative edema. This is because a longer surgery requires a greater manipulation of the tissues, and consequently, a greater inflammatory process [4, 13–16].

The increasing of the surgical time can occur due to factors related to the surgery and intrinsic to the patient, such as age and anatomical variations. In addition of it, the surgeon's experience is related to the increasing or decreasing of the operative time [13, 15, 17].

The operative time is predictive not only for the amount of edema, but also to the intensity of pain and trismus. This is due to a bigger trauma or intraoperative complications, which is directly related to the increase in surgical time [4, 14, 16]. Thus, although studies indicate that there is a correlation between high surgical time and greater postoperative edema, factors that caused an increasing of the surgical time must be considered.

#### **2.4 Type of surgery and surgical trauma**

The type of surgery performed interferes directly in postoperative edema. Thus, large surgery (such as orthognathic surgery) is expected to cause a greater inflammatory process and, therefore, larger and more diffuse edema than minor surgery (third molar extraction, for example) [14, 18].

However, when it comes to the same type of surgery, variations can occur depending on the surgical difficulty level. It is expected that a major difficulty surgery occurs in a longer surgical time and causes a more intense and extensive surgical trauma. Therefore, the inflammatory process will be bigger, as well as the postoperative edema [4, 11, 14, 16, 19].

Some factors can contribute to the increasing of the surgery difficulty level, such as denser bones, teeth with roots formed and consolidated in the bone by masticatory stimuli, quantity of procedures, and unfavorable dental position [9, 11, 14].

The position of the third lower molar closer to the lingual wall appears to result in more severe postoperative edema, due to a more extensive surgical trauma in consequence of the bone amount removed [11]. In addition, the distal and horizontal position of the teeth is related to the greater postoperative edema, as the need to perform osteotomy and odontosection, which results in a greater surgical trauma [14].

In large surgeries such as orthognathic surgery, factors like the duration of the surgery, combined procedures (maxillary and mandibular osteotomy and mentoplasty) and bone density are related to the amount of postoperative edema. Thus, surgeries in only one of the jaws present less surgical trauma than the bimaxillaries, and therefore, develop smaller edema. When it comes to bone density, thicker and denser bones cause more difficulty in the osteotomies, increasing surgical trauma and inflammatory process [18, 20].

Surgeries involving maxilla, such as Le Fort I osteotomy, result in greater internal edema to the cavities, increasing the risk of airways obstruction [18].

#### **2.5 Surgeon's experience**

It is very difficult to evaluate the experience of one surgeon, since there are no preestablished protocols to separate experienced surgeons from inexperienced. Some papers use the classification based on the training phase in which the surgeon is, others how long the surgeon is graduated, or even the amount of surgeries already performed by the professional [14, 17, 21, 22].

Surgeon's experience indirectly interferes with postoperative edema. This is because it does not directly affect the factors that converge to the edema formation, but rather those that are related to the severity of the postoperative edema [14, 17, 21, 22].

The greater the experience of the surgeon, the lower is the occurrence of postoperative complications. In addition, the more experienced surgeon is capable to solve more quickly and efficiently intraoperative complications, as well as perform the surgical procedure accurately. And more, the surgeon's experience is closely related to possible planning errors (such as implant and orthognathic surgeries) and execution. Less experienced surgeons are more likely to make these mistakes, culminating in the prolongation in the surgery duration and even possible the need for surgical reintervention [17, 21, 22].

Therefore, the surgeon's experience interferes in the surgical time, trauma extension, and blood loss, which are decisive factors for the inflammatory process and, consequently, for postoperative edema [14, 21].

#### **2.6 Blood loss**

Although there are no studies relating the amount of transoperative bleeding to edema, it is known that there is a relation between blood loss and postoperative quality.

Lymphedema is characterized by the increasing volume of a body segment. However, this swelling is not always present only by edema, especially in the postoperative cases. Hematomas and clots also cause enlargement of the region volume. That way, trans- and postoperative bleeding contributes to swelling, as there is an increase in the body segment volume but, unfortunately, it is not possible to clearly distinguish whether it is edema, hematomas, or the combination of them [23].

Besides that, the amount of blood lost during surgery influences the inflammatory process. The greater the bleeding, the more intense and lasting is the inflammatory process, and the greater is the postoperative edema [18, 20].

Due the fact that these surgeries are performed in oral cavity, there is a possibility of swallowing blood during the surgical procedure. Besides the malaise caused by blood loss, postoperative vomiting increases the pressure in the newly operated region and causes an increasing of the edema. In addition, due to the bleeding caused by the pressure increasing, there may be formation and/or increasing of hematomas [24–26].

Therefore, strategies are necessary in order to reduce the amount of bleeding and, consequently, not only to improve postoperative quality, but also to help control facial edema and reduce the period of hospitalization after oral and maxillofacial surgeries.

#### **2.7 Induced hypotension**

The mean arterial pressure interferes directly in the bleeding and, thus in surgical time. Lower mean blood pressure reduces transoperative bleeding, reducing as well the amount of blood lost, improving the visualization of the surgical field, reducing surgical time, and the formation of hematomas and swelling [27–30].

*Edema Management in Oral and Maxillofacial Surgery DOI: http://dx.doi.org/10.5772/intechopen.80971*

The hypotension induced during surgery is a strategy to improve the surgical field through the reduction of bleeding and consequently reducing surgical time and postoperative inflammatory process [24, 27, 28, 31]. Induced hypotension, or controlled hypotension, is defined by the reduction of systolic blood pressure to 80–90 mmHg with a reduction in mean arterial pressure (MAP) to 50–65 mmHg or a 30% reduction in MAP [30, 32]. It is obtained through medicament during anesthesia. Despite being considered safe and presenting proven benefits, induced hypotension requires preparation and good skill of the anesthesiologist and should not be maintained for long time due to hypoperfusion risks of the organs such as the central nervous system (CNS), heart, liver, and kidneys [29, 32].

Although in the current literature have not yet been found studies that have investigated the correlation between hypotension induced in face surgeries and postoperative edema, hypotension is capable to improve several factors involved with the development and amount of edema.

#### **2.8 Age**

The age at which the patient is operated has been pointed out as one of the predictive factors for the development of bigger or smaller edema. Despite studies attempt to find this relation, there is still no consensus on the relation between age and severity of developed edema [4, 14, 15, 33, 34].

On the one hand, some authors argue that face surgery in younger individuals results in less difficulty in the procedure and consequently less surgical trauma and less edema [11]. On the other hand, there are authors who affirm that the reduction of the inflammatory response and diminution of the lymphatic system elasticity occur with the increase in the age. Thus, older individuals develop less edema and have less efficacy of the lymphatic system [4, 14–16, 33–35]. Besides that, older patients have a prolonged inflammatory process and, therefore, slower reduction of edema [15].

#### **2.9 Gender**

Another factor pointed as an influencer in the formation of edema and its quantity is gender. Although is expected that women develop greater swelling due to hormonal variations, use of oral contraceptives, and bigger risk for dry socket, the male gender is pointed out in studies as being more predisposed to a greater amount of postoperative edema [11, 13, 34].

Factors such as increased bone density and thickness and stronger muscles can do postoperative edema to be more severe in men than in women. This is because they are factors that directly interfere in the level of difficulty and quantity of surgical trauma, injuring more lymphatic structures and increasing the inflammatory response, generating more edema [11, 34].

However, the smaller thickness of the female mandible increases the chances of fracture of the mandibular ramus during third molar extraction, increasing surgical trauma [4, 13].

Anyway, this significant difference in the amount of developed edema is observed on the first postoperative day, but it is irrelevant on the seventh day [11, 34].

