Could Polyphenols Help in the Control of Rheumatoid Arthritis?

Rheumatoid arthritis (RA) is a chronic, systemic, joint-invading, autoimmune inflammatory disease, which causes joint cartilage breakdown and bone damage, resulting in functional impairment and deformation of the joints. The percentage of RA patients has been rising and RA represents a substantial burden for patients around the world. Despite the development of many RA therapies, because of the side effects and low effectiveness of conventional drugs, patients still need and researchers are seeking new therapeutic alternatives. Polyphenols extracted from natural products are effective on several inflammatory diseases, including RA. In this review polyphenols are classified into four types: flavonoids, phenolic acids, stilbenes and others, among which mainly flavonoids are discussed. Researchers have reported that anti-RA efficacies of polyphenols are based mainly on three mechanisms: their anti-inflammatory, antioxidant and apoptotic properties. The main RA factors modified by polyphenols are mitogen-activated protein kinase (MAPK), interleukin-1β (IL-1β), IL-6, tumor necrosis factor-α (TNF-α), nuclear factor κ light chain enhancer of activated B cells (NF-κB) and c-Jun N-terminal kinases (JNK). Polyphenols could be potent alternative RA therapies and sources for novel drugs for RA by affecting its key mechanisms.


Introduction
Rheumatoid arthritis (RA) is a notorious chronic autoimmune inflammatory joint disease, which can cause cartilage and bone damage [1]. This disease is characterized by synovial inflammation, swelling, autoantibody production, cartilage and bone destruction, and systemic features such as cardiovascular, pulmonary, and skeletal disorders. It is associated with progressive disability, systemic complications, early death and socioeconomic costs [2]. As of 2015 is estimated that RA affects about 24.5 million people [3]. This number includes 0.5 to 1% of adults in the developed world, 5 to 50 per 100,000 patients newly added each year [1]. Although the critical damage caused by this disease is well known and thus widely studied, the mechanism(s), underlying cause and pathway(s) of RA are not well-known.
The number of therapeutic solutions available for treating RA has continuously grown in the past 30 years. These solutions include non-steroidal anti-inflammatory drugs, glucocorticoids, disease-modifying anti-rheumatic drugs (DMARDs) of synthetic origin (e.g., methotrexate and c-Jun N-terminal kinase (JNK) inhibitors) and of biological origin (ex. tumor necrosis factor (TNF) inhibitors, interleukin (IL)-6 inhibitor, and B cell-depleting drugs) [4]. Recently, medications that suppress

Phenolic Acids
Hydroxybenzoic and hydroxycinnamic acids are characteristic phenolic acids. Phenolic acids account for about a third of the polyphenolic compounds in our diet and are found in all plant material, but they are particularly abundant in acidic-tasting fruits. Caffeic acid, gallic acid, and ferulic acid are some common phenolic acids. Phenolic acids showing anti-RA effects are arranged in Table 1. When monocyte and macrophage cells from rat were pre-exposed for 24 h to ferulic acid, which is found in grains, vegetables, fruits and nuts, nuclear factor of activated T cells c1 (NFATc1), c-Fos, NF-κB, tartrate-resistant acid phosphatase (TRAP), matrix metalloproteinases (MMP)-9, Cathepsin activities were depressed [10]. The natural polyphenol N-feruloylserotonin (N-f-5HT), extracted from Leuzea carthamoides, had RA-inhibitory effects via suppressing c-reactive protein (CRP), 12/15-lipoxygenase (LOX), TNF-α, inducible nitric oxide synthase (iNOS), IL-1β in liver and spleen cells of arthritic rats. This study was conducted for 28 days, with 3 mg/kg of N-f-5HT [11]. In the study of Lee, mRNA transcription of TNF-α was significantly attenuated in a human mast cell line (HMC-1) treated with gallotanin derived from Euphorbia [12]. Chlorogenic acid (CGA), derived from Gardenia jasminoides, inhibited the phosphorylation of p38, Akt, extracellular signal-regulated kinase (ERK) and IkB, also suppressed the mRNA expression of nuclear factor activated T cells cl (NFATcl). Furthermore, lipopolysaccharide (LPS)-induced bone erosion was alleviated in vivo when bone marrow macrophages (BMMs) were exposed to 10, 25, 50 µg/mM of CGA for 4 days [13]. p-Coumaric Acid (CA), which can be extracted from Gnetm cleistostachyum, was used in two studies. Both of them used the same dose of 100 mg/kg of CA to treat an adjuvant-induced arthritis (AIA) rat model. One trial with a duration of 8 days presented degradation of TNF-α and circulating immune complexes (CIC) levels while inducing alleviation of immunoglobulin G (IgG) [14]. In the other 16 day trial, CA treatment also reduced TNF-α activation, suggest an anti-RA effect via attenuation of cytokines, chemokines, osteoclastogenic factors, transcription factors, and mitogen-activated protein kinase (MAPK). In detail, the affected cytokines and chemokines are IL-1β, IL-6, monocyte chemoattractant protein (MCP)-1, the osteoclast factors are receptor activator of nuclear factor kappa-B ligand (RANKL), TRAP, the pro-inflammatory cytokines are IL-1b, IL-6, IL-17, the inflammatory enzymes are iNOS and cyclooxygenase (COX)-2, the transcription factors are NF-κB-p65, p-NF-κB-p65, NFATc-1, c-Fos, MAP kinases are JNK, p-JNK, ERK1/2. However, osteoprotegerin (OPG) elevation was shown [15].

