类泛素蛋白FAT10通过促进炎症反应介导肾缺血再灌注损伤的机制研究

Inflammation play an important role in the development of renal ischemia-reperfusion injury (RIRI), but its regulation mechanism is unclear. Our preliminary studies suggest that the ubiquitin-like protein FAT10 expression was elevated in tubular epithelial cell after RIRI；Knockout of FAT10 alleviated morphologic damage after RIRI, protecting from deterioration of kidney function. Furthermore, In vitro experiments indicate that FAT10 silence abrogated NF-κB signaling activation and reduced the induction of chemokine RANTES in tubular epithelial cells. Consequently, we hypothesize that the upregulation of FAT10 in renal tubular cells may promotes RANTES expression via activation of NF-κB signaling, and then aggravates renal inflammation, leading to tissue damage after RIRI. To determine the critical roles of FAT10 in post-RIRI inflammatory response leading to kidney injury, we will use WT and FAT10 knockout mice undergoing renal artery clamp models of ischemia-reperfusion injury in combining with RANTES neutralizing antibody injection in vivo, flow cytometry and pathological analysis. Moreover, to evaluate the effect of FAT10 on RANTES expression via NF-κB signaling pathway, we will employ the Lentiviral vector, Co-Immunoprecipitation (Co-IP), Cytometric bead array (CBA) and other methods. The proposed studies will provide new knowledge regarding the molecular mechanisms of how FAT10 participates the pathogenesis of renal ischemia-reperfusion injury through promoting inflammation, and provide the theoretical and experimental basis for intervention of acute kidney injury after RIRI.

炎症反应在肾缺血再灌注损伤的发生发展中起关键作用，但其发生和精确调控仍不完全清楚。我们前期研究发现：缺血再灌注损伤后肾脏组织中类泛素蛋白FAT10的表达上调，敲除FAT10明显减轻肾脏组织损伤以及改善肾功能；在肾小管上皮细胞中，沉默FAT10可抑制NF-κB信号通路的活化，而且减少趋化因子RANTES的产生。据此我们假设：在肾缺血再灌注损伤中，FAT10通过激活NF-κB信号通路促进趋化因子RANTES的产生，进而加重炎症反应，导致肾脏组织损伤。本申请通过体内注射RNATES中和抗体、流式细胞术结合病理学等技术探讨FAT10调控肾缺血再灌注损伤的作用和分子机制；采用慢病毒载体、流式微球分析技术及Co-IP等手段研究FAT10通过NF-κB信号通路调控RANTES产生的分子机制。本研究将阐明FAT10通过调控炎症反应促进肾缺血再灌注损伤的分子机制，为寻找和筛选新的药物靶点提供重要的科学依据。

缺血再灌注损伤（renal ischemia-reperfusion injury, IRI）是急性肾损伤（acute kidney injury, AKI）最常见病因之一，目前尚无理想的治疗措施。大量研究表明，炎症症反应在肾缺血再灌注损伤的发生发展中起关键作用。因此，深入研究炎症反应的调节机制以及如何有效抑制炎症反应强度，对于防治IRI导致的AKI具有重要意义。在本课题中，通过一系列体内体外实验，我们发现：① IRI后肾脏组织中类泛素蛋白FAT10的表达明显上调，且主要表达于肾小管上皮远端细胞；② 敲除FAT0明显减轻IRI导致的肾脏组织损伤，并显著改善肾功能；③ 敲除FAT0明显抑制IRI诱导的肾脏组织炎症反应；④ FAT10通过激活NF-κB信号通路的活化介导趋化因子RANTES的产生，进而导致IRI诱导的肾脏组织炎症反应；⑤RANTES是FAT10促进IRI诱导的肾脏组织损伤的关键分子。综合以上研究成果，本课题阐明了FAT10 在IRI过程中的作用及分子机制，揭示了在IRI过程中FAT10活化通过激活 NF-κB信号通路，促进趋化因子 RNATES 的产生，放大炎症反应，加重肾脏损伤的生物学过程。这些发现将为IRI导致急性肾损伤的防治提供新的思路，为相关药物的开发提供新的依据，具有重要的科学意义和临床价值。