**4. Plant-derived molecules as emerging therapies for arthritis**

Current arthritis treatments result in unwanted side effects and tend to be expensive, and natural products devoid of such disadvantages offer a novel opportunity. The use of natural products represents a promising alternative to treat rheumatic diseases, in particular by acting as therapeutic adjuvants to reduce the daily doses of conventional drugs that RA patients administer [148–150]. In this section, we highlight future perspectives in the treatment of RA with natural compounds, mainly herbal compounds, to minimize the harmful effects of the over-activation of neutrophils.

Decreased inflammation and joint destruction have been directly correlated with reduced neutrophil influx into the joints, as observed in mouse models by means of antibody blockade or the gene deletion of chemoattractant receptors such as CXCR1, CXCR2, and BLT1 (LTB<sup>4</sup> receptor) [15, 79]. The prospect of new drugs obtained from herbal products (or from structures of herbal products) plays a compelling role in drug discovery and development [151].

As previously mentioned, pharmacologic treatment options for arthritis are diverse and present several side effects. Furthermore, the high costs and increased risk of malignancies limit the use of such agents. Because of these limitations, there is a growing interest in the use of natural products as therapies or adjunct therapies [22]. Plant-derived products such as polyphenols, sesquiterpenes, flavonoids, and tetranortriterpenoids, which are herbal metabolites, are considered to have potential activity to block inflammation, and they may provide new therapeutic agents and cost-effective treatments [22, 23]. These natural products have attracted considerable interest over the past decade because of their multiple beneficial effects, such as their antioxidant, anti-inflammatory, antiproliferative, and immunomodulatory properties. In this section, we discuss the plant-derived products that have been most studied in RA experimental models and/or clinical trials (**Table 2**).

#### **4.1. Quercetin**

Quercetin (**Figure 2a**) is the major dietary flavonol found in fruits, vegetables, and beverages, such as tea and red wine [152]. Several epidemiological and experimental studies support the antioxidant, anti-inflammatory, antiangiogenic, antiproliferative, and proapoptotic effects of this molecule [153–155]. Preclinical studies on primary cells and animal models, as


**Table 2.** Herbal products that exhibit anti-arthritic potential in animal models.

Neutrophils in Rheumatoid Arthritis: A Target for Discovering New Therapies Based on Natural Products http://dx.doi.org/10.5772/intechopen.68617 99

**Figure 2.** Chemical structure of (a) quercetin, (b) methyl gallate, (c) gedunin, (d) epigallocatechin gallate, and (e) curcumin.

well as clinical studies, suggest an inhibitory action of quercetin in RA. Quercetin has been reported to lower the levels of IL-1β, C-reactive protein, and monocyte chemotactic protein-1 (MCP-1), and restore plasma antioxidant capacity. In addition, quercetin increased the expression of hemeoxygenase-1 in the joints of arthritic rats. Finally, quercetin inhibited the twofold increase in NF-κB activity observed in joints after arthritis induction [156].

There are divergent data on the effect of quercetin in neutrophils. For instance, *in vitro*, quercetin inhibited myeloperoxidase activity [157] but had no effect on lipopolysaccharide-induced neutrophil surface expression of the adhesion molecules L-selectin (CD62L) and β2 integrin (CD11b/Mac1), [158] which are related to rolling and firm adhesion, respectively [159]. In paw edema induced by carrageen, quercetin did not inhibit the increase in myeloperoxidase, which is used as a marker of neutrophil recruitment [160]. Therefore, it seems unlikely that quercetin would inhibit neutrophil recruitment [158]. On the other hand, quercetin inhibits the fMLPinduced increase in intracellular calcium, [158] which is necessary for actin polymerization and consequently neutrophil migration [159]. In addition, *in vitro*, quercetin blocked human neutrophil mobilization through the inhibition of the cellular signaling responsible for actin polymerization in association with the down-regulation of adhesion molecules [161], indicating that treatment with this flavonoid is a conceivable approach to control excessive neutrophil recruitment during inflammation and to prevent neutrophil-mediated tissue lesions [162] (**Table 3**).

#### **4.2.** *Schinus terebinthifolius* **and methyl gallate**

**4. Plant-derived molecules as emerging therapies for arthritis**

over-activation of neutrophils.

98 Role of Neutrophils in Disease Pathogenesis

**4.1. Quercetin**

experimental models and/or clinical trials (**Table 2**).