Discussion
Rheumatoid arthritis (RA) is an autoimmune disease that induces chronic joint inflammation, which causes cartilage and bone damage [1]. Synovial inflammation, swelling, autoantibody production, cartilage and bone destruction, and systemic features such as cardiovascular, pulmonary, and skeletal disorders are the main symptoms of this disabling autoimmune disease. Currently, non-steroidal anti-inflammatory drugs (NSAIDS), glucocorticoids, DMARDs, immunosuppressants, and biologic agents have been used to treat this autoimmune disease. DMARDs, especially, has been acknowledged as an effective early intervention for RA, their efficacy being validated by several randomized trials [58,59]. However, each DMARD showed different toxicity that causes side-effects such as diarrhea and rashes [59], and therefore various studies have been conducted to find a better solution for RA treatment. We saw the potential of finding the solution in natural products, especially polyphenols.
Flavonoids demonstrate anti-RA effects through diverse mechanisms [24]. α-Glucosyl-hesperidin showed results in an animal model study, but it lacked a specific discussion on the mechanism of the effect [27]. A study on cocoa polyphenol extract (CPE) discusses in depth the effect of the compound on several different inflammatory routes such as VEGF regulation, PI3K-Akt, and MAPK pathways [29]. Epigallocatechin-3-gallate (EGCG), from green tea is the most widely studied polyphenol related to RA. Lee et al. conducted an in-depth study on this compound using CIA rats and found specific elements that regulate and are regulated by Th17 cells and p-STAT3. This study further has observed gene-level events, which showed that control of Nrf 2 gene may lead to anti-RA effects [32]. Yun et al., on the other hand, noted the mechanism of EGCG's effect from a different perspective, comprehensively focusing on MMP production via the MAPK and AP-1 pathways [31]. Morinobu et al. focused their study on the role of nuclear factor of activated T cells c1 (NF-ATc1) in EGCG's effect on osteoclasts [35]. On the other hand, a study by Leichsenring, et al. lacked a detailed discussion on the mechanism of EGCG [34]. A study of Oka, et al. on both EGCG and TFDG also gave an incomplete description of the mechanism of the compounds' anti-RA effects [36]. A study on a flavonol-rich residual layer of hexane fraction (RVHxR) gave a poor examination of the role of elements in the MAPK pathway of angiogenesis [37]. The study also descriptively illustrated the malvidin-3-O-β-glucoside-related RA pathogenic pathway targeting macrophages [42]. Thymoquinone (TQ) was studied systematically in two studies. Vaillancourt, et al conducted an in-depth study on TQ, setting three stages of experiments, which included in vivo experiments on human RAFLS and a rat model and in vitro tests on an animal model. This study observed the effect of TQ on RA based on three different pathogenic pathways, which are lipid peroxidation, inflammation, and bone destruction. Interactions of elements that comprise each pathway are described in detail through an organized experiment process [48]. Umar et al, suggested a new point of view in studying the effects of polyphenols on RA pathogenesis. The study focuses on the role of apoptosis signal-regulating kinase 1 (ASK1) in the TNF-α signaling pathway and explains the role of its related factors in RA reduction [46]. In a study by Tekeoglu et al. three different experimental groups and a control group were used, but the results was unhelpful in explaining the molecular mechanism of the regulating effect of TQ [47].
Phenolic acids, plant metabolites that are widely spread throughout the plant kingdom, also possess anti-RA effects. A study on ferulic acid made profound observations on the effect of the compound in the RA pathogenic pathway, especially targeting the relation of RANKL, an osteogenic factor, and NF-κB signaling pathway [10]. Kwaket al. conducted an incomplete study on chlorogenic acid. Considering that RA's pathological pathway contains various immunological factors, only using osteoclasts as the experiment cell line is limiting [13]. Neog et al. conducted a thoughtful study on the effect of p-coumaric acid (CA), also focusing on the system related to RANK and its interaction with T cell immune system factors [15]. Another study of CA, designed by Pragasam et al., was conducted under similar experimental conditions as used by Neoget al., but was imperfect in elucidating the molecular mechanism of CA's anti-Ra effect [14].
Stilbenes are polyphenols that have two phenyl moieties connected by a two-carbon methylene bridge. Most of the studies on stilbenes that showed anti-RA effects were made on resveratrol. Three noticeable studies were made on resveratrol. Tsai et al. particularly noted resveratrol's role in regulating COX-2 and PGE2 interaction. This study is unique because it focuses on the effect of particulate matter (PM) from air pollution on RA, and sees how resveratrol affects the inflammatory pathways of RA caused by PM [19]. Wahba et al. reported the effect of this compound from three perspectives. They observed immunological changes, inflammatory systemic changes, and oxidative stress changes.
