ASC was reported as a potent antiviral and
ASC was reported as a potent antiviral and antifungal agent, and its derivatives could modulate a variety of physiological activities, such as anti-tumor, anti-metastatic, and anti-diabetic activities in animals. Mie Tsuruga et al. found that 4-o-methylascochlorin had immunosuppressive activity, but the mechanism was not revealed (Tsuruga et al., 2007). In this study, we demonstrated that ASC showed notable anti-inflammatory and immunosuppressive effects in vitro and in vivo through reversible inhibition of hDHODH.
In inhibition kinetics study, the double-reciprocal plots of data showed ASC was an uncompetitive inhibitor with respect to DHO and a competitive inhibitor with respect to CoQ10. ASC resembled CoQ and the CoQ analogues (Fig. 1), which could explain that it exhibited competitive inhibition relative to CoQ10. As CoQ analogues, ASC should be uncompetitive inhibitors versus DHO, due to the two-site ping-pong enzyme mechanism (McLean et al., 2001). Similarly, ASC was also a specific inhibitor of mitochondrial cytochrome bc1 complex, as structurally related to CoQ10 (Berry et al., 2010).
IL-2 plays an important role in T lymphocytes activation and proliferation (Thornton et al., 2004), as most T lymphocytes activation pathways are mediated by it. IFN-γ promotes T lymphocytes differentiation and activates macrophages to secrete various cytokines. Their abnormal expressions are associated with many auto-inflammatory and auto-immune diseases (Malek, 2008, Marwaha et al., 2014). Our results showed that ASC could significantly inhibit T lymphocytes proliferation and decrease the IL-2 and IFN-γ production that were simulated by ConA, suggesting that ASC exerted potent immunosuppressive activity. A recent study indicated that ASC exhibited anti-inflammatory effects in LPS-stimulated RAW 264.7 macrophage serine threonine protein kinase through inhibition of nuclear factor-κB pathway (Lee et al., 2016), while our findings provided another anti-inflammatory mechanism by inhibition of hDHODH.
Furthermore, the in vivo anti-inflammatory and immunosuppressive effect of ASC was demonstrated by results from animal models. The xylene-induced ear swelling is a common inflammatory model for evaluating vascular permeability. Our studies proved that ASC (50mg/kg) strongly inhibited xylene-induced ear swelling in mice.
HDHODH could be a potential target for development of anti-rejection drugs, since inhibition of hDHODH suppressed lymphocytes activity against specific alloantigen. Several hDHODH inhibitors had been proved to have beneficial effects on rejection and prolong survival time of organ (Rusai et al., 2012, Wennberg et al., 2000). In allogeneic skin graft study, we found that ASC could markedly inhibit skin graft necrosis and increase mean graft survival time.
RA is a chronic inflammation characterized by painful joints and progression to irreversible joint destruction (McInnes and Schett, 2011). hDHODH is an effective target for RA chemotherapy, as it plays an important role in inflammation and immunoregulation. In the present study, the CIA model was used to investigate the anti-arthritic effect of ASC. Our results showed that 40mg/kg ASC treatment significantly attenuated ankle joint swelling in rats and exhibited sustained anti-arthritis effect over 4 weeks.
In summary, ASC from microbial metabolites was identified as a new structural class of hDHODH inhibitors with in vitro and in vivo effects on anti-inflammation and immunosuppresion. It may be a promising candidate for development of new therapy for the treatment of autoimmune diseases.
Conflicts of interest
Dihydroorotate dehydrogenase (DHODH) is responsible for the conversion of dihydroorotate to orotate, which is the rate-limiting step in pyrimidine biosynthesis. Inhibitors of DHODH show immunosuppressant and antiproliferative activities, which are most pronounced on activated T-cells. Leflunomide, an inhibitor of DHODH, is currently used to treat rheumatoid arthritis, and analogs are in clinic to treat graft versus host disease and multiple sclerosis.