**Compound Chemical class Arthritis experimental** 

**Table 2.** Herbal products that exhibit anti-arthritic potential in animal models.

Quercetin Flavonoid Adjuvant-induced arthritis [156] Methyl gallate Polyphenol Zymosan-induced arthritis [171] Gedunin Tetranortriterpenoid Zymosan-induced arthritis [176] Epigallocatechin gallate Polyphenol Collagen-induced arthritis [179] Curcumin Polyphenol Collagen-induced arthritis [191]

Current arthritis treatments result in unwanted side effects and tend to be expensive, and natural products devoid of such disadvantages offer a novel opportunity. The use of natural products represents a promising alternative to treat rheumatic diseases, in particular by acting as therapeutic adjuvants to reduce the daily doses of conventional drugs that RA patients administer [148–150]. In this section, we highlight future perspectives in the treatment of RA with natural compounds, mainly herbal compounds, to minimize the harmful effects of the

Decreased inflammation and joint destruction have been directly correlated with reduced neutrophil influx into the joints, as observed in mouse models by means of antibody blockade or the gene deletion of chemoattractant receptors such as CXCR1, CXCR2, and BLT1 (LTB<sup>4</sup> receptor) [15, 79]. The prospect of new drugs obtained from herbal products (or from structures of herbal products) plays a compelling role in drug discovery and development [151]. As previously mentioned, pharmacologic treatment options for arthritis are diverse and present several side effects. Furthermore, the high costs and increased risk of malignancies limit the use of such agents. Because of these limitations, there is a growing interest in the use of natural products as therapies or adjunct therapies [22]. Plant-derived products such as polyphenols, sesquiterpenes, flavonoids, and tetranortriterpenoids, which are herbal metabolites, are considered to have potential activity to block inflammation, and they may provide new therapeutic agents and cost-effective treatments [22, 23]. These natural products have attracted considerable interest over the past decade because of their multiple beneficial effects, such as their antioxidant, anti-inflammatory, antiproliferative, and immunomodulatory properties. In this section, we discuss the plant-derived products that have been most studied in RA

Quercetin (**Figure 2a**) is the major dietary flavonol found in fruits, vegetables, and beverages, such as tea and red wine [152]. Several epidemiological and experimental studies support the antioxidant, anti-inflammatory, antiangiogenic, antiproliferative, and proapoptotic effects of this molecule [153–155]. Preclinical studies on primary cells and animal models, as

**model**

**Reference**

*S. terebinthifolius* Raddi (Anacardiaceae) is a native plant from South America. It has been used in folk medicine as teas, infusions, or tinctures, as an anti-inflammatory, febrifuge, analgesic,


**Table 3.** Major molecular targets and anti-arthritic mechanisms of herbal products.

and depurative agent and to treat urogenital system illnesses [163]. Scientific reports demonstrated that *S. terebinthifolius* extracts and fractions are rich in polyphenols and display antioxidant, antibacterial, and antiallergic properties in different experimental models [164–166]. The HPLH chromatograms of hydroalcoholic extracts from *S. terebinthifolius* leaves (ST-70) reveal that methyl gallate (MG, **Figure 2b**) is one of the major polyphenol components of the ST-70 extract [167]. Methyl gallate has been extensively studied because of its antioxidant, antitumor, and antimicrobial activities [168–170]. Pharmacological studies have shown that ST-70 and MG also have an anti-inflammatory effect and may have potential activity against arthritis. Pretreatment with ST-70 or MG markedly reduced knee-joint thickness, total leukocyte (mainly neutrophil) infiltration, and reduced the production of inflammatory mediators associated with arthritis such as CXCL-1/KC, IL-6, TNF-α, IL-1β, LTB4, and PGE<sup>2</sup> . ST-70 and MG also inhibited murine neutrophil chemotaxis induced by CXCL-1/KC *in vitro*, and MG impaired the adhesion of these cells to TNF-α-primed endothelial cells [167, 171]. These results provide some evidence that MG inhibits neutrophil activation and adhesion molecules expression and consequently prevents the neutrophil entry into inflammatory sites (**Table 3**).

Moreover, unlike potassium diclofenac, the long-term oral administration of ST-70 does not induce lethality or gastric damage in mice, which suggests that ST-70 could be used to treat inflammatory conditions such as arthritis with less toxicity [167].

