**3. Newer experimental therapies with antioxidants evaluated in AA**

Oxygen metabolism has an important role in the pathogenesis of RA. ROS produced in the course of cellular oxidative phosphorylation and by activated phagocytic cells during oxidative bursts exceed the physiological buffering capacity and result in OS. The excessive production of ROS can damage proteins, lipids, nucleic acids, and matrix components. They also serve as important intracellular signaling molecules that amplify the synovial inflammatory–proliferative response. Repetitive cycles of hypoxia and reoxygenation associated with changes in synovial perfusion are postulated to activate hypoxia-inducible factor-1α and nuclear factor-κB, two key transcription factors that are regulated by changes in cellular oxygenation and cytokine stimulation, and that in turn orchestrate the expression of a spectrum of genes critical to the persistence of synovitis. Understanding of the complex interactions involved in these pathways might allow the development of novel therapeutic strategies for RA (Hitchon & El-Gabalawy, 2004). Free radicals from oxygen metabolism destroy antioxidant systems (Harris, 2003). Researchers such as Heliovaara et al. (1994) have suggested that enzymatic and/or nonenzymatic antioxidant systems are impaired in RA. RA patients are therefore exposed to oxidant stress (Harris, 2003). Consequently, a number of different activities of antioxidant enzymes, such as superoxide dismutase, glutathione peroxidase, catalase, and glutathione reductase have been reported to be effective in treating RA (Blake et al., 1981; Mazetti et al., 1996; Shah & Vohora, 2002). Some other researchers found that RA patients were more inclined to lipid peroxidation because of the reduced antioxidant defense system (Rowley et al., 1984; Gambhir et al., 1997). The synovial fluids from patients with RA were found to present high levels of antioxidant damage (Miyata et al., 1998; Chapman et al., 1999). According to Babior (2000) reactive oxidants are essential tools for the pathogenesis of RA.

There is widespread availability and interest in the use of antioxidant supplementation by patients with inflammatory arthritis, although proof of efficacy is modest. A traditional Mediterranean diet relatively high in antioxidants improved RA disease and functional status after 3 months compared with a standard 'Western' diet, although clinical improvement was not associated with any significant change in plasma levels of antioxidants (Heliovaara et al., 1994; Skoldstam et al., 2003). In a separate study of patients with RA, supplementation with the antioxidants vitamin A, E, and C increased plasma antioxidant levels with a corresponding decrease in MDA, a key marker of OS; however, a clinical response was not reported (Jaswal et al., 2003). Specific supplementation of oral vitamin E, the major lipid-soluble antioxidant in human plasma, erythrocytes, and tissue, had no effect on RA disease activity or indices of inflammation but did improve pain, suggesting a role in central analgesia mechanisms (Edmonds et al., 1997). Various forms of antioxidant therapy have demonstrated promising results in experimental arthritis models (Bandt et al., 2002; Cuzzocrea et al., 2000; Venkatraman & Chu, 1999).

IL-6, IL-8, and MIP- 1α. It was shown to suppress macrophage cytotoxic activity, parasite killing, and macrophage-derived nitric oxide production (Vannier, et al., 1992). In our experiments, detection of plasma IL-1α, IL-4, TNF-α, and MCP-1 was done by the flowcytometric (Cytomics FC 500, Beckman Coulter Inc. Fullerton, USA) fluorescent beadbased multiplex assay Rat Cytokine Flow Cytomix Multiplex (Bender Med System, GmbH., Austria). Additionally for determination of IL-1α in plasma an ELISA kit from Bender MedSystems or from R&D Systems Quantikine® was used and to as assessed MCP in

Oxygen metabolism has an important role in the pathogenesis of RA. ROS produced in the course of cellular oxidative phosphorylation and by activated phagocytic cells during oxidative bursts exceed the physiological buffering capacity and result in OS. The excessive production of ROS can damage proteins, lipids, nucleic acids, and matrix components. They also serve as important intracellular signaling molecules that amplify the synovial inflammatory–proliferative response. Repetitive cycles of hypoxia and reoxygenation associated with changes in synovial perfusion are postulated to activate hypoxia-inducible factor-1α and nuclear factor-κB, two key transcription factors that are regulated by changes in cellular oxygenation and cytokine stimulation, and that in turn orchestrate the expression of a spectrum of genes critical to the persistence of synovitis. Understanding of the complex interactions involved in these pathways might allow the development of novel therapeutic strategies for RA (Hitchon & El-Gabalawy, 2004). Free radicals from oxygen metabolism destroy antioxidant systems (Harris, 2003). Researchers such as Heliovaara et al. (1994) have suggested that enzymatic and/or nonenzymatic antioxidant systems are impaired in RA. RA patients are therefore exposed to oxidant stress (Harris, 2003). Consequently, a number of different activities of antioxidant enzymes, such as superoxide dismutase, glutathione peroxidase, catalase, and glutathione reductase have been reported to be effective in treating RA (Blake et al., 1981; Mazetti et al., 1996; Shah & Vohora, 2002). Some other researchers found that RA patients were more inclined to lipid peroxidation because of the reduced antioxidant defense system (Rowley et al., 1984; Gambhir et al., 1997). The synovial fluids from patients with RA were found to present high levels of antioxidant damage (Miyata et al., 1998; Chapman et al., 1999). According to Babior (2000) reactive oxidants are essential

There is widespread availability and interest in the use of antioxidant supplementation by patients with inflammatory arthritis, although proof of efficacy is modest. A traditional Mediterranean diet relatively high in antioxidants improved RA disease and functional status after 3 months compared with a standard 'Western' diet, although clinical improvement was not associated with any significant change in plasma levels of antioxidants (Heliovaara et al., 1994; Skoldstam et al., 2003). In a separate study of patients with RA, supplementation with the antioxidants vitamin A, E, and C increased plasma antioxidant levels with a corresponding decrease in MDA, a key marker of OS; however, a clinical response was not reported (Jaswal et al., 2003). Specific supplementation of oral vitamin E, the major lipid-soluble antioxidant in human plasma, erythrocytes, and tissue, had no effect on RA disease activity or indices of inflammation but did improve pain, suggesting a role in central analgesia mechanisms (Edmonds et al., 1997). Various forms of antioxidant therapy have demonstrated promising results in experimental arthritis models

(Bandt et al., 2002; Cuzzocrea et al., 2000; Venkatraman & Chu, 1999).

**3. Newer experimental therapies with antioxidants evaluated in AA** 

plasma by Instant ELISA kit from eBioscience®.

tools for the pathogenesis of RA.

As resulted also from our previous experiments (Bauerova et al., 2005a, 2005b, 2008a, 2008b, 2009, Drabikova et al., 2009; Drafi et al., 2010; Jancinova et al., 2009; Kogan et al., 2005; Macickova et al., 2010; Ponist et al., 2010; Sotnikova et al., 2009; Strosova et al., 2008, 2009), all performed in the AA model, substances with antioxidant properties have a high potency to be used in therapy of RA. They decreased the progression of AA when administered to rats with AA over the period of 28 days. For our experiments, we chose substances with antioxidative properties and low toxicity. These antioxidative substances were synthetic antioxidants, as pyridoindol derivatives, and natural substances possessing antioxidative properties. We chose compounds and extracts isolated from plants, polysaccharides isolated from yeast and mushrooms and finally also endogenous antioxidants. An overview of some selected results is given below along with new unpublished data to provide complex information about administration of antioxidants in AA.