Thus, even in studies in which the amount of postoperative edema does not present a significant difference between the genders, the extent of surgical trauma and the occurrence of intraoperative complications are indicated as the main influential factors for the severity of postoperative edema [10, 19]. However, the occurrence and intensity of these factors are difficult to predict, so that is the reason to consider the gender and its risk factors and predict the level of the surgery difficulty.

#### **2.10 Vomiting**

The presence of nausea followed or not by vomiting is a factor that can be observed in clinical practice. Increased patient effort during vomiting increases facial edema and also stimulates postoperative bleeding. However, although the relation between nausea and vomiting with edema is not mentioned in the literature, it is a fact that can be verified in clinical practice, especially in the postoperative period of orthognathic surgery.

#### **2.11 Postoperative rest**

Another important factor related to the control or prevention of edema formation consists on the postoperative rest and positioning of the patient. It is known that the dorsal decubitus tilted by approximately 30° decreases the pressure in the face blood vessels and helps to control the bleeding and edema.

After surgery, the periosteum is detached in the operated region. Thus, the mobilization of this periosteum, by movement or compression of this region, stimulates the inflammatory response potentiating the edema.

Although these factors are not in specific scientific studies, clinical observation makes it possible to affirm the importance of both bedside and resting care in the postoperative period of face surgeries.

#### **3. Forms of evaluation and edema measurement**

In maxillofacial surgery, the observation, control, and reduction of edema are important postoperative factors, due to the possibility of airway compromise. In this way, surgeries with potential formation of exacerbated edema should present evaluation and control of this condition, in order to assist the decision related to the maintenance or replacement of the edema treatment protocol.

Between the techniques described for evaluating edema, the most used ones clinically are subjective, and are totally dependent on the professional's experience and on the patient's report. Although there are more objective methods of clinical evaluation with good reproducibility, these are limited to the upper and lower limbs, making it impossible to apply to regions such as head and neck [35, 36].

In the head and neck regions, most of the methods reported in the literature measure the edema by the distance between two points, based on anatomical points, such as mandibular angle, lateral, and medial epicanto of the eyes and middle of the chin.

Other measurement devices that provide more accurate data about the changes related to edema values include imaging exams. However, due to the fact that it involves high cost and exposes the patient to ionizing radiation, these techniques need specific indication [37]. Ultrasonography (US), magnetic resonance imaging, and computed tomography are examples of usable exams [38]. The US presents changes in the echogenicity of its images, which are not specific for volume changes caused by increasing of subcutaneous fluids [39]. In addition, to the face part, the echographic measurement does not always point the more swollen site due to the reproduction of the distances from the skin to the bone, which leads to imprecise and disproportionate results [40].

Bioelectrical impedance is another method described in the literature for the measurement of edema. This technique measures the amount of peripheral and total fluid in the body. However, low-cost and easily applicable devices for measuring body edemas as well as limbs are still scarce [36].

*Edema Management in Oral and Maxillofacial Surgery DOI: http://dx.doi.org/10.5772/intechopen.80971*

The evaluation methods developed for use in researches have evolved greatly. The first studies used subjective methods and difficult reproducibility, which made them less reliable in relation to the real magnitudes and behavior of edema. Van Gool et al. and Album et al. demonstrated the lack of correlation between subjective evaluations and objective measures of edema [41–43]

The measurement methods should be capable of being used in clinical and patient tolerable trials. Thus, portable devices were studied with the objective that they could be easily used with precision and transported to the place where the patient is, making possible to obtain early measures and follow-up of the edema [42, 44].

Therefore, objective measurement methods represent a more appropriate approach to the problem. However, these measurements should be evaluated and validated by doing repeated measurements on untreated individuals to verify its accuracy.

The methods already tested and used in studies were [45, 46]:


#### **4. Complications related to the postoperative edema**

The early stage of inflammation presents accumulation of fibrin and polymorphonuclear neutrophils in the extracellular space of injured tissues. The processes that occur in this phase are vessel diameter change, increased vascular permeability, exudate formation and migration of neutrophil cellular exudates into the extravascular space. The chemical mediators of acute inflammation include histamine, prostaglandins, leukotrienes, serotonin, and various cytokines. It is known that prostaglandin associated to bradykinin has the most potent pain-activating effect [14, 47, 48].

The control of inflammation and, therefore, swelling aims to reduce pain and improve life quality in the postoperative period. The processes of the inflammatory mediator may last up to 96 hours.

Trismus occurs as a result of muscle spasm caused by the inflammatory process. In this process, there is compression of the nervous structures by the edema, leading to the limitation of movement accompanied by a painful sensation, which can be from discomfort to severe pain [14, 47, 49].

Although it is subjective and dependent on several factors, the evaluation of postoperative pain in maxillofacial surgeries is essential, since this is one of the main complaints of operated patients and is directly related to edema. Therefore,

#### *Inflammation in the 21st Century*

pain, edema and trismus are consequences of the formation and release of prostaglandins, bradykinins, and other mediators of inflammatory response [14, 47].

Patients with moderate and severe edema may be unable to discern pain from discomfort caused by stretching of the skin by increased facial volume. In addition, the pain is related to the patient's emotional state, being influenced directly by their mood, level of satisfaction, and well-being [18, 20, 50].

Therefore, edema can also cause psychological and emotional problems due to the esthetic alteration of the affected body segment [50]. The maxillofacial surgeries carry great esthetic and functional expectations. However, patients, although relieved to have undergone surgery, may present mood swings due to the difficulty of self-care, pain, and edema. Changes in body image are one of the major complaints related to edema [20].

Edema can also influence self-care. This is because it makes feeding and oral hygiene difficult because it prevents proper visualization of the oral cavity and limits the range of mandibular movement. In addition, patients submitted to orthognathic surgery have shown greater difficulty in removing and placing intermaxillary locking elastics according to the degree of edema they develop [20].

Internal edema to the cavities is a major concern in the postoperative period. This is because breathing may be affected by pressure and possible obstruction of upper airway structures, causing respiratory distress and discomfort, and even leading to the need for re-intubation or performing a tracheostomy in the most serious, life-threatening cases [6, 8].

Severe postoperative edema is an important complication that can affect upper airway permeability and may lead to obstruction in more severe cases. The procedure that presents the greatest risk of airway obstruction due to edema is the Le Fort I type osteotomy, performed in the maxilla and covering the floor of the nasal fossa [18]. Thus, severe edema can cause respiratory and functional problems, which increases hospitalization time and the need for ICU admission.

Peripheral nerve damage is the result of direct or indirect trauma to a nerve. The direct relationship between edema and paresthesia is known and can be explained by the spatial relationship of the nerve vessels with adjacent structures, such as muscles and bones.

Following the same mechanism of acute compressive neuropathies, facial edema caused by surgical trauma, infections, fractures, or injuries can compress the sensory and motor nerves of the face (trigeminal nerve and facial nerve). This compression, or even stretching of these nerve bundles, impairs the conduction of the nerve impulse, resulting in paresthesia and even temporary paralysis.

Studies on nerve conduction measured the magnitude of the conduction blockade of nerve action potentials and the focal slowing of conduction. Direct correlation between degree of changes and duration of compression was demonstrated. Another observation is related to local ischemia, which, in combination with direct pressure effects, contributes to the development of compressive neuropathies. In severe cases of acute compression, with direct relation to extensive and prolonged edema, remyelination of nerve fibers can take weeks or months after resolution of compression.

Another aspect in relation to the neurosensorial disorders is related to the inflammatory mediators that are released when a trauma to the tissue occurs. These are located in the edema region and act temporarily as chemical irritants to the nerves.