#### **4.3.** *Carapa guianensis* **and gedunin**

*C. guianensis* Aublet is a member of the Meliaceae family that is widely used in folk medicine in Brazil and other countries surrounding the Amazon rainforest [172]. Anti-inflammatory and analgesic activities are among the most remarkable properties attributed by ethnopharmacological research to the oil extracted from *C. guianensis* seeds, mainly for rheumatic pain and arthritis [172, 173]. *C. guianensis* oil and six different tetranortriterpenoids (TNTP) isolated from the oil were able to significantly inhibit zymosan-induced knee joint edema formation and protein extravasation. TNTP pretreatment inhibited the increase in total leukocyte and neutrophil numbers in the synovial fluid. TNTP also impaired the production of TNF-α, IL-1β, and CXCL-8/IL-8, and significantly inhibited the expression of the NF-κB p65 subunit [174].

Gedunin (**Figure 2c**) is a natural tetranortriterpenoid isolated from vegetal species of the Meliaceae family and is known to inhibit the stress-induced chaperone heat shock protein (Hsp) 90 [175]. Mouse pretreatment and posttreatment with gedunin impaired zymosaninduced edema formation and total leukocyte influx mainly due to the inhibition of neutrophil migration and reduced articular hypernociception. Gedunin also reduced the *in situ* expression of preproET-1 mRNA and IL-6, TNF-α, LTB<sup>4</sup> and PGE<sup>2</sup> production and prevented increases in the number of lipid bodies in synovial leukocytes [176]. Lipid bodies are important sites for the synthesis and storage of lipid mediators and they increase in number during inflammatory responses [177]. In neutrophils, gedunin impaired ET-1-induced shape changes, blocked ET-1- and LTB<sup>4</sup> -induced chemotaxis, decreased ET-1-induced lipid body formation and impaired neutrophil adhesion to TNF-α-primed endothelial cells [176]. The combined *in vitro* and *in vivo* effects of gedunin reveal its potential as an anti-arthritic candidate, especially its direct effect on key cells involved in articular inflammation such as neutrophils (**Table 3**).

### **4.4. Epigallocatechin gallate**

and depurative agent and to treat urogenital system illnesses [163]. Scientific reports demonstrated that *S. terebinthifolius* extracts and fractions are rich in polyphenols and display antioxidant, antibacterial, and antiallergic properties in different experimental models [164–166]. The HPLH chromatograms of hydroalcoholic extracts from *S. terebinthifolius* leaves (ST-70) reveal that methyl gallate (MG, **Figure 2b**) is one of the major polyphenol components of the ST-70 extract [167]. Methyl gallate has been extensively studied because of its antioxidant, antitumor, and antimicrobial activities [168–170]. Pharmacological studies have shown that ST-70 and MG also have an anti-inflammatory effect and may have potential activity against arthritis. Pretreatment with ST-70 or MG markedly reduced knee-joint thickness, total leukocyte (mainly neutrophil) infiltration, and reduced the production of inflammatory media-

**Compound Molecular targets/mechanisms Reference**

activity in joints

mobilization

formation

Epigallocatechin gallate Ameliorates the severity of arthritis

Curcumin Suppresses collagen-induced arthritis

**Table 3.** Major molecular targets and anti-arthritic mechanisms of herbal products.

and MCP-1 levels. Restores plasma antioxidant capacity, increases HO-1 expression, and inhibits NF-κB

[156, 157, 161]

[171]

[176]

[179–184, 186–189]

[191, 193]

. Affects

Inhibits myeloperoxidase activity in neutrophils and blocks neutrophil

leukocyte accumulation, neutrophil migration and IL-6, TNF-α, CXCL-1, IL-1β, LTB4, and PGE2 production in zymosan-induced arthritis. Impairs neutrophil chemotaxis and adhesion

articular edema, neutrophil migration, hypernociception, and the production of IL-6, TNF-α, LTB4, and PGE2 and prevents increases in lipid bodies. Decreases neutrophil shape changes, chemotaxis, and lipid body

and regulates the expression of cytokines, chemokines, MMPs, ROS, NO, COX-2, and PGE<sup>2</sup>

by reducing cellular infiltration, synovial hyperplasia, cartilage destruction, and bone erosion. Blocks

neutrophil recruitment

neutrophil functionality and inhibits IL-8 and MIP-3α expression

Quercetin Inhibits IL-1β, C-reactive protein,

100 Role of Neutrophils in Disease Pathogenesis

Methyl gallate Reduces edema formation, total

Gedunin Attenuates zymosan-induced

tors associated with arthritis such as CXCL-1/KC, IL-6, TNF-α, IL-1β, LTB4, and PGE<sup>2</sup>

and MG also inhibited murine neutrophil chemotaxis induced by CXCL-1/KC *in vitro*, and

