**5. Potential role of free radicals in rheumatoid cachexia**

Cachexia is one of the major causes of progressive weight loss and affects up to 20% of RA patients [92]. Unlike sarcopenia, which is a normal physiological process of body mass reduction affected by aging, cachexia appears to be a secondary manifestation of an already ongoing disease [93]. Cachexia associated with RA can occur in two forms. The first form is cachectic RA or rheumatoid cachectic obesity, which is manifested by severe muscle wasting, with little or no fat mass loss. It is a less threatening form of cachexia mainly because the energy demands of muscles can be compensated by lipid metabolism [94]. The second form is rheumatoid cachexia, which is manifested by severe muscle wasting as well as fat loss.

Rheumatoid cachexia (RC) is a progressive form of RA, which is primarily thought to be caused by the abnormal production of the pro-inflammatory cytokines produced by the immune cells localized in the synovial tissue of the affected joints. Excessive concentrations of several cytokines, especially TNF-α, IL-1β, IL-6, and INF-γ, could potentially affect the intracellular mechanisms of muscle fibers, leading to severe muscle atrophy and weakness [95]. The most dominant cytokine in RA and RC pathogenesis appears to be TNF-α which acts synergically with IL-1β. When bound to their specific receptors, these cytokines cause activation of NF-κB signaling cascade. A study by Cai et al. [96] suggests that muscle atrophy is

### *Impact of Oxidative Stress on Inflammation in Rheumatoid and Adjuvant Arthritis: Damage… DOI: http://dx.doi.org/10.5772/intechopen.89480*

predominantly promoted by the NF-κB pathway via the activation of MuRF1 transcription factor which ultimately induces immoderate proteolysis of muscle proteins by activating the ubiquitin-proteasome system. Moreover, Castillero et al. [97] observed overexpression of MuRF1 as well as several other myogenic factors, such as atrogin-1/MAFbx ubiquitin ligases in adjuvant arthritis.

Another important pathogenic factor of RC is reduced physical activity, which appears to be the result of either poor pain management of inflamed and swollen joints, metabolic changes, or merely general caution for physical activity. Lower physical activity leads to reduced muscle fiber stimulation, which significantly disrupts the cycle of muscle proteolysis and proteosynthesis in favor of proteolysis [98]. One of the possible triggers of RC could also be increased free radical concentration and onset of OS.

As mentioned in the previous text, ROS and RNS concentrations have been reported to be elevated in the joint area as well as plasma. This may indicate that an increase in free radicals levels could also be found in skeletal muscle tissue. There are several sites of free radical production in muscles including mitochondria, sarcoplasmic reticulum, and sarcolemma [99]. As metabolically highly active organs, muscles dramatically increase their oxygen consumption during physical activity in order to compensate various energy-dependent processes. Concurrently excessive amounts of oxidants are produced, which then serve as messenger molecules in multiple intracellular cascades. The main site of free radical generation is mitochondria during aerobic metabolism and oxidative phosphorylation. It has been shown that complexes I and III and more recently complex II of mitochondrial electron transport chain are key producers of ROS in muscle fibers [100]. Several authors suggest that the major ROS produced in muscle cells is superoxide anion (O2•), which is a very unstable radical and rapidly undergoes reduction resulting in dismutation into hydrogen peroxide (H2O2) [101]. Even though H2O2 is quite a stable nonradical molecule, excessive concentrations of H2O2 could ultimately result in increased generation of hydroxyl radical (•OH)–a highly reactive ROS which could potentially damage various cellular molecules and disrupt many intracellular mechanisms. Free radicals are also regularly produced by several enzymes such as nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (Nox) family as well as xanthine oxidase (XO) [99]. In skeletal muscles only Nox 2 and Nox 4 isoforms have been found, and it is believed that both of these isoforms are localized primarily in mitochondria [102]. However, the precise mechanism by which the increased activity of these enzymes is promoted is to date poorly understood. Under physiological conditions, excessive concentrations of free radicals are regularly scavenged and converted into non-radical molecules by antioxidant defense system molecules. However, several studies have observed low concentrations of some nonenzymatic antioxidants such as GSH [41] as well as low activity of enzymatic antioxidants such as SOD and glutathione peroxidase (GPX) in RA, which could potentially affect muscle tissue [103]. It has been proposed that decreased physical activity in RC patients could play a major role in oxidative damage of muscle cells since lower muscle stimulation reduces antioxidant capacity thus causing impaired balance in oxidant-antioxidant ratio [104].

