HLA-E单抗抑制B细胞激活减轻肾移植抗体介导排斥反应的机制

Antibody mediated rejection (AMR) is a major cause of long-term kidney allograft failure. IVIg is an effective way to treat AMR, however its mechanism has not been figured out. We found there contains high dose HLA-E antibodies in IVIg, and its monoclonal antibody could inhibit antibody secretion by activated B cells. And this phenomenon may work through its inhibition on B cell activation. Moreover, we found HLA-E monoclonal antibody could bind to the open conformer of MHC I on the surface of activated B cells. References demonstrated the elongated tale of MHC I could recruit and up-regulate SHP phosphorylation. B cells activation could be inhibited by SHP-1 and SHP-2 through Syk-Btk/MAPK and TRAF6/NF-kB signal pathway respectively. Hence we make hypothesis that HLA-E monoclonal antibody could bind to the open conformer of MHC I on the surface, which recruit and up-regulate SHP phosphorylation, to inhibit B cells activation by Syk-Btk/MAPK and TRAF6/NF-kB signal pathway. For this purpose, we plan to utilize rat kidney transplant model to figure out the effectiveness of HLA-E monoclonal antibody on inhibiting B cells activation and alleviating kidney allograft AMR, and illustrate the molecular mechanism of HLA-E monoclonal antibody on inhibiting B cells activation in vivo and in vitro. This program could provide theoretical basis and experimental support for the clinical application of HLA-E monoclonal antibody.

抗体介导的排斥反应（AMR）是影响移植肾长期存活的主要因素。静脉注射免疫球蛋白（IVIg）被广泛用于治疗AMR，机制亟待阐明。我们发现HLA-E抗体在IVIg中含量很高，其单抗能与B细胞表面的MHC I类抗原开放构象结合，并显著抑制B细胞激活与抗体分泌，具有减轻肾移植AMR的潜在作用。研究表明MHC I类抗原可募集并激活SHP，而SHP-1和SHP-2能分别通过抑制Syk-Btk/MAPK及TRAF6/NF-kB通路抑制B细胞激活。因此我们推测：HLA-E单抗通过与MHC I类抗原开放构象结合，募集并激活SHP-1与SHP-2，进而分别通过抑制Syk-Btk/MAPK及TRAF6/NF-kB通路抑制B细胞激活。为此，本项目拟通过大鼠肾移植模型，明确HLA-E单抗抑制B细胞激活减轻肾移植AMR的作用，阐明并验证其抑制B细胞激活的分子机制，为将来HLA-E单抗的临床应用提供理论基础和实验支持。

抗体介导的排斥反应（AMR）是影响移植肾长期存活的主要因素，近年来受到越来越多的关注。B细胞激活后分泌供者特异性抗体（DSA）是AMR发生机制中非常重要的环节，因此阻断B细胞激活是预防和缓解AMR的有效治疗手段。本项目的立项意在研究HLA-E单抗对AMR的缓解效果，以及其抑制B细胞激活、降低DSA分泌的内在机制。在本基金的资助下，我们构建了小鼠皮肤移植AMR模型，发现HLA-E单抗可以延长小鼠的皮肤移植物存活时间，降低DSA分泌，减轻AMR。在应用HLA-E单抗干预的情况下，B细胞表面CB80、MHC II类分子等表达明显降低，证明B细胞激活受到抑制。机制研究表明，HLA-E单抗通过SHP-2/TRAF6/NF-kB通路抑制了B细胞激活。在我们应用SHP抑制剂PP1后，HLA-E单抗抑制B细胞激活的作用明显减弱。结合我们前期发现的HLA-E单抗与MHC I类分子开放构象的结合靶点，这些结果揭示HLA-E单抗是通过与MHC I类分子结合后激活SHP阻断TRAF6/NF-kB通路进而抑制B细胞激活的。同时我们利用小鼠皮肤移植AMR模型，研究了环肽CHBP对AMR的作用。我们发现，环肽CHBP能降低DSA分泌，减缓小鼠移植皮肤AMR。应用环肽CHBP的小鼠，脾脏GC B细胞和Tfh细胞数量和比例明显降低。进一步研究发现Tfh激活相关的JAK-STAT3信号通路被阻断。这说明环肽CHBP抑制B细胞激活，且此作用与生发中心的Tfh细胞密切相关。课题组还在肾移植术后免疫状态监测上做了创新性研究，发现了尿液代谢特征谱在诊断排斥反应中的诊断价值，并构建了基于网络构架和蛋白表达的诊断模型，为临床无创诊断排斥反应提供了新的思路。