The Effect Of Purinergic Receptor P2Y12 On The Pathogenesis Of EAE And Its Mechanism

Autoimmune diseases arise from an abnormal immune recognition of substances and tissues normally present in the body, and launch an immune attack against ourselves. Multiple sclerosis is an important autoimmune disease which is mainly mediated by CD4+ T cells and is characterized by demyelination and neurodegeneration of central nervous system (CNS)—brain and spinal cord. Although plenty of researches had been done on MS by scientists, there still remains a number questions on the specific mechanisms until now. EAE (Experimental Autoimmune Encephalomyelitis) is a widely used animal model for MS, which shares similarities both in clinical phenotypes and pathological characteristics. Moreover, because of its mature methods, apparent clinical phenotypes and standardized evaluation system, EAE model is a classical autoimmune disease model for basic researches.P2Y12 receptor, a member of P2Y receptor family, is also a G protein-coupled receptor. It enjoys close and extensive attention for its importance in regulating the function of blood platelet. As is known to all, GPCR take an essential role in regulating multiple biological functions and signal transduction downstream. A previous study has proved that the expression level of P2Y12 receptor is altered in MS patients compared with the normal, but whether P2Y12 has the relevance with the pathogenesis of MS is not clear. Therefore, we mainly focused on the relationship between P2Y12 receptor and MS/EAE by constructing EAE animal model based on the wild type and P2Y12-knockout mice.In this study, we discovered that the expression of P2Y12 in the spleen of EAE mice was lower than that of wild type mice. Compared to the wild type mice, P2Y12-knockout mice developed much more severe EAE whatever in clinical scores, pathological characteristics or the analysis of CD4+ T cell subgroups. Naive CD4+ T cells were sorted from the spleen and cultured ex vivo for further study. It was demonstrated that P2Y12 did not influence differentiation and proliferation of naive CD4+ T cells in vitro. To further explore the specific mechanism of P2Y12 on the pathogenesis of EAE, bone marrow-derived dendritic cells (BMDCs) were prepared to and the biological functions of DCs were analyzed by detection of the cytokine secretion, surface molecule expression and T cell differentiation. Genetic deletion of P2Y12 significantly increased the expression of CD40, CD86 and MHCⅡ. In terms of the expression of cytokines, IL-23pl9 and TGF-β in P2Y12-knockout BMDCs had higher expression in contrast to the wild type BMDCs. FACS analysis indicated that the culture supernatant from P2Y12-knockout DCs promoted more naive CD4+ T cells to differentiatiate into Th17 cells compared to wild type DCs.In conclusion, genetic deletion of P2Y12 receptor broke the balance of Th subtypes, especially Th17 cells by affecting the function of BMDCs, resulting in the aggravated EAE. Our findings suggest that P2Y12 may be a potential target in treating MS/EAE, and also provides guidance for clinical medication.