Thus, studies attempt to relate the use of corticosteroids with the improvement of neurosensory symptoms after tissue trauma with considerable edema. However, due to the lack of standardization of the applied tests and classification, only the presence or absence of the disorder was considered [51]. More controlled clinical trials need to be performed to obtain data on neurosensory disorders.

#### *Edema Management in Oral and Maxillofacial Surgery DOI: http://dx.doi.org/10.5772/intechopen.80971*

Some local factors (directly related to the wound) and systemic (linked to the individual) can interfere in the cicatricial process, facilitating complications and sequels and causing esthetic and functional damages to the tissue.

Local factors: dimension and depth of the lesion; level of contamination; presence of net collections (bruises, ecchymosis, edema); tissue necrosis and local infection; poor vascular supply; surgical technique used, material and technique of suture, types of bandages; and traction or mechanical pressure on the scar [52–54].

Systemic factors: age group, ethnic origin, nutritional status, presence of chronic diseases, and use of medicines.

Angiogenesis is essential to healing wounds as it provides restoration of blood flow and transport of nutrients to cells as well as transporting the components of the immune system. Edema makes this stage difficult, because the excessive distension of the tissues leads to compression of the newly formed vessels, altering the blood flow. In this way, the body's capability to carry defense cells and administered antibiotics is impaired, making healing more difficult.

Hypoxia in the area of the lesion stimulates angiogenesis responsively, aiming formation and remodeling of the extracellular matrix for tissue repair. However, this process is limited to the first 48 hours of the beginning of the repair process, being detrimental to vascular neoformation and regulation of healing factors.

Fibroblasts are involved in deposition of the extracellular matrix and also in approaching the edges of the wound. Thus, the tissue distension caused by edema compromises this narrowing and tissue reepithelialization, making it difficult to form the fibrin network and providing a disordered growth of collagen, which leads to the formation of hypertrophic scars [53].

With excessive edema, a lesion that could have first-intention healing with contact between the edges becomes second intention, due to tissue tension, causing dehiscences of suture and separation of the wound edges. In addition, local edema obstructs the lymphatic vessels, facilitating the accumulation of catabolites and producing a greater level of inflammation.

#### **5. Medications used for edema control**

#### **5.1 Corticoids**

Inflammation is the local physiological response to tissue injury. Although some amount of inflammation is needed for proper wound healing, the excess of inflammation leads to severe edema and pain that causes discomfort to the patient.

The use of corticosteroids during orthognathic surgery is a fairly common practice for faster resolution of facial edema [55]. However, there is no consensus on its uses, its benefits, and adverse effects. The comparison of drugs in published studies is difficult due to the variety of parameters and methods used. Corticosteroids help reduce facial edema by acting as immunosuppressants that block the early and late stages of inflammation, decreasing the dilation and permeability of blood vessels. From this, there is a reduction of the amounts of liquid, proteins, macrophages, and other inflammatory cells present in the areas of tissue injury. In this terms, corticoids have a beneficial effect on the inflammation control, and consequently, on edema [51].

The use of steroids in patients can be by mouth, intramuscular injection, or intravenous methods. A recent study compared the effects of different routes of methylprednisolone uses on edema and trismus after extraction of third molars [56]. It was concluded that the systemic application of a steroid is more effective for improving the range of motion. However, direct injection of the steroid into the musculature had the best effect in reducing postoperative swelling.

Another study by Ehsan et al. [57] analyzed the effect of preoperative submucosal uses of dexamethasone on swelling and trismus on third molar extraction. They found out that this injection was very effective in reducing these postoperative conditions. In another study, it was found that the uses of corticosteroids in the preoperative period through the parenteral route have a greater impact in the reduction of postoperative swelling and trismus [58]. In addition, patients with zygomatic bone fractures usually present swelling, pain, and trismus before surgery, requiring prolonged treatment than removal of the third molars. Therefore, in order to benefit from steroid medication, patients with facial fractures should receive higher doses than patients undergoing minor surgeries [45].

The use of intravenous systemic corticosteroids before orthognathic surgery helps to reduce facial edema, but adverse effects are not well described in literature [59]. The use of corticosteroids before, during, and after orthognathic surgery, independently of the dosages, promotes reduction in facial edema, mainly until the third postoperative day. The most commonly used corticosteroids are dexamethasone, methylprednisolone, and betamethasone [51, 60]. Betamethasone is considered a potent steroid because it has high anti-inflammatory activity and does not cause fluid retention [60]. Dexamethasone is a highly selective and long-acting synthetic corticosteroid that has potent anti-inflammatory action [61].

In oral surgery, of all pharmacological agents tested, steroids seem to be the most successful for inflammation control. Corticosteroids, such as dexamethasone, may inhibit the early stage of the inflammatory process and have been widely used in different regimens and pathways to decrease inflammatory process after third molar surgery [62].

Although steroids seem to be the most successful in relieving edema after extraction of the third molar, the immunosuppressive effects of cortisol and its synthetic analogues are well known [63]. Previous studies about dexamethasone in third molar surgeries have concluded the need of accurate clinical research for better evaluation protocols for corticosteroid use [64].

#### **5.2 Analgesics**

The use of analgesics and nonsteroidal anti-inflammatory drugs alone or in combination with corticosteroids or opioids is common after third molar surgeries to reduce facial edema and pain [65]. When nonsteroidal anti-inflammatory drugs are given prior to surgery, they significantly reduce postoperative edema [66]. One study compared the use of diclofenac potassium, etodolac, and naproxen sodium given in preoperative of third molar surgery and concluded that diclofenac potassium showed better edema reduction [67]. Another study compared the use of diclofenac potassium alone or in combination with dexamethasone and concluded that combined therapy was more effective in reducing pain, trismus, and edema after third molar surgery [68]. There is no consensus in literature about which analgesics to use, for how long, and what is the best dosage with the least adverse effects.

#### **5.3 Hyaluronic acid (HA)**

A new drug trend that has been used to control edema development is hyaluronic acid (HA). Nowadays, few studies are found in literature and their actual efficacy as well as their use is not well established yet. HA is a high molecular weight glycosaminoglycan, a major component of the extracellular matrix [69]. It can be found in several tissues, and one of its properties is formation induction of early

granulation tissue, which helps the healing and improves inflammatory process [70]. HA turned out to be effective in reducing edema when used as spray after third molar extraction [70, 71]. The use of HA associated with platelet-rich fibrin was capable to decrease edema after third molar extraction surgery, compared to the isolated use of platelet fibrin [72]. Further studies using HA in larger groups and in other types of surgeries are necessary to establish a protocol use, consensus on its effects, and investigation of possible adverse effects.

#### **5.4 Adverse effects of medications at the doses used**

The adverse effects of corticosteroids are rare but important to evaluate. Complications are well known and include immune system suppression, hypertension, hyperglycemia, suppression of adrenal corticosteroid activity, allergic reactions, skin steroid acne, glaucoma, and psychiatric disorders. In addition, the use over 7 days may lead to development of Cushing's syndrome [54, 73].

Thus, it is noted that complications are related to prolonged use. In maxillofacial surgeries, it is generally used for a short time, at most 24–48 hours, so side effects are rare.

Also, it is known that anti-inflammatory drugs for edema control may increase bleeding by directly interfering in coagulation cascade. Thus, its benefit regarding edema control is compromised.

#### **6. Most commonly used forms of edema control**

#### **6.1 Cryotherapy**

Cryotherapy is the therapeutic use of cold applied for reducing skin and subcutaneous tissues temperature. It is indicated for inflammation control, pain, and edema after surgery or injury [65, 74]. Thus, physiological cooling exerts autonomic-mediated effect that induces vasoconstriction, favoring minimization and control of edema [75].

