The Effect Of GPR97 On The Pathogenesis Of EAE And The Immunoregulation Of DC Cells And Its Mechanism

G protein-coupled receptors (GPCRs), also known as 7-transmembrane receptors, are the largest family of cell surface receptors which play an important role in transmitting extracellular environmental signals into the cells. A mount of these receptors comprise approximately 2% of the human genome and mediate most of our physiological responses. Dysfunction of GPCRs leads to various diseases and remains a central focus in basic pharmacology studies and drug discovery efforts. GPR97 is a 549 amino acid multi-pass membrane protein and belongs to the G-protein coupled receptor 2 family. GPR97 possesses an exceptionally long extracellular region and is an orphan adhesion GPCR. Until now, its physiological function remains greatly unclear. Because adhesion GPCRs usually are related to the immune function, GPR97 may play an important role in immunoregulation.Multiple sclerosis is an autoimmune disease of the central nervous system, which is the leading cause of non-traumatic disability in young adults. Etiology of MS contains genetic, environmental and immune factors, but its exact pathogenesis is unknown. Although numerous studies have been made over the past few decades, there is no effective cure for MS. As a widely used animal model for MS, experimental autoimmune encephalomyelitis (EAE) shares many pathological and histological similarities with MS. MOG peptide is usually used for the induction of EAE and EAE model has been extensively used to study the pathogenesis of MS and find the new therapeutic targets for MS.Dendritic cells (DCs) are the most crucial antigen-presenting cells, which link innate and adaptive immune responses. Immature DCs have strong phagocytic ability and mature DCs increase the expression of MHC and co-stimulatory molecules such as CD40 and CD86. Mature DCs also secrete large amounts of cytokines that stimulate naive Th cells to differentiate into different cells. A number of studies have showed that DCs may involved in the pathogenesis of MS.GPR97-knockout mice are potent tool to study the functions of GPR97. We detected the immune cells and the subsets in the spleen of GPR97-knockout mice. The results showed that T cell numbers in spleen of GPR97-knockout mice were a little lower than those of wild-type mice, while a little higher cell numbers were indicated in NK, CD4+T cells, Thl and Th17 subsets. Dendritic cells were also detected and the results showed that there were significantly higher cell numbers and higher level of surface molecule expression on DCs in spleen of GPR97-knockout mice compared to wild-type mice.To study the role of GPR97 in the pathogenesis of MS, GPR97-knockout mice were used as the object of study, EAE model was established by immunizing mice with MOG peptide. The role of GPR97 gene knockout in the pathogenesis of EAE mice was analyzed by clinical score, histological staining of spinal cord and immunological index. Compared to EAE model of wild-type mice, the results showed that GPR97-knockout mice developed a higher clinical score, increased inflammatory cell infiltration in the CNS, more Thl and Th17 cell subsets, higher level of pro-inflammatory cytokines such as TNF-α、IL-17A and IL-6 and more MOG-specific T cells in the spleens. These data suggested that GPR97-knockout mice exhibited increased disease severity in a model of EAE.To elucidate the possible mechanism of GPR97 on the pathogenesis of EAE, DC cells were used as the target cells, the effect of GPR97 gene knockout on the biological functions was analyzed by detecting the cytokine secretion profile, surface molecule expression, induction of Th cell differentiation and antigen presentation, etc. RT-PCR and ELISA analysis showed that more IL-6, TGF-β,IL-12 but less IL-10 were detected in GPR97-knockout DCs. GPR97-knockout DCs induced the differentiation of Th into Thl and Th17 subsets. Higher expression level of surface molecules such as CD40 and CD86 on GPR97-knockout DCs. GRP97-knockout DCs had stronger ability to stimulate the proliferation of MOG-specific T cells. The data strongly suggested that the deficiency of GPR97 enhanced the biological function of DCs.In conclusion, GPR97 is closely relative to the pathogenesis of EAE and the possible mechanism of more severe EAE in GPR97-knockout mice is that the deficiency of GPR97 enhanced the immunological functions of DC cells. The study lay the theoretical and experimental basis for the MS treatment using GPR97 as the the therapeutic target.