Experimental autoimmune encephalomyelitis inhibited by Huangqi Guizhi Wuwu Decoction by enhancing Th2 cytokine

Abstract

Background: Current modern conventional medicine (MCM) on multiple sclerosis (MS) are non-specific immunosuppressive drugs, which remains many side effects. Huangqi Guizhi Wuwu decoction (HQGZWW) is a common formula of Chinese herbal medicine (CHM) has good effects on treatment of MS and its animal model, experimental autoimmune encephalomyelitis (EAE) very well, so it is very important to understand the precise mechanism. Our previous study suggested that CD8+ autoreactive T cells in EAE had a lower encephalitogenic function but were unique and independent on pathogenic of EAE rather than their CD4+ counterparts. The aims of current study were to determine the pathological interrelationship between CD4+ and CD8+ autoreactive T cells in MS/EAE upon the HQGZWW treatment.Methods: Female C57BL/6 mice (n=8, each group) were induced by myelin oligodendrocyte glycoprotein (MOG)35-55 peptide, and meantimely were treated with distilled water, prednisone, high dose or low dose HQGZWW. At 14 days after immunization, T cells were isolated from the spleen and purified as CD4+ and CD8+ T cells by using CD4 and CD8 isolation kits, and then the purity was determined by flow cytometric analysis. These cells were stimulated by MOG35-55 peptide and applied to proliferation assays. The interferon-gamma (IFN-g), interleukin (IL)-4 and IL-10 secretion of supernatant of cultured CD4+ and CD8+ T cells were measured by enzyme-linked immunosorbent assays (ELISA). For adoptive transfer, recipient mice were injected with MOG35-55 -specific CD8+ or CD4+ T cells. EAE clinical course was measured by EAE score at 0-5 scale and spinal cord was examined by staining with hematoxylin and eosin and Luxol fast blue staining.Results: For aEAE, there were significant improvement of EAE score in both HQGZWW high dose and prednisone groups by EAE score and pathological examination of spinal cord. CD8+ CD3+ and CD4+CD3+ cells were around 90% pure of total CD3+ cells after CD8/CD4 bead enrichment in 4 groups, respectively. These cells were stimulated by MOG35–55 peptide and applied to proliferation assays. There is lower antigen-specific responses of CD8+ as well as CD4+ T cells in HQGZWW high dose and prednisone group, compared with HQGZWW low dose and distilled water groups. For cytokine profiles, the CD4+ and CD8+ T cell supernatants contained lower levels of IFN-g and higher levels of IL-4 and IL-10 in HQGZWW high dose and prednisone groups compared with HQGZWW low dose and distilled water groups. Finally, HQGZWW had similar effect at high dose level to typical conventional medicine prednisone on tEAE, but no effect at low dose level.Conclusion: Our data suggested that HQGZWW had similar effect at high dose level to typical conventional medicine prednisone on EAE, but no effect at low dose level, and it suggested that the protection role of HQGZWW on EAE might be upon Th2 cytokine secretion profile by either MOG35–55 specific CD8+ or CD4+ T cells.