补体效应分子C5a在松鼠猴DCD肾移植后BK型多瘤病毒肾病模型中的作用及相关机制研究

Polyomavirus BK (BKPyV) infection and Polyomavirus BK nephropathy (BKPyVN) have become a major complication in kidney transplant recipients, which affect the graft survival greatly. A BKPyVN model in animal has never been developed, which can resemble human disease. Previously, we found donation after cardiac death(DCD) kidney transplantation was an independent risk factor for BKPyV infection and BKPyVN. The preliminary data showed BKPyV replication and antigen expression was regulated through the pathway of TNF-α－NF-kB and was in proportion to the level of C5aR in kidney graft. However, the mechanism of the enhancement of BKPyV replication due to DCD kidney transplantation is still not clear. Recent studies have found that C5a can be up-regulated greatly in the DCD donor and increases the expression of TNF-α. Based on the previous studies and our preliminary data, we hypothesize that up-regulation of C5a might activate BKPyV replication through the pathways of TNF-α－NF-kB after DCD kidney transplantation. Because BKPyV can only infect squirrel monkey and human at present, to test the above hypothesis, we simulate the process in cell culture, develop DCD kidney transplantation model in positive BKPyV serological donor and negative recipient squirrel monkey(D+R-).After the C5a in donor kidney is overexpressed or silenced, the level of BKPyV and antigen expression, C5a and important proteins expression in TNF-α－NF-kB pathway in the recipient will be tested. Then we use the inhibitors of above pathway to verify our hypothesis. The study will investigate the effect of C5a on BKPyV replication through the pathway of TNF-α－NF-kB after DCD kidney transplantation and its mechanism, and develop an animal model of BKPyVN in order to mimic the BKPyVN seen in human kidney transplant recipients. This will support a laboratory facility to investigate the pathogenesis of BKPyVN and develop antiviral drugs.

BK型多瘤病毒肾病(BKPyVN)严重影响肾移植预后，目前无动物模型。前期研究表明, DCD肾移植是PyV感染和PyVN的独立危险因素。预实验显示PyV复制受TNF-α－NF-kB通路调控，且肾组织中PyV与C5aR水平成正比，但机制不明。最近发现补体效应分子C5a可在DCD供体内大量活化上调下游TNF-α。由此，提出活化C5a经TNF-α—NF-kB通路激活DCD肾移植BKPyV复制。由于松鼠猴是目前证实的BKPyV唯一非人类宿主，本课题拟结合体内外实验，在术前BKPyV血清学状态为D+R-且供肾差异性表达C5a的松鼠猴间进行DCD肾移植，检测BKPyV复制、C5a 及TNF-α－NF-kB通路相关分子的表达，并用以上通路抑制剂，证实活化C5a上调TNF-α激活IkB/NF-kB通路增强DCD肾移植BKPyV的机制，构建与人类BKPyVN类似的动物模型，用于研究发病机制、开发抗病毒药物。

多瘤病毒(PyV)肾病(PyVAN)严重影响肾移植受者预后，但尚没有相关有效的动物模型。C3是补体系统中含量最高的成分，是系统的核心组成成分，其激活会进一步转化C5生成C5a。我们通过小鼠的动物实验建立PyV感染的动物模型，比较缺血再灌注损伤（IRI）和免疫抑制剂对于PyV再激活的影响。实验发现，肾IRI后第三天，小鼠多瘤病毒（MPyV）可在肾组织中再次激活载量明显增加且幅度大于右肾，效应大于免疫抑制组，随后逐渐减弱。GO功能显著性富集和KEGG信号通路显著性富集分析显示：病毒再激活复制跟NF-kB信号通路激活有关。随后引入公共数据库中移植肾BKPyVAN和正常移植肾数据做对比，我们观察到了C3、FN1、NF-κB在BKPyVAN中高表达。本研究探索了PyV致病的可能相关通路，可用于抗病毒药物的开发。