It is a treatment modality widely used because it is simple, inexpensive, and can be applied many times. Its therapeutic effects are due to alterations in blood flow, consequent vasoconstriction, and reduction of metabolism, also providing restriction of bacterial growth.

However, information concerning cryotherapy effects on edema is controversial [74]. Few studies report the effects of cryotherapy in maxillofacial surgeries, although its use is consecrated by the great majority of surgeons and in several types of surgeries.

Considering that during the first 10 minutes of ice application, most of the local temperature reduction occurs, most studies recommend the application for 10–20 minutes, having a rest period of the same time or twice as long [74]. The use of cryotherapy for 30 minutes every 1½ hours, for 48 hours after third molar extraction was quite effective in facial edema control [76].

Cryotherapy is contraindicated for patients with peripheral vascular disease, hypersensitivity or cold intolerance, as in Raynaud's phenomenon and in areas with impaired circulation. A disadvantage of cryotherapy is that its use normally starts at 0° and rapidly reaches room temperature [75].

The cryotherapy protocols use differ greatly from each other, especially regarding duration and application form [74]. Its efficacy has been questioned because despite its common and daily use in clinical practice after maxillofacial surgeries, there is no consensus or protocols on its use, so new studies are needed.

#### **6.2 Hilotherapy**

Hilotherapy began to be used recently in postoperative of maxillofacial surgeries for control and reduction of facial edema. It is a preformed polyurethane face mask, in which cold and sterile water stream passes through, promoting cryotherapy at regulated and maintained temperatures [77].

A recent systematic review showed that hilotherapy is used immediately after surgery, with temperatures of 14–15°C. However, in third molar extraction, single application was used for 45 minutes, and after orthognathic surgeries, the application was for continuous period from 48 to 72 hours. Both protocols had positive effect in reducing facial edema [78]. Therefore, it can be concluded that extensive surgeries require longer application.

Hilotherapy, when compared to facial cryotherapy performed using ice blocks, was more efficient in facial edema control and reduction after maxillofacial surgeries [77–80].

A recent study has shown that the use of facial hilotherapy performed at home after third molar extraction surgery is safe, easy to apply, brings benefits in reducing facial edema and also improves quality of life [75].

One of the difficulties in using hilotherapy is the cost of the device, which can reach high values. However, once this is resolved, its use will probably replace conventional cryotherapy in a few years as studies have shown beneficial effects in reducing edema and postoperative pain with greater patient comfort.

#### **6.3 Laser**

Low-power laser is a relatively recent method and has been used as an alternative to edema control because it is capable of promoting modulation of the inflammatory response, reducing pain, edema and trismus, in addition to accelerate tissue repair [71, 81]. It is considered easy to apply and does not cause adverse effects [65].

Laser acts in reduction of edema by controlling and decreasing inflammatory response. So, it promotes faster recovery of injured lymphatic vessels and potentiates the action of lymph nodes [82].

Despite this, there is still no consensus about which is the best protocol for use in maxillofacial surgeries, so that its effects can be better utilized. However, different protocols can be found in literature, especially regarding to which postoperative moment laser should be applied and how many sessions are necessary. In laboratory tests, low-power laser was able to improve pain by regulating inflammatory factors at doses around 7.5 J/cm2 . In addition, application in an area using more than one point promotes better results than the concentrated application in a single point.

The need to control inflammation in preoperative period is known. However, using laser before third molar extraction surgery seems to have only analgesic response [83].

The laser can be applied in minor surgeries, such as dental extractions and also larger, such as orthognathic surgery. Although the application of intraoral and extraoral laser at the end of the surgery does not show benefits in the immediate reduction of edema, when evaluated in the following days, the patients present a reduction in facial edema [82–84]. That occurs due to the latency period in which there is the biomodulation caused by the laser on the inflammatory response, with prolonged and residual effect [83], not requiring more than one application [84].

Therefore, the use of laser is questioned in small and controlled inflammatory processes, since benefits to patient do not justify treatment costs [85, 86]. Still, in some cases, laser seems to have analgesic effect only, not helping to reduce facial edema [87].

Thus, although low-power laser has potential to control inflammatory process and reduce complications, results depend on an indication that justifies its use and, mainly, the protocol used.

#### **6.4 Manual lymphatic drainage (MDL)**

Manual lymphatic drainage is a resource that, if applied correctly and by a trained professional, helps in the resolution of edema. By means of slow movements and gentle pressure (30–40 mmHg) following the lymph pathway, the MLD proposes to potentiate the function of the lymphatic system [88, 89]. Thus, it is a nondrug option in the treatment of edema.

The benefit of manual lymphatic drainage is undeniable; however, in maxillofacial surgeries, it is still little used and little known, due to the scarcity of studies that demonstrate its effectiveness in this type of surgery and also prove the safety of its application. In surgeries in other regions of the body, the use of MLD to decrease edema is quite consistent, with well-established protocols and benefits. In maxillofacial surgery, there are still no protocols for beginning and no consensus regarding their benefits due to the amount of work done so far.

The MLD had proven efficacy in the postoperative period of third molar extraction, alveolar bone graft, and orthognathic surgery [90–92]. In a clinical trial with a split mouth model, third molar extraction was performed by adding MLD on one side only in the postoperative period. Using reproducible facial measures and Visual Analogue Scale (VAS) for pain, it was concluded that MLD is able to significantly reduce postoperative swelling and pain in this surgery [93].

The same effect was observed in the postoperative period of alveolar bone graft with filling of the bone defect by spongy bone of the iliac crest. However, this study compared the MLD performed by a physiotherapist to an adapted drain that was taught and applied by the patient. Both groups showed improvement over the course of the day, but MLD applied by physiotherapist had better results on edema and pain compared to self-drainage [92]. Despite that, attention should be paid to the absence of a control group so that the study would effectively prove the benefits of MLD. However, it is possible to conclude the importance of the physical therapist in the postoperative period of this surgery, since this professional has skills that can contribute to the improvement of the discomfort caused by the edema and the referred pain.

In the orthognathic surgeries, MLD was very effective in reducing postoperative edema when compared to a placebo, both applied by a physiotherapist. In these cases, not only was drainage capable to accelerate the regression process of edema, but also to anticipate its peak. It was also observed that the maximum edema was lower in the patients who received the MLD. Thus, MLD is able to promote the control of edema when applied during its development period and also to accelerate the process of regression of swelling in the postoperative period [91, 94].

However, even in this study, MLD was not effective in relation to pain perception. The authors attribute this to two factors: the application of a placebo, which may have interfered in patients' perception of pain and the fact that the patients did not develop severe edema, and therefore, the pain or discomfort related to the edema may have been lower, as well as the perception of relief in the group that received the MLD [91].

Although the benefits of MLD in the postoperative period of oral and maxillofacial surgeries have been studied, there is still no agreement as to when the application of MLD should begin. However, it is known that the peak of edema in maxillofacial surgeries occurs between 48 and 72 hours after surgery, and therefore, the beginning of MLD before this period seems to anticipate the peak of edema and regression, causing the amount of edema at the peak being lower [91, 94].

It can be concluded that MLD represents a safe nondrug option in the treatment of postoperative edema, when well indicated and applied by a qualified professional. Despite all the proven benefits, it is necessary to observe the need for MLD

in various oral and maxillofacial surgeries. It is known that it is able to accelerate the process of regression of edema and provide relief of pain, but the need should be questioned in cases of small surgeries with the formation of discrete and local edema. In those cases, typical of a small controlled inflammatory process, MLD can be an unnecessary treatment to the patient, increasing the costs of the treatment and not having all its benefits observed.

