Preface

The mitochondrion is a unique and ubiquitous organelle that contains its own genome, encoding essential proteins that are major components of the respiratory chain and energy production system. It is reasonable therefore that mitochondria play a dominant role in the life and function of eukaryotic cells including neurons and glia, as their survival and activity depend almost exclusively upon mitochondrial energy production and supply.

Thus, mitochondrial functional impairment is related to a substantial number of neurological disorders, most of them having high incidence in the range of neurodegenerative diseases, such as Parkinson's disease, Alzheimer's disease, Friedreich's ataxia, and Huntington's disease.

New techniques improving mitochondrial DNA analysis and detecting mutation penetrance patterns have also increased the awareness of the rate of mitochondrial involvement in the pathogenetic mechanisms of a substantial number of neurological diseases. In addition, since mitochondria synthesize ATP through oxidative phosphorylation (OXPHOS) as the main energy providers for neurons and glia, their dysfunction induces an energy crisis with tragic consequences for cells of high energy demands.

Besides energy production, mitochondria also play a vital role in neuronal and glial calcium homeostasis due to their high capacity for accumulating Ca2+. On the other hand, mitochondria comprise reactive oxygen species (ROS), which control redox status, intracellular Ca2+ levels, and may induce apoptosis by activating the mitochondrial permeability transition pore (mtPTP), a crucial point of excitotoxic neuronal injury.

In a parallel manner, ROS may play an important role in inflammatory conditions of the brain such as Multiple Sclerosis (MS) either by decreased activity of mitochondrial enzymes, or by peroxidation of lipids, proteolipid protein or myelin basic protein, therefore destabilizing the anoxic myelin sheath.

In reality, the role of the mitochondria in neuronal survival and function is multidimensional. Thus metabolic intermediates derived from Krebs cycle in the mitochondria may behave as co-factors for epigenetic modifications, playing an essential role in histone acetylation and in the addition and removal of epigenetic marks, controlling the expression of a substantial number of genes in the neuronal genome.

Mitochondria also contribute to the synthesis of steroid hormones such as glucocorticoids, mineralocorticoids, progestogens, estrogens, and androgens; and facilitate the interaction of catecholamines with their receptors, therefore promoting neuronal plasticity by stress mediators. Thyroid hormones also stimulate mitochondrial activity by enhancing oxidative phosphorylation. In turn, steroid hormones regulate mitochondrial respiratory capacity and oxidative stress, since mitochondria are particularly sensitive and dependable on hormonal stimulation. Given that sex hormones regulate mitochondrial biogenesis and function, it is expected that

mitochondria express sexual dimorphism and show different reactions to stress and disease in males and females.

Mitochondria modulate systemic energy homeostasis in both sexes by the production and release of mitokines, such as humanin, and by operating as the central organelle of adaptation and stress-response modulation.

From the morphological point of view, mitochondrial alterations are accurately visualized in light microscopy in a properly fixed material by means of a number of special staining reactions. Transmission electron microscopy has also contributed substantially in the morphological study of mitochondria in neurological diseases revealing the disruption of the cristae and the disarrangement of their inner architecture, taking in consideration also that mitochondria exhibit high morphological variability according to activity and metabolic demands of the cell. Using transmission electron microscopy, it was also observed that in Alzheimer's disease, mitochondrial cristae are disrupted from the initial stages of the disease even in areas of the brain with minimal Alzheimer's pathology.

A considerable number of syndromes have been described with marked neurological phenomena in the spectrum of mitochondrial disorders. It must be highlighted that the severity of the clinical manifestation of mitochondrial dysfunction varies considerably, given the threshold in the degree of mitochondrial deficiency for the clinical expression of the disease. Thus, the symptoms and clinical phenomena are continuously aggravated, in the majority of the cases of mitochondrial diseases, as the age of the patient advances and the energy deficiency increases in various organs. However, the most frequent clinical manifestations that have been reported in mitochondrial diseases include cognitive impairment, epilepsy, myopathies, renal and liver dysfunction, cardiopathy and disorders of inner secretion resulting in hormonal imbalance.

Molecular genetic testing and muscle biopsy for histochemical investigation in light microscopy and ultrastructural study of the specimens in electron microscopy are essential diagnostic procedures for the documentation and final verification of the diagnosis of mitochondrial disorders. In addition, biochemical testing in blood, urine, and spinal fluid associated with neuroimaging would be useful diagnostic procedures in following up the progression of mitochondrial diseases.

A final escape from the labyrinth of mitochondrial related neurological disorders is extremely difficult and less pragmatic under the present circumstances. Thus the treatment for the majority of mitochondrial diseases remains mostly symptomatic. Therapeutic factors enhancing mitochondrial biogenesis and restoring nitric oxide production might be beneficial at the initial stages of the diseases. The administration of cardiolipin protector and agents aiming at ameliorating mitochondrial function, such as antioxidants, riboflavin, idebenone, CoQ10 (ubiquinone) and thiamin might be reasonably prescribed in attempting to improve the physical and mental condition of the patients who suffer from mitochondrial disorders.

Prospectively, an efficient treatment could be based on a stable modulation of mtDNA heteroplasmy, whereas gene therapy, liver transplantation, gene transfer and tRNA-targeted therapeutic attempts as well as stem cell therapy for nuclear DNA mutations are promising therapeutic endeavors.

In conclusion, an efficient and easy treatment of mitochondrial dysfunction would open new bright horizons in the therapy of inflammatory and neurodegenerative disorders, being beneficial in the amelioration of the quality of life of a substantial number of patients.

In this book, the authors attempted to describe the complex relationship of mitochondrial disorders with neurodegeneration for further clarification and analysis of the multidimensional background of the pathogenetic mechanisms of neurological diseases.

> **Stavros J. Baloyannis MD, PhD** Professor Emeritus, Aristotelian University, Thessaloniki, Greece

Section 1 Introduction

#### **Chapter 1**

## Introductory Chapter: Mitochondrial Alterations and Neurological Disorders

*Stavros J. Baloyannis*

#### **1. Introduction**

Mitochondria (from Greek mito, μίτος, thread; and chondrion, χόνδριον, thick granule) are principal cell organelles, which participate in a wide spectrum of essential cellular functions, being the main energy providers for living eukaryotic cells, especially for neurons and glia, which are characterized by high metabolic activity and energy consumption.

Thus, it is expectable that mitochondrial dysfunction, having pleotropic effect on the cell, may play a crucial role in a substantial number of serious neurological disorders including Alzheimer's disease (AD) [1, 2], Parkinson's disease (PD) [3] Huntington's disease [4, 5], amyotrophic lateral sclerosis (ALS) [6], multiple sclerosis (MS) [7, 8], as well as some of the major psychiatric diseases [9], given that both, neurons and glia, are particularly sensitive and vulnerable to energy decline [10].

Mitochondria hypothesis of those devastating diseases advocates reasonably in favor of the important role that mitochondrial dysfunction may play in the early stages of neurodegeneration by inducing energy deficiency and oxidative stress [11].

However, the majority of the mitochondrial diseases, being maternally inherited, which are designated as mitochondrial encephalomyopathies [12], are closely connected either with the impairment of nucleus-to-mitochondria signaling or with mutations in mtDNA or nuclear genome that affect seriously the mitochondrial respiratory function even from the initial steps of the life [13], inducing defective oxidative phosphorylation (OXPHOS).

#### **2. The genetic background of mitochondrial dysfunction**

It is well known that mitochondria, as very specific organelles, include several copies (2–10 copies) of their own DNA (mtDNA), which consists of a 16.5 kb circular DNA molecule, being particularly prone to mutation [14]. mtDNA encodes for 37 genes, 13 of them encoding 13 polypeptides, which all are major components of OXPHOS complexes I, III, IV, and V, along with 22 tRNAs and 2 rRNAs, which play an essential role for the expression of the 13 subunits [15].

Mutations in mtDNA may be related to 25% of childhood-onset diseases [16] and to 75% of adult-onset ones [17], depending on the existing homoplasmy or heteroplasmy. In addition, the accumulation of mtDNA mutations can also induce or facilitate the aging process [18], since a common phenomenon in mammalian aging is the substantial decrease of electron transfer in mitochondria [19, 20].

#### **3. Biological consequences of mitochondrial dysfunction**

The mitochondria in addition to energy production compose also reactive oxygen species (ROS), which control redox status and intracellular Ca2+ levels and may induce apoptosis, by activating the mitochondrial permeability transition pore (mtPTP) [21]. In addition, mitochondria play a very important role in neuronal and glial calcium homeostasis due to their high capacity to accumulating Ca2+ [22].

Resting neurons contain usually minimal Ca2+ that can be increased by the activation of NMDA glutamate receptors, which induce a massive entry of Ca2+ into neurons, resulting in its high accumulation in the mitochondria [23]. Continuous activation of NMDA receptors would therefore induce Ca2+ overload of the mitochondria with the tragic consequence of the cell apoptosis, which frequently occurs as an epilogue of the excitotoxicity [24].

The apoptosis consists of a wide spectrum of biological phenomena [25] including the release of caspase activators [26], the alterations of the electron transport system, the change of mitochondrial transmembrane potential, the disruption of the cellular oxidation-reduction equilibrium, and the activation of the pro-apoptotic Bcl-2 family proteins [27, 28].