Choosing specific biomarkers for each part, this study specifically elucidated the role of resveratrol in each pathway [23]. A study by Xuzhu et al. observed three different levels of the object, which included the CIA animal model, DLN cells, and Th17 cells. Despite the effort to observe the result in diverse ways, this study failed to identify the specific mechanism of resveratrol's effect on RA regulation at a molecular level [22].
In addition to flavonoids, phenolic acids, and stilbenes there are several other polyphenols that are hard to classify. Among them, curcumin (CM)-related molecules and emodin are the most actively studied polyphenols. Ramadan et al. systemically examined the anti-inflammatory and anti-oxidant effects of curcuminoids shrewdly considering diverse factors related to the pathways [51].
Kloesch et al. comprehensively tested the effect of CM on various inflammation pathway factors, but the duration of this study was too short [52]. A study by Zheng, et al. suggested a new way to increase the bioavailability of CA by forming CM-loaded Ns (CM-Ns). They also skillfully designed their experiments with three different experimental groups and one control group. However, a study on the molecular mechanism of CM-Ns' anti-Ra effect was lacking [54]. Park et al. conducted an n in depth study on how emodin targets the apoptosis pathway, mainly focusing on Bax/Bcl-2 imbalance and activation of caspase-9 and caspase-3 [53]. A study of Ha, et al. on emodin thoroughly studied different aspects of the RA pathogenic pathway and tested the compound in vivo, which signified its role in inflammatory conditions [57]. Zhu et al. further conducted an animal model study on the effect of emodin on RA symptoms, but their explanation on the therapeutic mechanism in the pathogenic pathway was deficient [56].
Polyphenol inhibit RA progress mainly by acting on three pathways: the inflammatory pathway, the oxidative pathway, and the apoptotic pathway. The inflammatory pathway regulated by polyphenols is mainly via the MAPK pathway and through regulation of NFATC1 gene in osteoblasts.
Although they are not mentioned frequently among the studies, the oxidative and apoptotic pathways are also attributed a role in the reduction of RA symptoms by polyphenols (Figure 2). The key elements in the oxidative pathway which are controlled by polyphenols are mostly in the PI3-K/Akt pathway that produces HO-1 through transcription of the Nrf-2 gene. Other than this pathway, iNOS is frequently mentioned as the target of polyphenols. For the apoptotic pathway, only the pathway that involves Bcl-2 is indicated among many studies.
Studies on polyphenols' anti-RA effects have mainly focused on their influence on inflammation pathways. There are some studies that concentrate on the anti-oxidative and apoptotic effect of polyphenols which result in a reduction of RA symptoms, but those are few in number. Further studies are needed in clarify the molecular studies mechanism of polyphenols' anti-oxidative and apoptotic effects that regulate RA's pathogenic pathways.  In this review we have organized and summarized the role of each polyphenol compound in diverse pathogenic pathways of RA. This work will be significant in providing systematized information for developing natural-product-based RA therapeutic solutions.

Methods
Searches regarding the anti-RA effects of polyphenol were conducted on PubMed and Google Scholar in August of 2018. When searching for appropriate studies, we included "rheumatoid arthritis", and "polyphenol" as keywords. Only articles written in English, published from 2006 to 2018 were selected for further review. We selected studies which met the following criteria: (i) studies based on in vitro or in vivo experiments that demonstrate the anti-RA effects of polyphenols; (ii) studies that show statistically significant analysis data (p < 0.05); (iii) studies that were not shown to have errors by subsequent studies; (iv) studies written in English. For classifying the type of polyphenol, we used the method of Soto et al. [60].

Conclusions
Polyphenols reduce rheumatoid arthritis symptoms by regulating an extensive collection of RA-related molecules, including MAPK, IL-1β, IL-6, TNF-α, NF-κB, JNK, ERK1/2, AP-1 and COX-2. Studies on polyphenols' anti-RA effect were mainly focused on their influence on inflammation pathways. Further studies are needed for clarifying the molecular mechanism of polyphenol's anti-oxidative and apoptotic effects that also regulate RA's pathogenic pathways. Based on these preclinical data, clinical trials could be conducted.