#### **6.5 Kinesio taping (KT)**

Elastic bandage, or Kinesio taping, was first used in athletes, to aid in the recovery of muscle injuries, provide more stability to the joints, and provide relief from pain. However, it was realized that due to its way of functioning, it could be beneficial in the treatment of lymphedema.

KT, through the formation of convolutions in the skin, increases the interstitial space. Thus, through this increased space, fluids tend to move from higher pressure areas (congesta) to areas of lower pressure, improving blood and lymphatic flow. This occurs following the placement of the KT, which is positioned according to the path of the lymphatic system. In that way, KT may be able to relieve swelling caused by bruising and edema [23, 45, 95, 96].

In maxillofacial surgeries, its efficacy has already been tested in several surgeries: surgical reduction of mandible fracture, surgery to reduce fractures of the zygomatic-orbital complex, third molar extraction, and orthognathic surgery [97].

In the surgical reduction of mandibular fracture and zygomatic-orbital complex, KT is effective in reducing edema, anticipating the day of peak edema, the amount of edema formed on this day, and accelerating its reduction. However, despite the more rapid resolution of edema, no effects on trismus or pain relief were found [95].

In third molar extraction surgeries, KT anticipates the day of maximum edema and the amount of edema formed on this day. However, the rate of edema reduction is lower when compared to patients who did not use KT. Despite that, patients who use KT postoperatively seem to have resolution of the edema earlier. Furthermore, KT was effective in relieving pain, but not in trismus [96].

Even so, in the exodontia, when compared to the placement of drains for the treatment of lymphedema, KT is not as effective. Drain placement at the surgical site is shown to be much more effective not only at the faster reduction of edema but also in relation to pain, although it is an invasive approach. Despite this, none of the treatments helped reduce trismus in this study. It should also be considered that drainage placement, despite being effective in reducing edema, may lead to other complications, in relation to the possibility of subcutaneous emphysema, infection, and external facial scar [98].

In orthognathic surgeries, the application of KT is beneficial in the treatment of postoperative edema, being capable to anticipate the day of maximum edema, reduce the maximum amount of edema formed, and accelerate the regression process of edema. However, it does not appear to have significant effects with regard to pain or trismus [97].

Thus, KT is a nonmedicated treatment option for the control and treatment of postoperative lymphedema of maxillofacial surgeries. However, its effects on pain and trismus need to be better elucidated. Although one of the goals of KT is to prevent the formation of bruises and/or to treat them, there is still no proof of it. Therefore, it is a function to be explored with great interest, since the increase in volume of a body segment is not only due to edema but also due to hematomas.

Therefore, KT is a relatively inexpensive treatment option, but it requires specific training and professional habilitation, as well as presurgery testing to check for allergy to the components of the bandage.

#### **7. Conclusions**

In this chapter, factors related to edema development in maxillofacial surgeries and alternatives for its control and treatment were presented. It is known that this condition is strictly related to the inflammatory process, and therefore, controlling edema also requires controlling postoperative inflammation.

Several factors contribute to edema severity, and knowing which factors cause these and their influence on inflammatory process, it is possible to predict the quality of the postoperative period. The inflammatory process control, and consequently edema restriction, is fundamental for the quality of healing process and postoperative. Thus, it is necessary to have attention and intervention of surgical team on controllable factors that lead to a most severe or mild formation of edema, such as surgical time and precise surgical planning.

In addition, knowing about the risks for each factor related to the edema development makes individual and personalized treatment possible, which brings great benefits to the patient. Aiming at reducing complications related to edema, better postoperative quality, increased satisfaction and reduction of hospitalization time and treatment costs, and several drug and nondrug methods may be employed. Currently, there is a tendency in reducing medicament use in order to reduce the occurrence and severity of adverse effects. In this way, nondrug methods are increasingly study targets and used in clinical practice.

Therefore, more studies are needed to prove the efficacy and safety of these methods. Also, the formation of a well-trained and integrated multiprofessional team is necessary, aiming for safety, comfort, and faster patient recovery in postoperative period of maxillofacial surgeries.

#### **Conflict of interest**

The authors declare that they have no conflict of interest.

#### **Author details**

Renato Yassutaka Faria Yaedu1 \*, Marina de Almeida Barbosa Mello2 , Juliana Specian Zabotini da Silveira2 and Ana Carolina Bonetti Valente2

1 Hospital for Rehabilitation of Craniofacial Anomalies, University of São Paulo and Bauru School of Dentistry, Bauru, São Paulo, Brazil

2 Hospital for Rehabilitation of Craniofacial Anomalies, University of São Paulo, Bauru, São Paulo, Brazil

\*Address all correspondence to: yaedu@usp.br

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

### **References**

[1] Ebert JR, Joss B, Jardine B, Wood DJ. Randomized trial investigating the efficacy of manual lymphatic drainage to improve early outcome after total knee arthroplasty. Archives of Physical Medicine and Rehabilitation. 2013;**94**(11):2103-2111

[2] Korpan MI, Crevenna R, Fialka-Moser V. Lymphedema: A therapeutic approach in the treatment and rehabilitation of cancer patients. American Journal of Physical Medicine & Rehabilitation. 2011;**90**(5 Suppl 1): S69-S75

[3] Smith BG, Lewin JS. Lymphedema management in head and neck cancer. Current Opinion in Otolaryngology & Head and Neck Surgery. 2010;**18**(3):153-158

[4] de Santana-Santos T, de Souza-Santos A-A-S, Martins-Filho P-R-S, da Silva L-C-F, De Oliveira E Silva E-D, Gomes A-C-A. Prediction of postoperative facial swelling, pain and trismus following third molar surgery based on preoperative variables. Medicina Oral, Patología Oral y Cirugía Bucal. 2013;**18**(1):e65-e70

[5] Bork K, Barnstedt S-E. Laryngeal edema and death from asphyxiation after tooth extraction in four patients with hereditary angioedema. Journal of the American Dental Association (1939). 2003;**134**(8):1088-1094

[6] Cifuentes J, Palisson F, Valladares S, Jerez D. Life-threatening complications following orthognathic surgery in a patient with undiagnosed hereditary angioedema. Journal of Oral and Maxillofacial Surgery. 2013;**71**(4):e185-e188

[7] Morcavallo PS, Leonida A, Rossi G, Mingardi M, Martini M, Monguzzi R, et al. Hereditary angioedema in oral surgery: Overview of the clinical

picture and report of a case. Journal of Oral and Maxillofacial Surgery. 2010;**68**(9):2307-2311

[8] Puricelli E, Ponzoni D, Artuzi FE, Martins GL, Calcagnotto T. Clinical management of angioneurotic oedema patient post-orthognathic surgery. International Journal of Oral and Maxillofacial Surgery. 2011;**40**(1):106-109

[9] Carvalho RWF, do Egito Vasconcelos BC. Assessment of factors associated with surgical difficulty during removal of impacted lower third molars. Journal of Oral and Maxillofacial Surgery. 2011;**69**(11):2714-2721

[10] van der Vlis M, Dentino KM, Vervloet B, Padwa BL. Postoperative swelling after orthognathic surgery: A prospective volumetric analysis. Journal of Oral and Maxillofacial Surgery. 2014;**72**(11):2241-2247

[11] Pérez-González JM, Esparza-Villalpando V, Martínez-Rider R, Noyola-Frías MÁ, Pozos-Guillén A. Clinical and radiographic characteristics as predictive factors of swelling and trismus after mandibular third molar surgery: A longitudinal approach. Pain Research & Management. 2018;**2018**:7938492