In the majority of the mitochondria-related neurological disorders, the functional or morphological alteration of the mitochondrial may be induced by increased ROS production, abnormal protein aggregates (Ab, tau) [29, 30], mutations in genes encoded by the mitochondrial and nuclear genome, and exposure of the cell to toxic factors [31].

#### **4. The morphology of mitochondria in health and disease**

Cell mitochondria could be visualized in light microscopy in properly fixed material by means of a number of special staining reactions [32–34]. It is observed that their size generally ranges from 0.5 to 1 micron in diameter, being changeable due to frequent divisions and fusions, which are controlled by mitofusin activity [35]. The shape of the mitochondria is also continuously changed due to their impressive active motility, controlled by calcium signal [36, 37], given that they are in constant flux, especially in brain's areas of high energy consumption in order to contribute in energy supply and to participate in the intracellular signaling actively [38].

Electron microscopy has been contributing greatly in the study of mitochondria in health and disease [39, 40]. Each mitochondrion in healthy condition is surrounded by a limiting double membrane and includes numerous longitudinal or tubular invaginations called mitochondrial cristae that are folds of the inner layer of the double membrane [41], which is four times greater than the outer one.

The cristae are mostly arranged perpendicularly to the long axis of the organelle, exhibiting a high morphological variability according to metabolic demands of the cell [42], being frequently lamellar, tubular, or triangle-shaped. In the majority of the mitochondria, the cristae are arranged parallel to one another inside a structureless matrix, which is clearly seen among the cristae.

Cardiolipin seems to play a crucial role in the morphology of cristae, since the disruption of cardiolipin biosynthesis induces obvious alteration of the cristae morphology [43]. In addition, Opa1, which is a GTPase, demonstrating dynaminlike properties, plays a substantial role in the modulation of the cristae structure and in their remodeling during mitochondrial fusion and fission [44] and apoptotic process [45]. The cristae have a high protein content [46], being also the principal site of the oxidative phosphorylation [47].

*Introductory Chapter: Mitochondrial Alterations and Neurological Disorders DOI: http://dx.doi.org/10.5772/intechopen.91051*

Electron microscope tomography, revealing the three-dimensional appearance of the cristae, shows that they are connected with the inner mitochondrial membrane by a narrow, tubular opening, characterized as "crista junction" (CJ), which is associated with protein import [48] and mitochondrial inner compartmentalization [49].

In neurodegeneration such as in Alzheimer's disease, mitochondrial cristae are disrupted even from the initial stages of the disease, and concentric patterns of cristae membranes are frequently seen [50].

#### **5. Mitochondrial trafficking and concentration**

Mitochondria, like many other cell organelles, are oriented and positioned properly in neurons and glia in order to be able to fulfill the energy demands of the cells perpetually. Thus, neurons, axons, dendrites, and synapses, which are characterized by high ceaseless activity, have intensive mitochondrial motility and impressive concentrations [51], via various trafficking patterns [52].

Axonal transport of mitochondria [53] requires microtubules (MTs) [54, 55] or actin filaments in axons [56], which facilitate the movement of the mitochondria in areas of high metabolic demands and increased energy consumption [57]. It is noticed that disruption of axonal transport of mitochondria occurs as an early phenomenon in cases of neuroinflammation [58], including multiple sclerosis [59–61].

#### **6. Clinical expression of mitochondrial dysfunction**

A considerable number of syndromes have been described with marked neurological phenomena in the spectrum of mitochondrial disorders [62]. The severity of the clinical manifestation of mitochondrial dysfunction varies considerably, given that there exists a threshold in the degree of mitochondrial deficiency for the clinical expression of the disease [63, 64]. Thus, the symptoms and clinical phenomena are continuously aggravated, in the majority of the cases of mitochondrial diseases, as the age of the patients advances [65]. It is reasonable to accept that organs with high energy demand would be more seriously affected by the mitochondrial dysfunction than others with low level of energy necessity. Thus the brain, the skeletal muscles, and the heart have a typical involvement in adolescence and adulthood, though multi-system manifestation is not also an uncommon phenomenon, especially in childhood.

Many clinical syndromes have been described that are associated with mitochondrial dysfunction including encephalomyopathy, stroke-like episodes, myoclonic epilepsy, neuro-gastrointestinal phenomena, cranial or peripheral neuropathy, ataxia, retinitis pigmentosa, chronic progressive external ophthalmoplegia which are associated frequently with lactic acidosis, mental retardation, or progressive mental decline [66].

In addition, oxidative stress, due to mitochondrial dysfunction, plays a principal role, as causative factor, in the neurodegeneration [67] and in Alzheimer's disease particularly [68, 69], and it is considered as been among the potential risk factors for the neurometabolic and neoplastic diseases, as well as obesity [70].

Molecular genetic testing on one hand and muscle biopsy on the other hand for the histochemical investigation in light microscopy and the ultrastructural study in electron microscopy of the muscle tissue are essential diagnostic procedures for approaching the diagnosis of mitochondrial disorders [71]. In addition, biochemical testing in blood, urine, and spinal fluid associated with neuroimaging [72]

would be useful diagnostic procedures in following in time the progression of mitochondrial diseases [71].

#### **7. The final escape**

A final escape from the labyrinth of mitochondrial-related neurological disorders is extremely difficult and less pragmatic under the present circumstances. Prospectively, an efficient treatment could be based on a stable modulation of mtDNA heteroplasmy [73], whereas gene therapy, gene transfer, and tRNA-targeted therapeutic attempts [74] as well as stem cell therapy for nuclear DNA mutations [75, 76] are very promising therapeutic endeavors with substantial medical and scientific value [77, 78].

In addition, an efficient and easy to apply treatment of mitochondrial dysfunction would open new bright horizons in the therapy of the inflammatory and neurodegenerative disorders [79], being beneficial in the amelioration of the quality of life of a substantial number of seriously suffering human beings.

#### **Author details**

Stavros J. Baloyannis Aristotle University of Thessaloniki, Greece

\*Address all correspondence to: sibh844@otenet.gr

© 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.

*Introductory Chapter: Mitochondrial Alterations and Neurological Disorders DOI: http://dx.doi.org/10.5772/intechopen.91051*

#### **References**

[1] Baloyannis SJ. Mitochondrial alterations in Alzheimer's disease. Journal of Alzheimer's Disease. 2006;**9**(2):119-126

[2] Wang X, Wang W, Li L, Perry G, Lee HG, Zh'u X. Oxidative stress and mitochondrial dysfunction in Alzheimer's disease. Biochimica et Biophysica Acta. 2014;**1842**(8):1240-1247

[3] Schapira AHV. Mitochondria in the aetiology and pathogenesis of Parkinson's disease. The Lancet Neurology. 2008;**7**(1):97-109

[4] Browne SE. Mitochondria and Huntington's disease pathogenesis: Insight from genetic and chemical models. Annals of the New York Academy of Sciences. 2008;**1147**(1):358-382

[5] Damiano M, Galvan L, Déglon N, Brouillet E. Mitochondria in Huntington's disease. Biochimica et Biophysica Acta. 2010;**1802**(1):52-61

[6] Orrell RW, Schapira AHV. Mitochondria and amyotrophic lateral sclerosis. International Review of Neurobiology. 2002;**53**:411-426

[7] Dutta R, McDonough J, Yin X, et al. Mitochondrial dysfunction as a cause of axonal degeneration in multiple sclerosis patients. Annals of Neurology. 2006;**59**(3):478-489

[8] de Barcelos IP, Troxell RM, Graves JS. Mitochondrial dysfunction and multiple sclerosis. Biology. 2019;**8**(2):37

[9] Rezin GT, Amboni G, Zugno AI, Quevedo J, Streck EL. Mitochondrial dysfunction and psychiatric disorders. Neurochemical Research. 2009;**34**(6):1021-1029

[10] Chrzanowska-Lightowlers ZMA, Lightowlers RN. How much does a

disrupted mitochondrial network influence neuronal dysfunction? EMBO Molecular Medicine. 2019;**11**(1):e9899

[11] Area-Gomez E, Guardia-Laguarta C, Schon EA, Przedborski S. Mitochondria, OxPhos, and neurodegeneration: Cells are not just running out of gas. The Journal of Clinical Investigation. 2019;**129**(1):34-45

[12] DiMauro S. Mitochondrial encephalomyopathies--fifty years on: The Robert Wartenberg lecture. Neurology. 2013;**81**:281-291

[13] DiMauro S. Mitochondrial diseases. Biochimica et Biophysica Acta (BBA) - Bioenergetics. 2004;**1658**(1-2):80-88

[14] Taylor RW, Turnbull DM. Mitochondrial DNA mutations in human disease. Nature Reviews. Genetics. 2005;**6**(5):389

[15] Frazier AE, Thorburn DR, Compton AG. Mitochondrial energy generation disorders: Genes, mechanisms, and clues to pathology. Journal of Biological Chemistry. 2019;**294**(14):5386-5395