Several long-term studies have reported a number of negative effects of free radicals in muscles at the molecular level. Oxidative damage of lipids, particularly in cell membranes [105], as well as nucleic acids in the DNA [106] is of great importance to normal cellular functioning, lately there has been a great deal of emphasis on protein modifications caused by ROS in multiple diseases.

Proteins as functional units of the cell can cause great damage to the cell itself if its space conformation is disrupted. Perhaps the most common protein modification caused by imbalance of oxidative status is carbonylation of side chains of multiple

the synovial membrane of mice with collagen-induced arthritis [85]. Catalase cata-

Activity of glutathione peroxidase (GPx) in blood serum and muscles of rats with AIA increased and activity of glutathione reductase (GR) in these tissues increased in comparison with the control. Probably, changes in enzyme activity are a defense response of the body to ROS generation in RA and can be a result of ROS activation or stimulation of their synthesis [87]. Similarly in the study of Sahu et al. [88], CIA increased antioxidant enzyme GPx and GR activities in joints, liver,

overexpression of heme oxygenase-1 HO-1 in RA lesions. In addition to superoxides and pro-inflammatory cytokines, hypoxia may play an important role in HO-1 expression in the lesions [89, 90]. AIA is an experimental model widely used to evaluate etiopathogenetic mechanisms in chronic inflammation. Devesa et al. [91] have examined the participation of HO-1 in AIA. They have found an increased nitric oxide (NO) production in the paw preceded the upregulation of HO-1, whereas selective inhibition of inducible NO synthase (iNOS) after the onset of arthritis lowered HO-1 expression, suggesting that this enzyme may depend on NO produced by iNOS. Administration of the HO-1 inhibitor protoporphyrin IX ameliorated the symptoms of arthritis. This compound significantly decreased leukocyte infiltration, erosion of articular cartilage, and osteolysis, as well as the production of inflammatory mediators. In this model, HO-1 can be involved in vascular endothelial growth factor production and angiogenesis. These results support a role for HO-1 in mediating the progression of the disease in this model of chronic arthritis [91]. Our research group showed that extra-articular manifestations of AIA are present also in lung, where the expression of heme oxygenase-1 was

lyzes the decomposition of hydrogen peroxide to water and oxygen, thus preventing the oxidation of biological structures by hydrogen peroxide. Authors demonstrated the elevated and LPO activity and NO level and decreased GSH, SOD, and catalase activities in AIA rats [86]. OS in AIA model is depleting antioxidant

Several pathologic factors have been suggested to be involved in the

**5. Potential role of free radicals in rheumatoid cachexia**

Cachexia is one of the major causes of progressive weight loss and affects up to 20% of RA patients [92]. Unlike sarcopenia, which is a normal physiological process of body mass reduction affected by aging, cachexia appears to be a secondary manifestation of an already ongoing disease [93]. Cachexia associated with RA can occur in two forms. The first form is cachectic RA or rheumatoid cachectic obesity, which is manifested by severe muscle wasting, with little or no fat mass loss. It is a less threatening form of cachexia mainly because the energy demands of muscles can be compensated by lipid metabolism [94]. The second form is rheumatoid cachexia, which is manifested by severe muscle wasting as well as fat loss. Rheumatoid cachexia (RC) is a progressive form of RA, which is primarily thought to be caused by the abnormal production of the pro-inflammatory cytokines produced by the immune cells localized in the synovial tissue of the affected joints. Excessive concentrations of several cytokines, especially TNF-α, IL-1β, IL-6, and INF-γ, could potentially affect the intracellular mechanisms of muscle fibers, leading to severe muscle atrophy and weakness [95]. The most dominant cytokine in RA and RC pathogenesis appears to be TNF-α which acts synergically with IL-1β. When bound to their specific receptors, these cytokines cause activation of NF-κB signaling cascade. A study by Cai et al. [96] suggests that muscle atrophy is

enzymes, which is in good agreement with human RA studies.

kidney, and spleen tissues of rats.