[12] Kantor ED, Lampe JW, Kratz M, White E. Lifestyle factors and inflammation: Associations by body mass index. PLoS One. 2013;**8**(7):e67833

[13] Benediktsdóttir IS, Wenzel A, Petersen JK, Hintze H. Mandibular third molar removal: Risk indicators for extended operation time, postoperative pain, and complications. Oral Surgery, Oral Medicine, Oral Pathology, Oral Radiology, and Endodontics. 2004;**97**(4):438-446

*Edema Management in Oral and Maxillofacial Surgery DOI: http://dx.doi.org/10.5772/intechopen.80971*

[14] Bello SA, Adeyemo WL, Bamgbose BO, Obi EV, Adeyinka AA. Effect of age, impaction types and operative time on inflammatory tissue reactions following lower third molar surgery. Head & Face Medicine. 2011;**7**:8

[15] Pogrel MA. What is the effect of timing of removal on the incidence and severity of complications? Journal of Oral and Maxillofacial Surgery. 2012;**70**(9):S37-S40

[16] Mobilio N, Vecchiatini R, Vasquez M, Calura G, Catapano S. Effect of flap design and duration of surgery on acute postoperative symptoms and signs after extraction of lower third molars: A randomized prospective study. Journal of Dental Research, Dental Clinics, Dental Prospects. 2017;**11**(3):156-160

[17] Blondeau F, Daniel NG. Extraction of impacted mandibular third molars: Postoperative complications and their risk factors. Journal of the Canadian Dental Association. 2007;**73**(4):325

[18] Khattak ZG, Benington PCM, Khambay BS, Green L, Walker F, Ayoub AF. An assessment of the quality of care provided to orthognathic surgery patients through a multidisciplinary clinic. Journal of Cranio-Maxillo-Facial Surgery. 2012;**40**(3):243-247

[19] Osunde OD, Saheeb BD. Effect of age, sex and level of surgical difficulty on inflammatory complications after third molar surgery. Journal of Oral and Maxillofacial Surgery. 2015;**14**(1):7-12

[20] Robinson RC, Holm RL. Orthognathic surgery for patients with maxillofacial deformities. AORN Journal. 2010;**92**(1):28-49; quiz 50-2

[21] Jerjes W, El-Maaytah M, Swinson B, Banu B, Upile T, D'Sa S, et al. Experience versus complication rate in third molar surgery. Head & Face Medicine. 2006;**2**:14

[22] Cassetta M, Bellardini M. How much does experience in guided implant surgery play a role in accuracy? A randomized controlled pilot study. International Journal of Oral and Maxillofacial Surgery. 2017;**46**(7):922-930

[23] Ristow O, Pautke C, Kehl V, Koerdt S, Hahnefeld L, Hohlweg-Majert B. Kinesiologic taping reduces morbidity after oral and maxillofacial surgery: A pooled analysis. Physiotherapy Theory and Practice. 2014;**30**(6):390-398

[24] Lin S, Chen C, Yao C-F, Chen Y-A, Chen Y-R. Comparison of different hypotensive anaesthesia techniques in orthognathic surgery with regard to intraoperative blood loss, quality of the surgical field, and postoperative nausea and vomiting. International Journal of Oral and Maxillofacial Surgery. 2016;**45**(12):1526-1530

[25] Silva AC, O'Ryan F, Poor DB. Postoperative nausea and vomiting (PONV) after orthognathic surgery: A retrospective study and literature review. Journal of Oral and Maxillofacial Surgery. 2006;**64**(9):1385-1397

[26] Thomas JS, Maple IK, Norcross W, Muckler VC. Preoperative risk assessment to guide prophylaxis and reduce the incidence of postoperative nausea and vomiting. Journal of Perianesthesia Nursing. 23 Jun 2018. DOI: 10.1016/j.jopan.2018.02.007

[27] Yu CN, Chow TK, Kwan AS, Wong SL, Fung SC. Intra-operative blood loss and operating time in orthognathic surgery using induced hypotensive general anaesthesia: Prospective study. Hong Kong Medical Journal. 2000;**6**(3):307-311

[28] Praveen K, Narayanan V, Muthusekhar MR, Baig MF. Hypotensive anaesthesia and blood loss in orthognathic surgery: A clinical study. The British Journal of Oral & Maxillofacial Surgery. 2001;**39**(2):138-140

[29] Choi WS, Samman N. Risks and benefits of deliberate hypotension in anaesthesia: A systematic review. International Journal of Oral and Maxillofacial Surgery. 2008;**37**(8):687-703

[30] Prasant MC, Kar S, Rastogi S, Hada P, Ali FM, Mudhol A. Comparative study of blood loss, quality of surgical field and duration of surgery in maxillofacial cases with and without hypotensive anesthesia. Journal of International Oral Health. 2014;**6**(6):18-21

[31] Phillips C, Brookes CD, Rich J, Arbon J, Turvey TA. Postoperative nausea and vomiting following orthognathic surgery. International Journal of Oral and Maxillofacial Surgery. 2015;**44**(6):745-751

[32] Degoute C-S. Controlled hypotension. Drugs. 2007;**67**(7): 1053-1076

[33] Olmedo-Gaya MV, Vallecillo-Capilla M, Galvez-Mateos R. Relation of patient and surgical variables to postoperative pain and inflammation in the extraction of third molars. Medicina Oral. 2002;**7**(5):360-369

[34] Yuasa H, Sugiura M. Clinical postoperative findings after removal of impacted mandibular third molars: Prediction of postoperative facial swelling and pain based on preoperative variables. The British Journal of Oral & Maxillofacial Surgery. 2004;**42**(3):209-214

[35] Stanton AWB, Northfield JW, Holroyd B, Mortimer PS, Levick JR. Validation of an optoelectronic limb volumeter (Perometer®). Lymphology. 1997;**30**(2):77-97

[36] Nuutinen J, Ikäheimo R, Lahtinen T. Validation of a new dielectric device to assess changes of tissue water in skin and subcutaneous fat. Physiological Measurement. 2004;**25**(2):447-454

[37] Haaverstad R, Nilsen G, Rinck PA, Myhre HO. The use of MRI in the diagnosis of chronic lymphedema of the lower extremity. International Angiology. 1994;**13**(2):115-118

[38] Lahtinen T, Nuutinen J. A new device for clinical and radiobiological research to measure local development of edema induced by radiotherapy, drugs or surgery. Radiotherapy and Oncology. 2005;**76**:S47

[39] Gniadecka M, Quistorff B. Assessment of dermal water by highfrequency ultrasound: Comparative studies with nuclear magnetic resonance. The British Journal of Dermatology. 1996;**135**(2):218-224. Available from: https://onlinelibrary.wiley.com/doi/ abs/10.1111/j.1365-2133.1996.tb01150.x

[40] Piso DU, Eckardt A, Liebermann A, Gutenbrunner C, Schäfer P, Gehrke A. Early rehabilitation of head-neck edema after curative surgery for orofacial tumors. American Journal of Physical Medicine & Rehabilitation. 2001;**80**(4):261-269

[41] Van Gool AV, Ten Bosch JJ, Boering G. A photographic method of assessing swelling following third molar removal. International Journal of Oral Surgery. 1975;**4**(3):121-129

[42] Album B, Olsen I, Løkken P. Bilateral surgical removal of impacted mandibular third molar teeth as a model for drug evaluation: A test with oxyphenbutazone (Tanderil®). International Journal of Oral and Maxillofacial Surgery. 1977;**6**(3):177-189