[16] Skladal D, Halliday J, Thorburn DR. Minimum birth prevalence of mitochondrial respiratory chain disorders in children. Brain. 2003;**126**:1905-1912

[17] Gorman GS, Schaefer AM, Ng Y, Gomez N, Blakely EL, Alston CL, et al. Prevalence of nuclear and mitochondrial DNA mutations related to adult mitochondrial disease. Annals of Neurology. 2015;**77**:753-759

[18] Müller-Höcker J. Mitochondria and ageing. Brain Pathology. 1992;**2**(2):149-158

[19] Fang EF, Scheibye-Knudsen M, Chua KF, Mattson MP, Croteau DL, Bohr VA. Nuclear DNA damage signaling to mitochondria in ageing. Nature Reviews. Molecular Cell Biology. 2016;**17**(5):308-321

[20] Breitenbach M, Rinnerthaler M, Hartl J, et al. Mitochondria in ageing: There is metabolism beyond the ROS. FEMS Yeast Research. 2014;**14**(1):198-212

[21] Wallace DC, Fan W. Energetics, epigenetics, mitochondrial genetics. Mitochondrion. 2010;**10**:12-31

[22] Marchi S, Patergnani S, Missiroli S, et al. Mitochondrial and endoplasmic reticulum calcium homeostasis and cell death. Cell Calcium. 2018;**69**:62-72

[23] Abeti R, Abramov AY. Mitochondrial Ca2+ in neurodegenerative disorders. Pharmacological Research. 2015; **99**:377-381

[24] Sattler R, Xiong Z, Lu WY, MacDonald JF, Tymianski M. Distinct roles of synaptic and extrasynaptic NMDA receptors in excitotoxicity. The Journal of Neuroscience. 2000;**20**(1):22-33

[25] Green DR, Reed JC. Mitochondria and apoptosis. Science. 1998;**281**(5381):1309-1312

[26] Fan TJ, Han LH, Cong RS, Liang J. Caspase family proteases and apoptosis. Acta Biochimica et Biophysica Sinica Shanghai. 2005;**37**(11):719-727

[27] Friedlander RM. Apoptosis and caspases in neurodegenerative diseases. The New England Journal of Medicine. 2003;**348**(14):1365-1375

[28] Antonsson B, Martinou J-C. The Bcl-2 protein family. Experimental Cell Research. 2000;**256**(1):50-57

[29] Hashimoto M, Rockenstein E, Crews L, Masliah E. Role of protein aggregation in mitochondrial dysfunction and neurodegeneration in Alzheimer's and Parkinson's diseases. Neuromolecular Medicine. 2003;**4**(1-2):21-36

[30] Wang X, Su BO, Siedlak SL, Moreira PI, Fujioka H, et al. Amyloid-β overproduction causes abnormal mitochondrial dynamics via differential modulation of mitochondrial fission/ fusion proteins. Proceedings of the National Academy of Sciences. 2008;**105**(49):19318-19323

[31] Lee HK, Cho YM, Kwak SH, Lim S, Park KS, Shim EB. Mitochondrial dysfunction and metabolic syndromelooking for environmental factors. Biochimica et Biophysica Acta. 2010;**1800**(3):282-289

[32] Cain AJ. An easily controlled method for staining mitochondria. Journal of Cell Science. 1948;**3**(6):229-231

[33] Roels F. Cytochrome c and cytochrome oxidase in diaminobenzidine staining of mitochondria. The Journal of Histochemistry and Cytochemistry. 1974;**22**(6):442-444

[34] Neto BA, Carvalho PH, Santos DC, Gatto CC, Ramos LM, et al. Synthesis, properties and highly selective mitochondria staining with novel, stable and superior benzothiadiazole fluorescent probes. RSC Advances. 2012;**2**(4):1524-1532

[35] Santel A, Fuller MT. Control of mitochondrial morphology by a human mitofusin. Journal of Cell Science. 2001;**114**(5):867-874

[36] Yi M, Weaver D, Hajnócky G. Control of mitochondrial motility and distribution by the calcium signal: a homeostatic circuit. The Journal of Cell Biology. 2004;**167**(4):661-672

#### *Introductory Chapter: Mitochondrial Alterations and Neurological Disorders DOI: http://dx.doi.org/10.5772/intechopen.91051*

[37] Wang X, Schwarz TL. The mechanism of Ca2+−dependent regulation of kinesin-mediated mitochondrial motility. Cell. 2009;**136**(1):163-174

[38] Rizzuto R, Bernardi P, Pozzan T. Mitochondria as all-round players of the calcium game. The Journal of Physiology. 2000;**529**:37-47

[39] Claude A, Fullam EF. An electron microscope study of isolated mitochondria: Method and preliminary results. The Journal of Experimental Medicine. 1945;**81**(1):51

[40] Sun MG, Williams J, Munoz-Pinedo C, et al. Correlated threedimensional light and electron microscopy reveals transformation of mitochondria during apoptosis. Nature Cell Biology. 2007;**9**(9):1057-1065

[41] Lea PJ, Hollenberg MJ. Mitochondrial structure revealed by high-resolution scanning electron microscopy. The American Journal of Anatomy. 1989;**184**:245-257

[42] Patten DA, Wong J, Khacho M, Soubannier V, Mailloux RJ, et al. Slack OPA1-dependent cristae modulation is essential for cellular adaptation to metabolic demand. The EMBO Journal. 2014;**33**:2676-2691

[43] Xu Y, Sutachan JJ, Plesken H, Kelley RI, Schlame M. Characterization of lymphoblast mitochondria from patients with Barth syndrome. Laboratory Investigation. 2005;**85**:823-830

[44] Chan DC. Fusion and fission: Interlinked processes critical for mitochondrial health. Annual Review of Genetics. 2012;**46**:265-287

[45] Frezza C, Cipolat S, Martins de Brito O, Micaroni M, Beznoussenko GV, et al. OPA1 controls apoptotic cristae remodeling independently

from mitochondrial fusion. Cell. 2006;**126**:177-189

[46] Neupert W, Herrmann JM. Translocation of proteins into mitochondria. Annual Review of Biochemistry. 2007;**76**:723-749

[47] Gilkerson RW, Selker JM, Capaldi RA. The Cristal membrane of mitochondria is the principal site of oxidative phosphorylation. FEBS Letters. 2003;**546**:355-358

[48] Perkins GA, Renken CW, van der Klei IJ, Ellisman MH, Neupert W, Frey TG. Electron tomography of mitochondria after the arrest of protein import associated with Tom19 depletion. European Journal of Cell Biology. 2001;**80**:139-150

[49] Mannella CA, Marko M, Buttle K. Reconsidering mitochondrial structure: New views of an old organelle. Trends in Biochemical Sciences. 1997;**22**:37-38

[50] Baloyannis SJ. Mitochondrial alterations in Alzheimer's disease. Journal of Alzheimer's Disease. 2006;**9**:119-126

[51] Chang DTW, Reynolds IJ. Mitochondrial trafficking and morphology in healthy and injured neurons. Progress in Neurobiology. 2006;**80**(5):241-268

[52] Sheng ZH. Mitochondrial trafficking and anchoring in neurons: New insight and implications. The Journal of Cell Biology. 2014;**204**(7):1087-1098

[53] Hollenbeck PJ, Saxton WM. The axonal transport of mitochondria. Journal of Cell Science. 2005;**118**(23):5411-5419

[54] Grafstein B, Forman DS. Intracellular transport in neurons. Physiological Reviews. 1980;**60**:1167-1283

[55] Hollenbeck PJ. The pattern and mechanism of mitochondrial transport in axons. Frontiers in Bioscience. 1996;**1**:91-102

[56] Langford GM, Kuznetsov SA, Johnson D, Cohen DL, Weiss DG. Movement of axoplasmic organelles on actin filaments assembled on acrosomal processes: Evidence for a barbed-enddirected organelle motor. Journal of Cell Science. 1994;**107**:2291-2298

[57] Bridgman PC. Myosin-dependent transport in neurons. Journal of Neurobiology. 2004;**58**:164-174

[58] Errea LO, Moreno B, González FA, García-Roves PM, Villoslada P. The disruption of mitochondrial axonal transport is an early event in neuroinflammation. Journal of Neuroinflammation. 2015;**12**:152

[59] Ghafourifar P, Mousavizadeh K, Parihar MS, Nazarewicz RR, Parihar A, Zenebe WJ. Mitochondria in multiple sclerosis. Frontiers in Bioscience. 2008;**13**:3116-3126

[60] Nikic I, Merkler D, Sorbara C, Brinkoetter M, Kreutzfeldt M, Bareyre FM, et al. A reversible form of axon damage in experimental autoimmune encephalomyelitis and multiple sclerosis. Nature Medicine. 2011;**17**:495-499

[61] Sorbara CD, Wagner NE, Ladwig A, Nikic I, Merkler D, Kleele T, et al. Pervasive axonal transport deficits in multiple sclerosis models. Neuron. 2014;**84**:1183-1190

[62] McFarland R, Taylor RW, Turnbull DM. A neurological perspective on mitochondrial disease. Lancet Neurology. 2010;**9**:829-840

[63] Wallace DC. Mitochondrial diseases in man and mouse. Science. 1999;**283**(5407):1482-1488

