In vitro evaluation of physiologically relevant concentrations of teriflunomide on activation and proliferation of primary rodent microglia

Tanja Wostradowski, Chittappen Kandiyil Prajeeth, Viktoria Gudi, Jessica Kronenberg, Sina Witte, Marina Brieskorn, Martin Stangel

Teriflunomide, an inhibitor of dihydroorotate dehydrogenase, is thought to ameliorate multiple sclerosis by reducing activation-induced proliferation of lymphocytes, which is highly dependent on de novo pyrimidine synthesis. Nevertheless, its immunomodulatory effects on resident glial cells in the central nervous system are only poorly understood. In this study, we employed physiologically relevant concentrations of teriflunomide and investigated its effects on survival, proliferation, activation, and function of primary rat microglia in vitro. We demonstrate that teriflunomide had no cytotoxic effect on microglia and had only a minor impact on microglial activation. In a concentration- and time-dependent manner, teriflunomide significantly downregulated surface expression of the co-stimulatory molecule CD86. Furthermore, in the highest concentration applied (5 μM), it slightly increased the expression of interleukin-10 in microglia in response to lipopolysaccharide. Treatment with low concentrations of teriflunomide (0.25-1 μM) did not have any impact on the activation or proliferation of microglia. At 5 μM concentration of teriflunomide, we observed a reduction of approximately 30 % in proliferation of microglia in mixed glial cell cultures. Taken together, our in vitro findings suggest that at higher concentrations, teriflunomide potentially exerts its effects by reducing microglial proliferation and not by modulating the M1-/M2-like cell differentiation of primary rat microglia. Thus, teriflunomide has no major impact on the plasticity of microglia; however, the anti-proliferative and minimal anti-inflammatory effects might be clinically relevant for immune modulation in the treatment of neuroinflammatory CNS diseases such as multiple sclerosis.