[43] Holland CS. The development of a method of assessing swelling following *Edema Management in Oral and Maxillofacial Surgery DOI: http://dx.doi.org/10.5772/intechopen.80971*

third molar surgery. The British Journal of Oral Surgery. 1979;**17**(2):104-114

[44] Ågren E. High-speed or conventional dental engines for the removal of bone in oral surgery: I. A study of the reactions following removal of bilateral impacted lower third molars. Acta Odontologica Scandinavica. 1963;**21**(6):585-625

[45] Ristow O, Pautke C, Kehl V, Koerdt S, Schwärzler K, Hahnefeld L, et al. Influence of kinesiologic tape on postoperative swelling, pain and trismus after zygomatico-orbital fractures. Journal of Cranio-Maxillo-Facial Surgery. 2014;**42**(5):469-476

[46] Yamamoto S, Miyachi H, Fujii H, Ochiai S, Watanabe S, Shimozato K. Intuitive facial imaging method for evaluation of postoperative swelling: A combination of 3-dimensional computed tomography and laser surface scanning in orthognathic surgery. Journal of Oral and Maxillofacial Surgery. 2016;**74**(12):2506.e1-2506.e10

[47] Garcia AG, Sampedro FG, Rey JG. Trismus and pain after removal of impacted lower third molars. Journal of Oral and Maxillofacial Surgery. 1997;**55**(11):1223-1226. Available from: https://www.joms.org/article/ S0278-2391(97)90172-5/abstract

[48] Gunes N. A comparison of the effects of methylprednisolone and tenoxicam on pain, edema, and trismus after impacted lower third molar extraction. Medical Science Monitor. 2014;**20**:147-152

[49] Laureano Filho JR, Maurette PE, Allais M, Cotinho M, Fernandes C. Clinical comparative study of the effectiveness of two dosages of Dexamethasone to control postoperative swelling, trismus and pain after the surgical extraction of mandibular impacted third molars. Medicina

Oral, Patología Oral y Cirugía Bucal. 2008;**13**(2):E129-E132

[50] Ridner SH. The psychosocial impact of lymphedema. Lymphatic Research and Biology. 2009;**7**(2):109-112

[51] de Lima VN, Lemos CAA, Faverani LP, Santiago Júnior JF, Pellizzer EP. Effectiveness of corticoid administration in orthognathic surgery for edema and neurosensorial disturbance: A systematic literature review. Journal of Oral and Maxillofacial Surgery. 2017;**75**(7):1528.e1-1528.e8

[52] Deodhar AK, Rana RE. Surgical physiology of wound healing: A review. Journal of Postgraduate Medicine. 1997;**43**(2):52-56

[53] Steinbrech DS, Longaker MT, Mehrara BJ, Saadeh PB, Chin GS, Gerrets RP, et al. Fibroblast response to hypoxia: The relationship between angiogenesis and matrix regulation. The Journal of Surgical Research. 1999;**84**(2):127-133

[54] Lawrence WT, Diegelmann RF. Growth factors in wound healing. Clinics in Dermatology. 1994;**12**(1):157-169

[55] Chegini S, Dhariwal DK. Review of evidence for the use of steroids in orthognathic surgery. The British Journal of Oral & Maxillofacial Surgery. 2012;**50**(2):97-101

[56] Koçer G, Yuce E, Tuzuner Oncul A, Dereci O, Koskan O. Effect of the route of administration of methylprednisolone on oedema and trismus in impacted lower third molar surgery. International Journal of Oral and Maxillofacial Surgery. 2014;**43**(5):639-643

[57] Ehsan A, Ali Bukhari SG, Ashar AM, Manzoor A, Junaid M. Effects of pre-operative submucosal

dexamethasone injection on the postoperative swelling and trismus following surgical extraction of mandibular third molar. Journal of the College of Physicians and Surgeons– Pakistan. Jul 2014;**24**(7):489-492

[58] Herrera-Briones FJ, Prados Sánchez E, Reyes Botella C, Vallecillo CM. Update on the use of corticosteroids in third molar surgery: Systematic review of the literature. Oral Surgery, Oral Medicine, Oral Pathology, Oral Radiology. 2013;**116**(5):e342-e351

[59] Jean S, Dionne P-L, Bouchard C, Giasson L, Turgeon AF. Perioperative systemic corticosteroids in orthognathic surgery: A systematic review and meta-analysis. Journal of Oral and Maxillofacial Surgery. 2017;**75**(12):2638-2649

[60] Widar F, Kashani H, Alsén B, Dahlin C, Rasmusson L. The effects of steroids in preventing facial oedema, pain, and neurosensory disturbances after bilateral sagittal split osteotomy: A randomized controlled trial. International Journal of Oral and Maxillofacial Surgery. 2015;**44**(2):252-258

[61] Tripathi KD. Essentials of Medical Pharmacology. New Delhi, India: JP Medical Ltd; 2013. p. 1002

[62] ElHag M, Coghlan K, Christmas P, Harvey W, Harris M. The antiinflammatory effects of dexamethasone and therapeutic ultrasound in oral surgery. The British Journal of Oral & Maxillofacial Surgery. 1985;**23**(1):17-23

[63] Asimakopoulos G, Thompson R, Nourshargh S, Lidington EA, Mason JC, Ratnatunga CP, et al. An anti-inflammatory property of aprotinin detected at the level of leukocyte extravasation. The Journal of Thoracic and Cardiovascular Surgery. 2000;**120**(2):361-369

[64] Alexander RE, Throndson RR. A review of perioperative corticosteroid use in dentoalveolar surgery. Oral Surgery, Oral Medicine, Oral Pathology, Oral Radiology, and Endodontics. 2000;**90**(4):406-415

[65] Osunde OD, Adebola RA, Omeje UK. Management of inflammatory complications in third molar surgery: A review of the literature. African Health Sciences. 2011;**11**(3):530-537

[66] Zor ZF, Isik B, Cetiner S. Efficacy of preemptive lornoxicam on postoperative analgesia after surgical removal of mandibular third molars. Oral Surgery, Oral Medicine, Oral Pathology, Oral Radiology. 2014;**117**(1):27-31

[67] Akbulut N, Üstüner E, Atakan C, Çölok G. Comparison of the effect of naproxen, etodolac and diclofenac on postoperative sequels following third molar surgery: A randomised, double-blind, crossover study. Medicina Oral, Patología Oral y Cirugía Bucal. 2014;**19**(2):e149-e156

[68] Bamgbose BO, Akinwande JA, Adeyemo WL, Ladeinde AL, Arotiba GT, Ogunlewe MO. Effects of co-administered dexamethasone and diclofenac potassium on pain, swelling and trismus following third molar surgery. Head & Face Medicine. 2005;**1**:11

[69] Gocmen G, Gonul O, Oktay NS, Yarat A, Goker K. The antioxidant and anti-inflammatory efficiency of hyaluronic acid after third molar extraction. Journal of Cranio-Maxillo-Facial Surgery. 2015;**43**(7):1033-1037

[70] Koray M, Ofluoglu D, Onal EA, Ozgul M, Ersev H. Efficacy of hyaluronic acid spray on swelling, pain, and trismus after surgical extraction of impacted mandibular third molars. International Journal of Oral and Maxillofacial Surgery. 2014;**43**(11):1399-1403. Available from: *Edema Management in Oral and Maxillofacial Surgery DOI: http://dx.doi.org/10.5772/intechopen.80971*

https://www.sciencedirect.com/science/ article/pii/S0901502714001799

[71] Drew SJ. Best practices for management of pain, swelling, nausea, and vomiting in dentoalveolar surgery. Oral and Maxillofacial Surgery Clinics of North America. 2015;**27**(3):393-404