[64] Finsterer J. Central nervous system manifestations of mitochondrial disorders. Acta Neurologica Scandinavica. 2006;**114**(4):217-238

[65] Calabrese V, Scapagnini G, Stella AG, Bates TE, Clark JB. Mitochondrial involvement in brain function and dysfunction: Relevance to aging, neurodegenerative disorders and longevity. Neurochemical Research. 2001;**26**(6):739-764

[66] Ng YS, Turnbull DM. Mitochondrial disease: Genetics and management. Journal of Neurology. 2016;**263**(1): 179-191

[67] Islam MT. Oxidative stress and mitochondrial dysfunctionlinked neurodegenerative disorders. Neurological Research. 2017;**39**(1):73-82

[68] Baloyannis SJ. Mitochondria: Strategic point in the field of Alzheimer's disease. Journal of Alzheimers and Neurodegenerative Diseases. 2016;**2**:004

[69] Baloyannis SJ. What has electron microscopy contributed to Alzheimer's research? Future Neurology. 2015;**10**(6):515-527

[70] Greaves LC, Reeve AK, Taylor RW, Turnbull DM. Mitochondrial DNA and disease. The Journal of Pathology. 2012;**226**:274-286

[71] Parikh S, Goldstein A, Koenig MK, Scaglia F, Enns GM, Saneto R, et al. Diagnosis and management of mitochondrial disease: A consensus statement from the mitochondrial medicine society. Genetics in Medicine. 2015;**17**(9):689-701

[72] Morava E, van den Heuvel L, Hol F, et al. Mitochondrial disease criteria: Diagnostic applications in children. Neurology. 2006;**67**:1823-1826

*Introductory Chapter: Mitochondrial Alterations and Neurological Disorders DOI: http://dx.doi.org/10.5772/intechopen.91051*

[73] Minczuk M, Papworth MA, Miller JC, Murphy MP, Klug A. Development of a single-chain, quasidimeric zinc-finger nuclease for the selective degradation of mutated human mitochondrial DNA. Nucleic Acids Research. 2008;**36**:3926-3938

[74] Yarham JW, Al-Dosary M, Blakely EL, Alston CL, Taylor RW, Elson JL, et al. A comparative analysis approach to determining the pathogenicity of mitochondrial tRNA mutations. Human Mutation. 2011;**32**:1319-1325

[75] Hussein E. Non-myeloablative bone marrow transplant and platelet infusion can transiently improve the clinical outcome of mitochondrial neurogastrointestinal encephalopathy: A case report. Transfusion and Apheresis Science. 2013;**49**:208-211

[76] Spendiff S, Reza M, Murphy JL, Gorman G, Blakely EL, Taylor RW, et al. Mitochondrial DNA deletions in muscle satellite cells: Implications for therapies. Human Molecular Genetics. 2013;**22**:4739-4747

[77] Kerr DS. Review of clinical trials for mitochondrial disorders: 1997-2012. Neurotherapeutics. 2013;**10**:307-319

[78] Nightingale H, Pfeffer G, Bargiela D, Horvath R, Chinnery PF. Emerging therapies for mitochondrial disorders. Brain. 2016;**139**(6):1633-1648

[79] Baloyannis SJ, Baloyannis JS. Mitochondrial alterations in Alzheimer's disease. Neurobiology of Aging. 2004;**25**:405-406

Section 2

## Mitochondria and Hormones

#### **Chapter 2**

### Pathology Associated with Hormones of Adrenal Cortex

*Lovelesh K. Nigam, Aruna V. Vanikar, Rashmi D. Patel, Kamal V. Kanodia and Kamlesh S. Suthar*

#### **Abstract**

Adrenal gland is an endocrine organ comprising of an outer cortex and inner medulla. These secrete various hormones that have a vital role in maintaining the normal homeostasis of the body. Lesions of adrenal cortex are quite common to encounter and most of these are related to the hormones secreted by three layers of adrenal cortex: the zona glomerulosa, the zona fasciculata, and the zona reticularis. Also it is very infrequent to encounter metastatic lesions in the adrenal glands too. So it is very important as a part of a clinician as well as a pathologist to know the pattern in which these hormones are secreted along with their physiological roles. Thus this chapter includes the disease that are related to excess as well as deficiencies of the hormones secreted by adrenal cortex. The chapter also includes various genetic syndromes that are associated with the disorders associated with hormones of adrenal cortex. The last part of the chapter includes a brief description of various benign as well as malignant lesions, the pathological as well as the etiological aspects and the hormonal abnormalities associated. This chapter thus mainly focuses on the pathology associated with the adrenal cortex and hormones secreted by the various layers of adrenal cortex.

**Keywords:** adrenal cortex, hormones, Cushing syndrome

#### **1. Introduction**

Adrenal gland consists of an outer cortex and inner medulla; the cortex is further subdivided into three distinct zones: the zona glomerulosa, the zona fasciculata, and the zona reticularis. Mineralocorticoids (aldosterone) secreted from the zona glomerulosa are essential for fluid and electrolyte balance and the reninangiotensin-aldosterone system. The fasciculata secretes glucocorticoids (mainly cortisol). The zona reticularis produces steroid sex hormones called androgens. These hormones play an important role in maintaining the normal homeostasis of the body [1–3]. However, it is quite common to encounter disorders related to the hormones of these three layers. These disorders could be possibly due to adrenal cortical masses secondary to cortical hyperplasia. It is very infrequent to encounter metastatic lesions in the adrenal glands.

This chapter mainly focuses on the pathology related to adrenal cortex which includes various forms of adrenocortical hyperplasia and benign and malignant neoplasms of the adrenal gland which lead to various hormonal imbalances encountered in clinical practice. Hormonal deficiency is due to inherited glandular or enzymatic

disorder, destruction of pituitary gland by autoimmune disorders, infection, infarction, or others [4, 5]. The major disorders of the adrenal cortex are characterized by excessive or deficient secretion of each type of adrenocortical hormone.

#### **2. Functional manifestations**

The lesions of the adrenal cortex could be functional as well as nonfunctional, which means that patients with these lesions may exhibit clinical symptoms that are due to hypersecretion of hormones released. Usually cortical hyperplasia and adenomas are nonfunctioning. The functional syndromes associated with pathology of adrenal cortex are hypercortisolism (Cushing's syndrome), adrenal insufficiency (Addison's disease), hyperaldosteronism, and androgen excess (adrenogenital syndrome) [4–9].

#### **3. Adrenal hyperplasia**

Adrenal hyperplasia is characterized as a smooth, diffuse, bilateral enlargement of the adrenal glands, wherein the glands retain their adreniform shape. Hyperplasia can be either macronodular or micronodular. They are commonly unilateral; however bilateral cases are also observed [7, 8]. Broadly adrenal cortical hyperplasia can be grouped into three main categories: ACTH-dependent (adrenocorticotropic hormone), ACTH-independent, and congenital adrenal hyperplasia (CAH). Cushing's syndrome is one of the common functional manifestations of adrenal gland hyperplasia and therefore is discussed first [8, 10, 11].

#### **3.1 Cushing's syndrome**

#### *3.1.1 Definition*

It is a syndrome which encompasses various clinical features due to chronic excess of glucocorticoids. The incidence is nearly 1–2 per 100,000 population per year. Harvey Cushing was the first to observe pituitary adenomas associated with hypercortisolism in 1932 [10–13]. Cushing's syndrome, caused by prolonged exposure of tissues to high levels of cortisol, presents as constellation of symptoms including central obesity, muscle fatigue/atrophy, hirsutism, infertility, osteoporosis, moon facies, dorsocervical and supraclavicular fat pads, and wide purple striae [8, 10, 12]. The syndrome may be ACTH-dependent or ACTH-independent. A fair number of cases attributed to iatrogenic causes are also identified. Most of the cases of Cushing's syndrome are due to ACTH hypersecretion from the anterior pituitary and are associated with pituitary cortical adenoma. Majority of the cases, about 80–90%, show diffuse hyperplasia of the adrenal cortex [9, 10]. Nearly 15% of cases do present with ectopic ACTH secretion associated with small cell lung carcinoma or bronchial carcinoid. Thymic carcinoids, pancreatic islet cell tumor, pheochromocytomas, and medullary carcinoma of thyroid form minor group of tumors associated with ectopic ACTH secretion [12]. In a study by Ejaz et al., lung tumors constituted 44.4% of all cases of neoplasm-related ectopic ACTH secretion causing Cushing's syndrome [14]. Clinically patients with Cushing's syndrome present with diastolic hypertension, hypokalemia, and edema. Hypogonadism and amenorrhea can also be seen in these patients which are attributed to suppression of gonadotropin secretion secondary to excess glucocorticoid secretion [10–14].

*Pathology Associated with Hormones of Adrenal Cortex DOI: http://dx.doi.org/10.5772/intechopen.84815*

#### *3.1.2 Cushing disease*

Cushing disease, resulting from a pituitary corticotropic adenoma, and rarely carcinoma, makes up to 80–85% of endogenous Cushing's syndrome cases [8, 10, 15].