*Animal Models in Medicine and Biology*

reduced during AIA [60].

**206**

amino acids such as arginine, lysine, threonine, and proline [107]. Moreover, carbonylation of proteins that are part of the contractile apparatus could be crucial in RC muscle dysfunction. Fedorova et al. [108] showed that carbonylation of actin could very much affect actomyosin ATPase activity, thus promoting subsequent muscle atrophy. Taken together, action of pro-inflammatory cytokines, mitochondrial dysfunction, and enhanced activity NADPH oxidase and xanthine oxidase contribute to the overproduction of ROS/RNS.

sufficiently understand precise mechanism by which this serious condition occurs. This could greatly improve quality of RA patients and thus contribute to modern

*Impact of Oxidative Stress on Inflammation in Rheumatoid and Adjuvant Arthritis: Damage…*

The animal model adjuvant arthritis gives a broad spectrum of possibilities to study different pathological mechanism of rheumatoid arthritis. One important pathological pathway is the connection between inflammation and oxidative stress, which is studied on both systemic and local levels. From our original results as well as from results reported by other authors, it is evident that treatment with compounds possessing redox balance modulating properties might be of great relevance for new strategies for therapy of rheumatoid arthritis. For this purpose, adjuvant arthritis seems to be an ideal animal model. Moreover, this animal model has also a good potential in the research of inflammatory cachexia and its pharmacological

Our experimental studies were supported by grants: APVV-15-0308, APVV SK-PT-18-0022, and VEGA 2/0115/19. We thank Martin Chrastina, MSc, for technical

The authors declare that they do not have any conflict of interest.

Center of Experimental Medicine, Institute of Experimental Pharmacology and

© 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,

Silvester Ponist\*, Miloslav Zloh and Katarina Bauerova

Toxicology, Slovak Academy of Sciences, Bratislava, Slovakia

\*Address all correspondence to: silvester.ponist@savba.sk

provided the original work is properly cited.

medicine.

**6. Conclusion**

*DOI: http://dx.doi.org/10.5772/intechopen.89480*

intervention.

assistance.

**Acknowledgements**

**Conflict of interest**

**Author details**

**209**

Decreased physical activity results in downregulation of antioxidants. Combination of these factors consequently leads to imbalance in protein synthesis and degradation resulting in muscle wasting (**Figure 4**).

#### **5.1 Animal models in rheumatoid cachexia**

Rheumatoid cachexia still remains a poorly investigated disease, and many scientists are trying to understand the exact mechanism by which the disease takes place. Several animal models of RA are used in the study of this condition. The best studied animal model to date has been CIA, which, by its characteristics, offers the most accurate comparison with humans, as the onset of this affection is relatively slow and the immune mechanisms driving the onset of cachexia are closest to rheumatoid cachexia in people [109].

Recently, Albarse et al. [110] have been investigating the development of cachexia in CIA in DBA1/J mice. In their study, they have observed significant increase in free exploratory locomotion as well as grip strength and endurance exercise performance. Additionally, they registered reduction of muscle weight in several muscles, which could indicate that mechanisms, which led to the onset of arthritis, could subsequently promote muscle atrophy and weakness.

Another model of rheumatoid arthritis and adjuvant arthritis was also used to investigate muscle wasting in male and female Lewis rats in the study of Roubenoff et al. [111]. It was shown that adjuvant-induced rats also manifested severe muscle loss when compared to control as well as pair-fed groups. This makes adjuvant arthritis suitable model for study of cachexia in chronic inflammatory diseases. Even though there have been multiple authors dedicated to unraveling the true cause of rheumatoid-induced cachexia, much more study is needed in order to

#### **Figure 4.**

*Mechanism of the effect of oxidative stress on the onset of cachexia in rheumatoid arthritis. NOX, nicotinamide adenine dinucleotide phosphate (NADPH) oxidase; XO, xanthine oxidase; ROS, reactive oxygen species; RNS, reactive nitrogen species.*

*Impact of Oxidative Stress on Inflammation in Rheumatoid and Adjuvant Arthritis: Damage… DOI: http://dx.doi.org/10.5772/intechopen.89480*

sufficiently understand precise mechanism by which this serious condition occurs. This could greatly improve quality of RA patients and thus contribute to modern medicine.
