PXR/VPS33A通路在CKD肾小管间质纤维化中的作用及机制研究

Renal fibrosis is a common event in many chronic kidney diseases. Investigation on the mechanisms of renal fibrosis is of importance for the development of novel therapeutic strategies of CKD. Nuclear receptor PXR have been investigated in the pathophysiology of many diseases, while its role in CKD has not been studied. Through the analysis of Nephroseq database, PXR was shown to be enhanced in the kidneys of CKD patients. In the kidneys of CKD patients and UUO mice, as well as in the renal fibroblasts treated by TGFβ, we further confirmed the upregulation of PXR under CKD condition. Activation or overexpression of PXR in renal fibroblasts strikingly blunted TGFβ-induced fibrotic response, suggesting an antifibrotic role of PXR in CKD. Furthermore, to explore the mechanism of PXR in antagonizing fibrotic response in kidney, we performed proteomics analysis using the kidney tissues from newly generated PXR knockout rats and found that VPS33A was strikingly downregulated. Overexpression of VPS33A attenuated TGFβ-induced fibrotic response, which suggests a protective role of VPS33A against renal fibrosis. Furthermore, bioinformatics analysis indicated that VPS33A is transcriptionally regulated by RXRαbut not PXR. Solid published evidence demonstrated that PXR and RXRαcould form a dimer to transcriptionally regulate their target genes. In agreement with this concept, we observed a significant upregulation of RXRα in the kidneys of UUO mice. Thus, these data highly suggested that PXR might regulate VPS33A via forming a dimer with RXRα to regulate the autophagic response to antagonize the fibrotic response in CKD. In the present proposal, we will fully investigate the role and mechanism of PXR/VPS33 pathway in protecting against renal tubulointerstitial fibrosis using the human CKD samples and models of animals, cells and molecules, which will offer novel insights into the prevention and therapy of CKD.

对肾脏纤维化机制进行深入研究有助于寻找新的CKD干预靶点。核受体PXR在机体防御机制中有重要作用，但在CKD中尚无报道。Nephroseq database显示PXR在CKD肾脏显著增加。预实验证实PXR在CKD患者和UUO小鼠肾脏及TGFβ处理的肾成纤维细胞明显上调，激动剂及过表达PXR均显著抑制TGFβ诱导的肾成纤维细胞活化。蛋白质组学分析发现PXR基因敲除大鼠肾脏VPS33A显著降低，过表达VPS33A抑制了TGFβ的促纤维化作用。进一步实验和分析发现，PXR可能通过与RXRα形成二聚体介导VPS33A转录；敲低VPS33A导致了自噬相关蛋白P62异常表达，结合PXR参与自噬的报道，推测：PXR/VPS33A通路可能通过自噬促进肾脏细胞内环境稳定，在CKD肾纤维化中发挥拮抗作用。本项目将在整体、细胞及分子层面研究活化PXR/VPS33A通路在拮抗CKD肾间质纤维化中的作用及机制。

慢性肾脏病（CKD）是临床常见多发性疾病，其持续进展将发展为终末期肾病。在慢性肾脏病的进展过程中，肾脏纤维化不但是其重要的病理表现，也是促进其进展的重要病理因素。因此，发现慢性肾脏病肾脏纤维化的发生机制及有效干预新靶点是本领域的研究热点之一。本项目主要研究了：1. 核受体PXR在CKD患者及小鼠肾脏组织的表达与定位。2. PXR在CKD肾间质纤维化中的功能。3. PXR拮抗CKD肾间质纤维化的作用机制。4. 新型PXR激动剂的开发及其在防治CKD肾间质纤维化中的应用。研究结果显示：1. 在CKD患者、 UUO及5/6Nx 小鼠肾脏，我们证实PXR在CKD肾脏组织中表达上调，且主要定位于近曲小管及间质成纤维细胞。2. 在肾脏敲除PXR可以显著加重CKD肾间质纤维化，而激活及过表达PXR则可以显著改善CKD肾间质纤维化。3.一系列组学及体内与体外实验发现，PXR可能是通过激活ASPP2抑制了β-catenin的入核活化，进而延缓了CKD肾间质纤维化的发生与发展。4. 现有的PXR激动剂主要作用于鼠类动物，尚无可做用于人类的特异性PXR激动剂。由此，我们开发了新型的可做用于人的PXR激动剂（Sophoricoside (25-F6) 及Calycosin (34-H4)），并验证了其在CKD肾间质纤维化中的治疗作用。本项目的完成，不但发现了CKD肾间质纤维化的新机制与干预新靶点，也为临床防治CKD提供了可能的新药物。