#### *3.1.3 Investigating a case of Cushing's syndrome*

A two-stage test is usually recommended in a patient to rule out Cushing's syndrome [8, 10, 15–17]:

	- a.Plasma ACTH measurement: Low plasma ACTH level suggests an adrenal cause of the disease; however normal/high [ACTH] level suggests ectopic ACTH secretion or hypersecretion of ACTH from pituitary (Cushing's disease).
	- b.High-dose dexamethasone suppression test: In this test the patient is administered with 2 mg of dexamethasone, 6 hourly for 48 h, following which plasma cortisol levels are measured. In the case of ectopic ACTH secretion or adrenal limited hypercortisolism, there is a failure of suppression of cortisol secretion. Also it is important to remember that cortisol is not suppressed with either low- or high-dose dexamethasone suppression in adrenal hyperplasia associated with ectopic ACTH production [18].

#### *3.1.3.1 The 24-h urinary-free cortisol test*

This investigation is used primarily for the diagnosis of hypercortisolism due to Cushing's syndrome, and reference ranges for this test with respect to age are 1.4–20 μg/24 h (3–8 years), 2.6–37 μg/24 h (9–12 years), 4–56 μg/24 h (13–17 years), and 3.5–45 μg/24 h in individuals ≥18 years of age. A 24-h urine sample with boric acid (10 g) as preservative is advisable for performing this analysis [10, 17–19].

#### *3.1.4 ACTH-independent Cushing's syndrome*

Nearly 15–20% of Cushing's syndrome are associated with ACTH-independent hypercortisolism and are secondary to a functioning adenoma or carcinoma. Diagnosis of ACTH-independent Cushing's syndrome includes clinical features of hypercortisolism, absence of serum cortisol diurnal rhythm, elevated late-night cortisol levels, and incomplete suppression of cortisol production with low-dose dexamethasone suppression test [10, 17–20].

#### *3.1.5 Pathological findings*

Adrenal glands from patients with Cushing's syndrome/hyperplasia appear variably enlarged in size and weigh approximately 6–12 g. The cortical width is widened as compared to the reticulosa. The zona fasciculata usually shows nodular hyperplasia. Nearly 10–20% of the patients reveal bilateral nodular hyperplasia, and up to 30% of patients may have normal adrenal morphology [2, 20, 21].

#### **3.2 Primary pigmented nodular adrenocortical disease (PPNAD)**

Primary pigmented nodular adrenocortical disease is a rare cause of childhood Cushing's disease having female preponderance, whereas Cushing's disease is common in prepubertal males [20–22]. It is the main endocrine manifestation of Carney complex (a multiple neoplasia syndrome caused by mutation in PRKAR1A gene) [23]. This is an autosomal dominant syndrome and is characterized by cutaneous lentigines, myxoma, schwannomas, and endocrinopathy [11, 23]. It was first described by Aidan Carney and co-workers in 1985. Almost 25–30% of patients with Carney complex have ACTH-independent Cushing's syndrome. Cutaneous pigmentation is the commonest manifestation of the disease [24]. Lentigines are seen in most patients, and this characteristic manifestation can be used to make the definitive diagnosis. The name is derived from the macroscopic appearance of the adrenals that show characteristic small pigmented micronodules in the adrenal cortex. The disease typically involves bilateral adrenal glands. Grossly the adrenal glands may have variable size. The most characteristic finding is the presence of multiple brown-black pigmented cortical nodules that measure 1 mm to 3 cm in diameter. The adjacent cortical tissue invariable shows atrophy. These pigmented nodules may extend into corticomedullary junction or peri-adrenal fat [9, 11, 24].

On microscopy these tumors appear as sharply circumscribed, unencapsulated tumors composed of large eosinophilic lipid-poor cells similar to the zona reticularis arranged predominantly in trabecular growth pattern. However the nucleus appears enlarged, with a variable degree of pleomorphism and prominent nucleoli. There is prominent lipofuscin deposit. Lipid-rich fasciculata-like cells are also seen invariably. The tumor may have focal areas of necrosis, mitotic activity, myelolipomatous change, and lymphocytic infiltrates [9, 11, 24].

#### **3.3 ACTH-independent macronodular adrenal hyperplasia (AIMAH)**

(Synonyms: ACTH-independent massive bilateral adrenal disease, massive macronodular hyperplasia, giant macronodular adrenal hyperplasia, macronodular adrenal hyperplasia, macronodular hyperplasia).

AIMAH is a disorder characterized by bilateral adrenocortical nodules, associated with ACTH-independent hypercortisolism, without any clinical features of pigmented nodular adrenocortical disease and histological features consistent with atrophic internodular cortex [25]. It is a rare cause of ACTH-independent Cushing's syndrome with slightly male preponderance. The patients present usually at later age (average: 48 years) [24–26]. In few patients with AIMAH, ectopic expression and/or increased sensitivity to gastric inhibitory peptide, vasopressin receptors, and beta-adrenergic receptors is also seen [25].

Grossly these lesions are characterized by nodules in the adrenal cortex, ranging from 1 to 4.2 cm. The adrenal gland weighs approximately 16.7–218 g. The adrenal gland may have a large mass of cortical tissue and multiple bilateral nodules measuring up to 5 cm. Combined adrenal gland weight of more than 300 g has also been noted (normal range: 8–12 g). Histology demonstrates large, yellow macronodules comprising of small cells with eosinophilic cytoplasm. Bilateral adrenalectomy and well-controlled glucocorticoid replacement is the most accepted treatment modality [2, 3, 9, 25, 26].

#### **3.4 Congenital adrenal hyperplasia**

CAH is an autosomal recessive disorder characterized by impaired steroidogenesis finally leading to mineralocorticoid and cortisol deficiency secondary to reduced activity of enzymes required for cortisol biosynthesis in the adrenal cortex. *Pathology Associated with Hormones of Adrenal Cortex DOI: http://dx.doi.org/10.5772/intechopen.84815*

These patients usually present during the perinatal period with ambiguous genitalia in females and salt wasting in males. The milder forms of disease may present later with virilization at puberty or even as irregular menses. Most of the cases (nearly 95%) are attributed to deficiency of the 21-hydroxylase enzyme [27, 28].

Abnormal growth and development, adverse effects on bone and the cardiovascular system, and infertility are few long-term effects seen in these patients. These patients are usually managed by reducing glucocorticoid exposure and improving excess hormone control [29, 30].

Congenital adrenal hyperplasia can be of four forms [8–10, 27–30]:


#### *3.4.1 Congenital adrenal hyperplasia: Classical 21-hydroxylase deficiency*

This form is the most common form of CAH, occurring due to 21-hydroxylase (21-OH) deficiency, accounting for almost 90% of the cases. It occurs with the frequency of 1:12000 to 1:15000 births, and nearly 75% of patients with classic 21-OH


#### **Table 1.** *Hereditary adrenocortical tumor syndromes.*

deficiency also have defect in synthesizing aldosterone. These patients die in the neonatal period due to shock from salt wasting. CAH is associated with multiple tumors like testicular tumors arising from ectopic adrenal cortical rests, testicular and ovarian Leydig cell tumor, and ovarian tumor of the adrenogenital syndrome as ovarian and paraovarian brown masses. Grossly the adrenal gland is marked enlarged having a cerebriform appearance. On cut surface the gland appears tan-brown in color. Under the microscope the adrenal gland reveals diffuse cortical hyperplasia. The cells are compactly arranged like how they are in the zona reticularis [2, 27–30].

**Table 1** illustrates various syndromes associated with adrenocortical lesions [31].

#### **4. Primary hyperaldosteronism (Conn's syndrome)**

This disease was defined first by Conn in 1955, with a prevalence of 5–13%. This syndrome is characterized by an inappropriate increase in production of aldosterone which is relatively independent from the renin-angiotensin mechanism and is non-suppressible by sodium loading. This is one of the leading causes of secondary hypertension in hypertensive adults [32]. Patients with primary aldosteronism may exhibit adrenal cortical hyperplasia or adenoma in 30% of sporadic cases, and nearly 1% of sporadic cases may have adrenocortical carcinoma [33]. Clinically these patients present most commonly as normokalemic hypertension, and severe cases do show hypokalemia (**Table 2**).



*Pathology Associated with Hormones of Adrenal Cortex DOI: http://dx.doi.org/10.5772/intechopen.84815*

#### **Table 2.** *Investigations.*

The aldosterone-to-renin ratio (ARR), a gold standard method to differentiate primary from secondary causes of hyperaldosteronism, is defined as the ratio of plasma aldosterone (expressed in ng/dL) to plasma renin activity (PRA, expressed in ng/mL/h). The cutoff value of ARR is 30 ng/dL per/mL per hour (or 750 pmol/L per ng/mL per hour). The principle behind this test is that as aldosterone secretion rises, PRA in ex vivo testing falls due to sodium retention. This negative feedback response should occur when the aldosterone levels are supraphysiologic for that individual patient, and PRA may fall well before plasma aldosterone is clearly increased. Primary aldosteronism is suspected if the ARR is >30 ng/dL per mL per hour. This method is also helpful in differentiating aldosterone-producing adenoma from bilateral adrenal hyperplasia [34].