[72] Afat İM, Akdoğan ET, Gönül O. Effects of leukocyte- and plateletrich fibrin alone and combined with hyaluronic acid on pain, edema, and trismus after surgical extraction of impacted mandibular third molars. Journal of Oral and Maxillofacial Surgery. 2018;**76**(5):926-932

[73] Semper-Hogg W, Fuessinger MA, Dirlewanger TW, Cornelius CP, Metzger MC. The influence of dexamethasone on postoperative swelling and neurosensory disturbances after orthognathic surgery: A randomized controlled clinical trial. Head & Face Medicine. 2017;**13**(1):19

[74] Greenstein G. Therapeutic efficacy of cold therapy after intraoral surgical procedures: A literature review. Journal of Periodontology. 2007;**78**(5):790-800

[75] Beech AN, Haworth S, Knepil GJ. Effect of a domiciliary facial cooling system on generic quality of life after removal of mandibular third molars. The British Journal of Oral & Maxillofacial Surgery. 2018;**56**(4):315-321

[76] Laureano Filho JR, de Oliveira e Silva ED, Batista CI, FMV G. The influence of cryotherapy on reduction of swelling, pain and trismus after third-molar extraction: A preliminary study. Journal of the American Dental Association (1939). 2005;**136**(6):774- 778 quiz 807

[77] Moro A, Gasparini G, Marianetti TM, Boniello R, Cervelli D, Di Nardo F, et al. Hilotherm efficacy

in controlling postoperative facial edema in patients treated for maxillomandibular malformations. The Journal of Craniofacial Surgery. 2011;**22**(6):2114-2117

[78] Glass GE, Waterhouse N, Shakib K. Hilotherapy for the management of perioperative pain and swelling in facial surgery: A systematic review and meta-analysis. The British Journal of Oral & Maxillofacial Surgery. 2016;**54**(8):851-856

[79] Rana M, Gellrich NC, Joos U, Piffkó J, Kater W. 3D evaluation of postoperative swelling using two different cooling methods following orthognathic surgery: A randomised observer blind prospective pilot study. International Journal of Oral and Maxillofacial Surgery. 2011;**40**(7):690-696

[80] Veitz-Keenan A. Continuous cooling mask devices reduce patient discomfort and postoperative pain and swelling in patients undergoing orofacial surgery. Evidence-Based Dentistry. 2016;**17**(4):121-122

[81] Oliveira Sierra S, Melo Deana A, Agnelli Mesquita Ferrari R, Maia Albarello P, Kalil Bussadori S, Porta Santos Fernandes K. Effect of low-level laser therapy on the post-surgical inflammatory process after third molar removal: Study protocol for a doubleblind randomized controlled trial. Trials. 2013;**14**(1):373

[82] He WL, Yu FY, Li CJ, Pan J, Zhuang R, Duan PJ. A systematic review and meta-analysis on the efficacy of lowlevel laser therapy in the management of complication after mandibular third molar surgery. Lasers in Medical Science. 2015;**30**(6):1779-1788

[83] Petrini M, Ferrante M, Trentini P, Perfetti G, Spoto G. Effect of preoperatory low-level laser therapy on pain, swelling, and trismus associated with third-molar surgery. Medicina Oral, Patología Oral y Cirugía Bucal. 2017;**22**(4):e467-e472

[84] Koparal M, Ozcan KA. Effects of low-level laser therapy following surgical extraction of the lower third molar with objective measurement of swelling using a three-dimensional system. Experimental and Therapeutic Medicine. 2018;**15**(4):3820-3826. Available from: https://www. spandidos-publications.com/ etm/15/4/3820

[85] Brignardello-Petersen R, Carrasco-Labra A, Araya I, Yanine N, Beyene J, Shah PS. Is adjuvant laser therapy effective for preventing pain, swelling, and trismus after surgical removal of impacted mandibular third molars? A systematic review and meta-analysis. Journal of Oral and Maxillofacial Surgery. 2012;**70**(8):1789-1801

[86] Farhadi F, Eslami H, Majidi A, Fakhrzadeh V, Ghanizadeh M, KhademNeghad S. Evaluation of adjunctive effect of low-level laser therapy on pain, swelling and trismus after surgical removal of impacted lower third molar: A double blind randomized clinical trial. Laser Therapy. 2017;**26**(3):181-187

[87] Raiesian S, Khani M, Khiabani K, Hemmati E, Pouretezad M. Assessment of low-level laser therapy effects after extraction of impacted lower third molar surgery. Journal of Lasers in Medical Sciences. 2017;**8**(1):42-45

[88] Kasseroller RG. The Vodder school: The Vodder method. Cancer. 1998;**83**(12 Suppl American):2840-2842

[89] Rockson SG, Miller LT, Senie R, Brennan MJ, Casley-Smith JR, Földi E, et al. American Cancer Society Lymphedema Workshop. Workgroup III: Diagnosis and management of lymphedema. Cancer. 1998;**83**(S12B):2882-2885

[90] Szolnoky G, Mohos G, Dobozy A, Kemény L. Manual lymph drainage reduces trapdoor effect in subcutaneous island pedicle flaps. International Journal of Dermatology. 2006;**45**(12):1468-1470

[91] Yaedú RYF, Mello MAB, Tucunduva RA, JSZ d S, Takahashi MPMS, ACB V. Postoperative orthognathic surgery edema assessment with and without manual lymphatic drainage. The Journal of Craniofacial Surgery. 2017;**28**(7):1816-1820

[92] Ferreira T, Sabatella MZ, Silva T. Facial edema reduction after alveolar bone grafting surgery in cleft lip and palate patients: A new lymphatic drainage protocol. RGO - Rev Gaúcha Odontol. 2013. Available from: http:// www.revistargo.com.br/viewarticle. php?id=2893&

[93] Szolnoky G, Szendi-Horváth K, Seres L, Boda K, Kemény L. Manual lymph drainage efficiently reduces postoperative facial swelling and discomfort after removal of impacted third molars. Lymphology. 2007;**40**(3):138-142

[94] Modabber A, Rana M, Ghassemi A, Gerressen M, Gellrich N-C, Hölzle F, et al. Three-dimensional evaluation of postoperative swelling in treatment of zygomatic bone fractures using two different cooling therapy methods: A randomized, observer-blind, prospective study. Trials. 2013;**14**:238

[95] Ristow O, Hohlweg-Majert B, Kehl V, Koerdt S, Hahnefeld L, Pautke C. Does elastic therapeutic tape reduce postoperative swelling, pain, and trismus after open reduction and internal fixation of mandibular fractures? Journal of Oral and Maxillofacial Surgery. 2013;**71**(8):1387-1396

[96] Ristow O, Hohlweg-Majert B, Stürzenbaum SR, Kehl V, Koerdt S, *Edema Management in Oral and Maxillofacial Surgery DOI: http://dx.doi.org/10.5772/intechopen.80971*

Hahnefeld L, et al. Therapeutic elastic tape reduces morbidity after wisdom teeth removal—A clinical trial. Clinical Oral Investigations. 2014;**18**(4):1205-1212

[97] Tozzi U, Santagata M, Sellitto A, Tartaro GP. influence of kinesiologic tape on post-operative swelling after orthognathic surgery. Journal of Oral and Maxillofacial Surgery. 2016;**15**(1):52-58

[98] Genc A, Cakarer S, Yalcin BK, Kilic BB, Isler SC, Keskin C. A comparative study of surgical drain placement and the use of kinesiologic tape to reduce postoperative morbidity after third molar surgery. Clinical Oral Investigations. 19 Apr 2018. DOI: 10.1007/s00784-018-2442-x

### Section 3