. ST-70

Epigallocatechin gallate (EGCG, **Figure 2d**) is one of the main components of green tea [178]. It has antioxidative, anti-inflammatory, antitumor, and chemopreventive properties. The potential disease-modifying effects of green tea on arthritis have been reported; for example, in a mouse model of RA, the induction and severity of arthritis was ameliorated by the prophylactic administration of green tea polyphenols [179]. Subsequent studies suggested that EGCG possesses remarkable potential to prevent chronic diseases like OA and RA [180–184]. The anti-inflammatory and anti-arthritic effects of EGCG are supported by *in vitro* and *in vivo* data indicating that EGCG can regulate the expression of cytokines, chemokines, MMPs, ROS, nitric oxide (NO), COX-2, and PGE<sup>2</sup> in cell types relevant to the pathogenesis of RA [179–184]. In *in vivo* studies, EGCG was found to inhibit inflammation in mouse models by affecting the functioning of T cells and neutrophils [185, 186]. IL-8 is the most powerful chemo-attractant for neutrophils in the target tissue. EGCG is a very effective inhibitor of IL-1β and of TNF-α-induced IL-8 and macrophage-inflammatory protein-3α (MIP-3α) expression in different cell types [187–189]. These *in vitro* and *in vivo* observations indicated the efficacy of EGCG and demonstrate that it can modulate multiple signal transduction pathways in a fashion that suppresses the expression of inflammatory mediators that play a role in the pathogenesis of arthritis (**Table 3**).

#### **4.5. Curcumin**

Curcumin (**Figure 2e**) is a yellow-colored polyphenol found in the rhizome of turmeric. It has antioxidant, anti-inflammatory, antiapoptotic, and anticarcinogenic properties [190]. Oral administration of curcumin suppressed type II collagen-induced arthritis (CIA) in mice by reducing cellular infiltration, synovial hyperplasia, cartilage destruction, and bone erosion. Moreover, the production of MMP-1 and MMP-3 was inhibited by curcumin in CIA and in TNF-α-stimulated RA fibroblast-like synoviocytes (RA-FLS) and chondrocytes [191].

*In vitro*, it has been reported that curcumin decreases IL-1β-induced expression of the pro-inflammatory cytokine IL-6 and vascular endothelial growth factor (VEGF) in RA-FLS [192]. In addition, curcumin blocks neutrophil recruitment through the inhibition of cellular signaling responsible for actin polymerization in association with the down-regulation of adhesion molecules [193]. It has also been shown to induce apoptosis of RA-FLS (which are resistant to apoptosis) by increasing the expression of the proapoptotic protein Bax and down-regulating the expression of the antiapoptotic protein Bcl-2 [190]. Some molecular mechanisms related to curcumin have been identified. In a human synovial fibroblast cell line (MH7A) stimulated with IL-1β, curcumin blocked the activation of the NF-κB pathway and induced deactivation of the ERK-1/2 pathway [192]. In addition, this polyphenol inhibited activating phosphorylation of protein kinase Cδ (PKCδ) in CIA, RA-FLS, and chondrocytes. Curcumin also suppressed JNK and c-Jun activation in those cells [191].

In a clinical trial with RA patients, curcumin reduced reported pain, tenderness, and swelling of joints [194]. A curcumin-based medicine, Meriva®, demonstrated efficacy in clinical trials with patients with osteoarthritis by reducing reported pain [195]. In another clinical trial, treatment with Meriva® reduced stiffness and physical signs of RA (treadmill test) along with IL-1, IL-6, and VCAM-1 production [196] (**Table 3**).

## **5. Conclusion**

In RA, neutrophils are key cells that are recognized to play an active role in orchestrating the progress of inflammation, through the release of pro-inflammatory cytokines, ROS, RNS, and NETs, which potentially affect the activities of both neutrophils and other cell types, such as resident mononuclear cells and chondrocytes. In addition, neutrophils participate in the cascade of events leading to mechanical hypernociception. Therefore, neutrophils participate in the pathogenesis of arthritis by promoting the inflammatory process, degradation of cartilage, and bone resorption. The modulation of neutrophil migration and functions in RA can be considered a potential target for pharmacological intervention in arthritis. The pharmacologic treatment options for arthritis are diverse. High costs and an increased risk of malignancies limit the use of these agents, in addition to the potential for side effects that all therapies possess. Nevertheless, herbal metabolites with anti-inflammatory activity and inhibitory action in neutrophils may provide new therapeutic agents and cost-effective treatments.