#### **4.1 Familial primary aldosteronism**

Familial primary aldosteronism is mainly of three types, all of which are inherited in an autosomal dominant manner [8, 10, 32, 33]:


#### *4.1.1 Gross features*

Grossly, the adrenal gland in cases of idiopathic hyperaldosteronism is rather unremarkable or may exhibit slight enlargement. The enlargement could be due to the presence of micronodules or macronodules. Usually, adenomas are unilateral and solitary. However few cases of bilateral disease have also been reported. These adenomas

**Figure 1.**

*Section from a 22-year-old patient, presented with a 2 cm mass in the right adrenal gland. Histology reveals adenoma with clusters of cells with enlarged lipid-rich cytoplasm (hematoxylin and eosin stain, ×200).*

are mostly intra-adrenal and do not show a capsule. Few cases may reveal the presence of a true capsule or a pseudocapsule [2, 3, 35]. The cut surface of this tumor appears homogenous and golden yellow and is classically described as "canary yellow" [2]. Focal areas of hemorrhage or cystic changes can be present in few cases [35].

#### *4.1.2 Microscopic features*

Microscopically these adenomas appear encapsulated by compressed fibrous rim or fibrous "pseudocapsule." The tumor cells are most commonly arranged in the form of nests or in alveolar pattern. Occasionally these cells may be arranged in short cords and trabeculae. Few cases may show mixed histological patterns. The tumor is composed of four different varieties of cells which may be present in varying proportions. More commonly seen are clear cells, having optically clear cytoplasm and centrally placed nuclei similar to those of the zona fasciculata cells; then there may be cells resembling to the zona glomerulosa or zona reticularis which appear small with compact eosinophilic cytoplasm. Then we have cells that are designated as "hybrid" cells. These hybrid cells have cytological features resembling both the zona fasciculata and glomerulosa (**Figure 1**). The uninvolved portion of adrenal cortex reveals atrophy. This atrophy is secondary to the negative feedback suppression effect of the hypothalamic–pituitary axis. Spironolactone bodies which appear as small, intracytoplasmic eosinophilic inclusions, round to oval, measuring 2–12 mm, are often encountered in adrenal cortical adenoma in patients on spironolactone treatment. These inclusions are delineated from the surrounding cytoplasm by a small, clear halo [2, 7, 9, 35].

#### **5. Adrenal insufficiency**

#### **5.1. Introduction and definition**

Adrenal insufficiency was first described by Thomas Addison in 1855 and was popularly known as Addison's disease. This disorder can occur either due to failure of the adrenal gland or impairment of the hypothalamic–pituitary axis [36]. Clinically this syndrome is characterized by weakness, fatigue, anorexia, abdominal pain, weight loss, orthostatic hypotension, and salt craving. Characteristic hyperpigmentation is seen in patients with primary adrenal failure [37]. This disease has been reported in three forms [7, 9, 37]:

*Pathology Associated with Hormones of Adrenal Cortex DOI: http://dx.doi.org/10.5772/intechopen.84815*


Most of the cases (80–90%) of primary adrenal insufficiency are caused by autoimmune adrenalitis. Most of the cases fall under the autoimmune polyendocrinopathy syndrome (60%) [1, 2, 19, 32–34]*.* Cell-mediated immune mechanisms are implicated in pathogenesis. Various antibodies have been identified, antibodies against steroid 21-hydroxylase (85% cases) and autoantigens like steroid 17α-hydroxylase and cholesterol side-chain cleavage enzyme. Other associations include cytotoxic T-lymphocyte antigen 4, protein tyrosine-phosphatase nonreceptor type 22, and the MHC class II transactivator. Secondary adrenal insufficiency results from any process that involves the pituitary gland and interferes with corticotropin secretion. Tertiary adrenal insufficiency results from processes that involve the hypothalamus and interfere with secretion of corticotropin-releasing hormone, arginine vasopressin, or both. Suppression of the hypothalamic–pituitary–adrenal (HPA) axis by long-term administration of high doses of glucocorticoids is the most common cause [9, 36, 38].

#### **5.2 Laboratory investigations**

The patients of AI usually present with hyponatremia and hyperkalemia due to decreased aldosterone. Hypoglycemia also occurs due to cortisol. Decreased levels of this hormone also lead to an increase in lymphocytes and eosinophils, as a result of decreased immune-modulatory action of hydrocortisone. Measurement of baseline cortisol levels between 8:00 and 9:00 AM is the test used to diagnose AI. A serum cortisol level of value less than 5 μg/mL favors diagnosis of AI. Stimulation test with cosyntropin which stimulates the cortex helps in differentiating primary and secondary AI. In this test 250 μg of cosyntropin is administered intramuscularly or intravenously, and serum cortisol is measured 30 min after infusion. Serum cortisol value of ≥18 μg/dL indicates a normal response. A cortisol peak <18 μg/dL confirms the diagnosis of AI. Serum cortisol level ≥ 100 pg/mL confirms the diagnosis of Addison's syndrome. Serum cortisol value of <10 pg/mL confirms diagnosis of secondary AI [35–39].

#### **6. Adrenocortical carcinoma**

#### **6.1 Definition and introduction**

ACC is a highly aggressive and a very rare malignancy. The incidence of this malignancy is approximately 0.72 per million cases per year according to the study by Surveillance, Epidemiology, and End Results (SEER) database [39]. The median age of diagnosis is usually fifth to sixth decade; however the German ACC Registry reports a median age at diagnosis of 46 years with a predilection for the female gender (female to male ratio: 1.5–2.5:1) [35, 38, 39].

Adrenocortical carcinomas (ACC) are rare tumors with an estimated annual inci¬dence of 0.7–2 cases by year and a global prevalence of 4–12 cases per million with a 5-year survival rate inferior to 35% in most of the studies published.

#### **6.2 Pathogenesis of adrenocortical carcinoma**

Various mutations have been implicated in association with ACC. Most common are germ-line *TP53* mutations, associated with childhood ACCs. The adult population shows a prevalence of 3–7% of similar mutation. Childhood ACC can be found in association with Li-Fraumeni syndrome, Beckwith-Wiedemann syndrome, Lynch syndrome, and multiple endocrine neoplasia type 1. Of late an association with familial adenomatous polyposis (FAP), neurofibromatosis type 1, Werner syndrome, and Carney complex has also been postulated [39, 40].

#### *6.2.1 Molecular mechanisms*

In ACCs, chromosomal gains were frequently observed in regions 4q, 4p16, 5p15, 5q12–13, 5q32-qter, 9q34, 12q13, 12q24, and 19p, and chromosomal losses were observed at 1p, 2q, 11q 17p, 22p, and 22q. Microsatellite studies identified frequent allelic losses in regions 17p13, 11q15, and 2p16 (85%, 92%, and 90% of samples, respectively) [41–43].

Signaling pathways involved in adrenal malignant carcinogenesis [44–47]:


#### *6.2.1.1 The TP53 alterations*

Protein p53, "guardian of genome," is located at the 17p13 locus, and alterations in this gene have been noticed in various cancers including adrenocortical carcinoma, more so at the somatic level. p53 gene mediates cellular response to stress, and adult sporadic ACCs usually reveal loss of heterozygosity at this locus (nearly 85%) [48]. Stress leads to inhibition of degradation of p53 by E3 ubiquitin ligase MDM2, leading to inhibition of cell cycle arrest in response to DNA damage as well as apoptosis. These tumors tend to be larger and present at more advanced stage of tumor progression with shorter disease-free survival. Various genetic alterations have been reported in patients with adrenal cortex carcinoma like loss of *PTTG1* has been reported in nearly 84%, mutation in retinoblastoma protein (pRb) in nearly 27% cases and mutation in RB1 gene in 7% of the cases. Inactivating mutations or homozygous deletions of CDKN2A have also been reported in 11–16% cases. High-level amplifications of CDK4 and MDM2 were reported in 2–7% ACCs [49–51]. It is surprising to see that majority of the TP53 mutations occur at the DNA-binding domain. Some tumors also have shown abnormalities in genes that encode for negative regulators of TP53, like PTTG1 which encodes for securin, noted in 84% of ACC. It is considered as a marker of poor survival [44, 46, 49].

#### *6.2.1.2 Wnt/beta-catenin signaling pathway*

Wnt family consists of highly conserved growth factors having similar amino acid sequences and is responsible for various developmental and homeostatic processes [4, 44, 46–48, 51]. A prevalence of 39 and 84% has been reported by various authors on immunohistochemistry for β-catenin. The Wnt receptor is composed of members of the frizzled family and low-density lipoprotein receptor-related

*Pathology Associated with Hormones of Adrenal Cortex DOI: http://dx.doi.org/10.5772/intechopen.84815*

protein. β-Catenin accumulates in the cytoplasm and gets translocated into the nucleus and then binds with Wnt receptor leading to inhibition of the axin-adenomatous polyposis coli—glycogen synthase kinase 3β (GSK-3) complex. This blocks the phosphorylation of β-catenin, leading to increased accumulation of β-catenin in the cytoplasm which further translocates into the nucleus. Interaction between β-catenin with the T cell-specific transcription factor/lymphoid enhancer-binding factor-1 family of transcription factors occurs in the nucleus, thus regulating transcription of Wnt target genes. If Wnt stimulation of GSK-3 phosphorylating β-catenin does not occur, degradation by proteosomes occurs following ubiquitylation of this receptor. Wnt pathway has been implicated in patients with familial adenomatous polyposis and in the development of colorectal carcinomas as well as ACCs. Wnt/beta-catenin pathway can be activated in both benign and malignant tumors by *CTNNB1* mutations and by *ZNRF3* inactivation in adrenal cancer. *ZNRF3 is a recent gene that* encodes a cell-surface transmembrane E3 ubiquitin ligase which acts as a negative feedback regulator of Wnt signaling. Recently, *ZNRF3* was found to be the most frequently altered gene in study cohorts of ACC investigated by integrated genomics, with a prevalence of 21 and 19% in studies by Assié et al. and Zheng et al., respectively [50, 51].

#### *6.2.1.3 Insulin growth factor II (IGF-II)*

Nearly 85–90% of the adult adrenocortical carcinomas are attributed to *IGF-II* overexpression. This molecular abnormality is associated with DNA demethylation at *IGF-II* locus in most of cases. Various transcriptome studies have confirmed that *IGF-II* is the most upregulated gene in ACC [52, 53].

#### *6.2.2 Biochemistry*

ACC are the tumors characterized by adrenocortical hormone production in nearly 45–70% of patients. Hypercortisolism is the most common presentation of patients presenting with hormone excess leading to a plethora of symptoms like diabetes mellitus, hypertension, hypokalemia, muscle weakness/atrophy, and osteoporosis [40–43]. Excess of androgens which comprise nearly 40–60% of hormone-secreting ACCs can cause rapid-onset male pattern baldness, hirsutism, virilization, and menstrual irregularities in women. Estrogen production occurs in 1–3% of male ACC patients, causing gynecomastia and testicular atrophy (through suppression of the gonadal axis). In the evaluation of adrenal tumors, regardless of size, androgen or estrogen production should always raise the suspicion of a malignant tumor [44].

#### *6.2.3 Gross findings*

ACCs are generally large tumors, measuring on average 10–13 cm. Only a minority of tumors are less than 6 cm (9–14%), with only 3% presenting as lesions less than 4 cm [2, 3, 6, 9, 35].

#### *6.2.4 Microscopy findings*

Microscopically these tumors have variable architectural patterns. The tumor cells are arranged in a trabecular, alveolar, or diffuse pattern. Occasionally mixed patterns are also noted. Some areas may also exhibit free-floating tumor cells forming balls [2, 3, 6, 9, 35] (**Figure 2A, B**).

#### **Figure 2.**

*(A) Section from a 45-year-old patient, presented with a 13 cm mass in the left adrenal gland. Histology reveals clusters of cells having anisocytosis and enlarged nuclei with prominent nucleoli. The fair number of darkly stained atypical mitosis is also evident (hematoxylin and eosin stain, ×200). (B) Histology reveals clusters of cells having anisocytosis and enlarged nuclei with prominent nucleoli. The cells are separated by myxoid stroma. The fair number of darkly stained atypical mitosis is also evident (hematoxylin and eosin stain, ×400).*

Histologic criteria for malignancy in adrenal cortical tumors are assessed as follows [2, 3, 6, 9, 35, 43–45]:


Weiss et al. proposed a scoring system which was further modified and is widely accepted to report adrenal cortex carcinomas. These criteria include [35, 43–45, 54] (**Table 3**).

#### *6.2.5 Interpretation*

Adrenal cortical adenoma: total score < 3. Adrenal cortical carcinoma: total score ≥ 3. Thus if the modified Weiss score is ≥ 3, then a diagnosis of adrenocortical carcinoma is given.


#### **Table 3.**

*Weiss scoring for adrenocortical carcinoma.*


#### **Table 4.**

*Differentiating features between adrenocortical adenoma and adrenocortical carcinoma.*

However there are other features that may help in differentiating between adenomas and carcinoma. These are listed in **Table 4** [35, 43–45].

#### **7. Conclusion**

Adrenal glands have an essential role in maintaining the normal hemostasis. However the three layers of adrenal cortex, the zona glomerulosa, zona fasciculata, and zona reticularis, secrete essential hormones that are involved in fluid and electrolyte balance, regulating renin-angiotensin-aldosterone system, production of glucocorticoids, and synthesis of sex hormones. These hormones play an important

#### *Mitochondria and Brain Disorders*

role in maintaining the normal homeostasis of the body. Various lesions in adrenal, benign as well as malignant, are known to cause disturbances in the internal milieu of our body. It is therefore essential to know the physiology as well as various types of disorders that can be encountered so as to define proper management of the patient. Also lesions of adrenal gland are attributed to various genetic abnormalities, knowledge of which can be implicated to study the pathogenesis and in applying this knowledge in prognosis as well as developing targeted therapy for these lesions.

#### **Author details**

Lovelesh K. Nigam1 \*, Aruna V. Vanikar1,2, Rashmi D. Patel1 , Kamal V. Kanodia1 and Kamlesh S. Suthar1

1 Department of Pathology, Laboratory Medicine, Transfusion Services and Immunohematology, G.R. Doshi and K.M. Mehta Institute of Kidney Diseases and Research Centre and Dr. H.L. Trivedi Institute of Transplantation Sciences, Civil Hospital Campus, Asarwa, Ahmedabad, India

2 Department of Cell Therapy and Regenerative Medicine, Asarwa, Ahmedabad, India

\*Address all correspondence to: drloveleshnigam@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.

*Pathology Associated with Hormones of Adrenal Cortex DOI: http://dx.doi.org/10.5772/intechopen.84815*

#### **References**

[1] Tischler AS. Paraganglia. In: Mills SE, editor. Histology for Pathologists. 3rd ed. Philadelphia: Lippincott Williams & Wilkins; 2012. pp. 1277-1294

[2] Zhang R, Lloyd RV. Chapter 2: The pathology of adrenal masses. In: Kebebew E, editor. Management of Adrenal Masses in Children and Adults. Cham: Springer; 2017. DOI: 10.1007/978-3-319-44136-8\_2

[3] Mangray S, De Lellis R. Adrenal glands. In: Mills SE, editor. Sternberg's Diagnostic Surgical Pathology. Philadelphia: Wolters Kluwer; 2015. pp. 585-646

[4] Else T, Hammer GD, McPhee SJ. Chapter 21: Disorders of the adrenal cortex. In: Pathophysiology of Disease: An Introduction to Clinical Medicine. 7th ed. USA: McGraw Hill Education; 2014. pp. 593-624

[5] Silverman ML, Lee AK. Anatomy and pathology of the adrenal glands. The Urologic Clinics of North America. 1989;**16**(3):417-432. PMID: 2665268

[6] Hughes S, Lynn J. Chapter 37: Surgical anatomy and surgery of the adrenal glands. In: Surgical Endocrinology. Philadelphia: Elsevier Ltd; 1993. pp. 458-467. DOI: 10.1016/ B978-0-7506-1390-.50042-8

[7] Zynger DL. Chapter 10: Adrenal Primary Tumors and non-tumors. In: Epstein JI, editor. Biopsy Interpretation of the Kidney and Adrenal Gland. 1st ed. Philadelphia: Wolters Kluwer; 2016. pp. 173-186

[8] Innes JA, Maxwell S. Chapter 10: Endocrine disease. In: Davidson's Essentials of Medicine. 23rd edition, Philadelphia: Elsevier; 2016. pp. 327-380

[9] Rosai J. Chapter 16: Adrenal gland and other paraganglia. In: Rosai and

Ackerman's Surgical Pathology. 10th ed. Vol. 2. Philadelphia: Elsevier; 2012. pp. 1057-1100

[10] Arlt W. Disorders of the adrenal cortex. In: Kasper DL, Hauser SL, Jameson JL, Fauci AS, Longo DL, Loscalzo, editors. Harrison's Principles of Internal Medicine. 19th ed. Vol. 2. USA: McGraw Hill Education; 2015. pp. 2309-2330

[11] Agrons MM, Jensen CT, Habra MA, Menias CO, Shaaban AM, Wagner-Bartak NA, et al. Adrenal cortical hyperplasia: Diagnostic workup, subtypes, imaging features and mimics. The British Journal of Radiology. 2017;**90**:20170330

[12] MvNicol AM. Lesions of adrenal cortex. Archives of Pathology & Laboratory Medicine. 2008;**132**:1263-1269

[13] Nieman L, Ilias I. Evaluation and treatment of Cushing's syndrome. The American Journal of Medicine. 2005;**118**:1340-1346.

[14] Ejaz S, Vassilopoulou-Sellin R, Busaidy NL, Hu MI, Waguespack SG, Jimenez C, et al. Cushing syndrome secondary to ectopic adrenocorticotropic hormone secretion. Cancer. 2011;**117**:4381-4389. DOI: 10.1002/ cncr.26029

[15] Chaudhary V, Bano S. Imaging of the pituitary: Recent advances. Indian Journal of Endocrinology and Metabolism. 2011;**15**(Suppl. 3):216- S223. DOI: 10.4103/2230-8210.84871

[16] Lila AR, Sarathi V, Jagtap VS, Bandgar T, Menon P, Shah NS. Cushing's syndrome: Stepwise approach to diagnosis. Indian Journal of Endocrinology and Metabolism. 2011;**15**:317-321

[17] Elamin MB, Murad MH, Mullan R, Erickson D, Harris K, Nadeem S. Accuracy of diagnostic tests for Cushing's syndrome a systematic review and meta-analyses. The Journal of Clinical Endocrinology and Metabolism. 2008;**93**:1553-1562

[18] Al-Saadi N, Diederich S, Oelkers W. A very high dose dexamethasone suppression test for differential diagnosis of Cushing's syndrome. Clinical Endocrinology. 1998;**48**(1):45-51

[19] Raff H, Sharma ST, Nieman LK. Physiological basis for the etiology, diagnosis, and treatment of adrenal disorders: Cushing's syndrome, adrenal insufficiency, and congenital adrenal hyperplasia. Comprehensive Physiology. 2014;**4**(2):739-769. DOI: 10.1002/cphy. c130035

[20] Manipadam MT, Abraham R, Sen S, Simon A. Primary pigmented nodular adrenocortical disease. Journal of Indian Association of Pediatric Surgeons. 2011;**16**:160-162. DOI: 10.4103/0971-9261.86881

[21] Pernick N. Cushing Syndrome. Available from: http://www. pathologyoutlines.com/topic/ adrenalcushings.html [Accessed: 22 December 2018]

[22] Katanić D, Kafka D, Živojinov M, Vlaški J, Budakov Z, Pogančev MK, et al. Primary pigmented nodular adrenocortical disease: Literature review and case report of a 6-year-old boy. Journal of Pediatric Endocrinology & Metabolism. 2017;**30**(5):603-609. DOI: 10.1515/jpem-2016-0249

[23] Ngow HA, Khairin WM. Primary pigmented nodular adrenocortical disease. Endokrynologia Polska (Polish Journal of Endocrinology). 2011;**62**(3):268-270

[24] Bain J. Carney's complex. Mayo Clinic Proceedings. 1986;**61**:508. DOI: 10.1016/S0025-6196(12)61989-2

[25] Swain JM, Grant CS, Schlinkert RT, Thompson GB, vanHeerden JA, Lloyd RV, et al. Corticotropin-independent macronodular adrenal hyperplasia. Archives of Surgery. 1998;**133**:541-546

[26] New MI, Wilson RC. Steroid disorders in children: Congenital adrenal hyperplasia and apparent mineralocorticoid excess. Proceedings of the National Academy of Sciences of the United States of America. 1999;**96**:12790-12797. DOI: 10.1073/ pnas.96.22.12790

[27] Chung EM, Biko DM, Schroeder JW, Cube R, Conran RM. From the radiologic pathology archives: Precocious puberty: Radiologic-pathologic correlation. RadioGraphics. 2012;**32**:2071-2099. DOI: 10.1148/rg.327125146

[28] Witchel SF. Congenital adrenal hyperplasia. Journal of Pediatric and Adolescent Gynecology. 2017;**30**(5):520-534

[29] El Maouche D, Arlt W, Merke DP. Congenital adrenal hyperplasia. The Lancet. 2017;**390**(10108):2121-2241

[30] Speiser PW, White PC. Congenital adrenal hyperplasia. The New England Journal of Medicine. 2003;**349**(8): 776-788. PMID: 12930931

[31] Åkerström G, Hellman P. Genetic syndromes associated with adrenal tumors. In: Linos D, van Heerden JA, editors. Adrenal Glands. Berlin, Heidelberg: Springer; 2005. DOI: 10.1007/3-540-26861-8\_25

[32] Mattson C, Young WF. Primary aldosteronism: Diagnostic and treatment strategies. Nature Clinical Practice. Nephrology. 2006;**2**(4):198-208

*Pathology Associated with Hormones of Adrenal Cortex DOI: http://dx.doi.org/10.5772/intechopen.84815*

[33] Ganguly A. Primary aldosteronism. The New England Journal of Medicine. 1998;**339**(25):1828-1834

[34] Umakoshi H, Tsuiki M, Yokomoto Umakoshi M, Takeda Y, Kurihara I, et al. Correlation between lateralization index of adrenal venous sampling and standardized outcome in primary aldosteronism. Journal of the Endocrine Society. 2018;**2**(8):893-902. DOI: 10.1210/js.2018-00055

[35] Lack EE, Wieneke J. Chapter 19: Tumors of the adrenal gland. In: Fletcher C, editor. Diagnostic Histopathology of Tumors. 4th ed. Vol. 2. Philadelphia: Elsevier Saunders; 2013. pp. 1294-1352

[36] Fares AB, dos Santos RA. Conduct protocol in emergency: Acute adrenal insufficiency. Revista da Associação Médica Brasileira. 2016;**62**(8):728-734. DOI: 10.1590/1806-9282.62.08.728

[37] Addison T. On the Constitutional and Local Effects of Disease of the Supra-Renal Capsules. London: Samuel Highley; 1855

[38] Betterle C, Morlin L. Autoimmune Addison's disease. Endocrine Development. 2011;**20**:161-172

[39] Reiff E, Duh QY, Clark OH, McMillan A. Extent of disease at presentation and outcome for adrenocortical carcinoma: Have we made progress? World Journal of Surgery. 2006;**30**:872-878

[40] Lack EE. Tumors of the adrenal gland and extra-adrenal paraganglia. In: Atlas of Tumor Pathology—3rd Series. Washington, DC: Armed Forces Institute of Pathology; 1997. pp. 102-104

[41] Aubert S, Wacrenier A, Leroy X, Devos P, Carnaille B, Proye C, et al. Weiss system revisited: A clinicopathologic and immunohistochemical study

of 49 adrenocortical tumors. The American Journal of Surgical Pathology. 2002;**26**(12):1612-1619

[42] Stojadinovic A, Brennan MF, Hoos A, Omeroglu A, Leung DNY, Dudas ME, et al. Adrenocortical adenoma and carcinoma: Histopathological and molecular comparative analysis. Modern Pathology. 2003;**16**(8):742-751. DOI: 10.1097/01.MP.0000081730.72305.81

[43] Giordano TJ. Adrenocortical tumors: An integrated clinical, pathologic, and molecular approach at the University of Michigan. Archives of Pathology & Laboratory Medicine. 2010;**134**:1440-1443

[44] Bonnet-Serrano F, Bertherat J. Genetics of tumors of the adrenal cortex. Endocrine-Related Cancer. 2018;**25**:131-152

[45] Soon PSH, Mcdonald KL, Robinson BG, Sidhu SB. Molecular markers and the pathogenesis of adrenocortical cancer. The Oncologist. 2008;**13**:548-561

[46] Lerario AM. Genetics and epigenetics of adrenocortical tumors. Molecular and Cellular Endocrinology. 2014;**386**:67-84. DOI: 10.1016/j. mce.2013.10.028

[47] Else T, Kim AC, Sabolch A, Raymond VM, Kandathil A, Caoili EM, et al. Adrenocortical carcinoma. Endocrine Reviews. 2014;**35**(2):282-326

[48] Gicquel C, Bertagna X, Gaston V, Coste J, Louvel A, Baudin E, et al. Molecular markers and long-term recurrences in a large cohort of patients with sporadic adrenocortical tumors. Cancer Research. 2001;**61**:6762-6767

[49] Ragazzon B, Libé R, Assié G, Tissier F, Barreau O, Houdayer C, et al. Mass-array screening of frequent

#### *Mitochondria and Brain Disorders*

mutations in cancers reveals RB1 alterations in aggressive adrenocortical carcinomas. European Journal of Endocrinology. 2014;**170**:385-391. DOI: 10.1530/EJE-13-0778

[50] Assié G, Letouzé E, Fassnacht M, Jouinot A, Luscap W, Barreau O, et al. Integrated genomic characterization of adrenocortical carcinoma. Nature Genetics. 2014;**46**:607-612. DOI: 10.1038/ng.2953

[51] Zheng S, Cherniack AD, Dewal N, Moffitt RA, Danilova L, Murray BA, et al. Comprehensive pan-genomic characterization of adrenocortical carcinoma. Cancer Cell. 2016;**29**: 723-736. DOI: 10.1016/j. ccell.2016.04.002

[52] De Reyniès A, Assié G, Rickman DS, Tissier F, Groussin L, René-Corail F, et al. Gene expression profiling reveals a new classification of adrenocortical tumors and identifies molecular predictors of malignancy and survival. Journal of Clinical Oncology. 2009;**27**:1108-1115. DOI: 10.1200/ JCO.2008.18.5678

[53] Else T. Association of adrenocortical carcinoma with familial cancer susceptibility syndromes. Molecular and Cellular Endocrinology. 2012;**351**(1): 66-70. DOI: 10.1016/j.mce.2011.12.008

[54] Scarpellia M, Algabab F, Kirkalic Z, Poppeld HV. Handling and pathology reporting of adrenal gland specimens. European Urology. 2004;**45**:722-729

Section 3
