Li Xiaoli, Chen Wenhui, Feng Jinfang, Zhao Bo
Department of Nephrology, Xiangyang No. 1 People's Hospital, Hubei University of Medicine, Xiangyang, China.
Department of Geriatrics, Xiangyang No. 1 People's Hospital, Hubei University of Medicine, Xiangyang, China.
Transl Androl Urol. 2020 Oct;9(5):2157-2165. doi: 10.21037/tau-20-918.
Renal ischemia/reperfusion (RI/R) injury are a common pathogenesis of acute kidney injury, which may cause renal parenchyma damage clinically. Hypoxia-inducible factor-1α (HIF-1α) has protective effects on cells in regulating the metabolism, angiogenesis, erythropoiesis, and anti-apoptosis of RI/R injury. However, the specific mechanisms for HIF-1α on RI/R injury are still unclear. This study aims to investigate the effects of HIF-1α overexpression on renal function injury, immune disorder, and mitochondrial apoptosis in RI/R rats.
The rat model of RI/R injury was set up. The lentivirus (LV) vector of HIF-1α overexpression was constructed, and then the LV was transfected to the model rats. The rats were randomly divided into four groups: the control group, RI/R group, RI/R + LV group, and RI/R + LV-HIF-1α group for later experiments. The mRNA levels of HIF-1α were detected by RT-PCR. Proteinuria, urea nitrogen, and serum creatinine levels were detected using the relative kit. Pathological damage was detected by HE staining. Apoptosis was detected by TUNEL staining. Levels of interleukin-6 (IL-6), interleukin-1β (IL-1β), tumor necrosis factor α (TNF-α) and interleukin-10 (IL-10) were detected by ELISA. Western blotting was used to detect the protein levels of HIF-1α, caspase-3, caspase-9, Bax, Bcl-2, and other proteins.
Compared with the control group, the mRNA and protein levels of HIF-1α in the RI/R group were increased significantly (P<0.05). Proteinuria, urea nitrogen, serum creatinine levels were increased significantly (P<0.05). The levels of IL-6, IL-1 beta, TNF-α were increased significantly (P<0.05). The ratios of cleaved caspase-3/caspase-3, cleaved caspase-9/caspase-9, and Bax/Bcl-2 were increased significantly (P<0.05). There was a significant increase in apoptosis rate and renal pathological tissue damage (P<0.05). Compared with RI/R+LV group, the mRNA and protein levels of HIF-1α in the RI/R+LV-HIF-1α group were increased significantly (P<0.05). Proteinuria, urea nitrogen, serum creatinine levels were decreased significantly (P<0.05). IL-6, IL-1 beta, TNF-α levels were significantly decreased (P<0.05). IL-10 level was significantly increased (P<0.05). The ratios of cleaved caspase-3/caspase-3, cleaved caspase-9/caspase-9, and Bax/Bcl-2 were significantly reduced (P<0.05), showing that the pathological damage degree and the apoptosis rate was significantly lower.
HIF-1α overexpression has protective effects on renal ischemia-reperfusion rats by improving pathological injury and immune function, reducing the release of inflammatory factors, and the expression of apoptotic proteins.
肾缺血/再灌注（RI/R）损伤是急性肾损伤的常见发病机制，临床上可导致肾实质损伤。缺氧诱导因子-1α（HIF-1α）在调节RI/R损伤的细胞代谢、血管生成、红细胞生成和抗凋亡方面对细胞具有保护作用。然而，HIF-1α对RI/R损伤的具体机制仍不清楚。本研究旨在探讨HIF-1α过表达对RI/R大鼠肾功能损伤、免疫紊乱和线粒体凋亡的影响。
建立RI/R损伤大鼠模型。构建HIF-1α过表达的慢病毒（LV）载体，然后将LV转染至模型大鼠。将大鼠随机分为四组：对照组、RI/R组、RI/R + LV组和RI/R + LV-HIF-1α组，用于后续实验。通过RT-PCR检测HIF-1α的mRNA水平。使用相关试剂盒检测蛋白尿、尿素氮和血清肌酐水平。通过HE染色检测病理损伤。通过TUNEL染色检测细胞凋亡。通过ELISA检测白细胞介素-6（IL-6）、白细胞介素-1β（IL-1β）、肿瘤坏死因子α（TNF-α）和白细胞介素-10（IL-10）的水平。采用蛋白质免疫印迹法检测HIF-1α、半胱天冬酶-3、半胱天冬酶-9、Bax、Bcl-2等蛋白的水平。
与对照组相比，RI/R组中HIF-1α的mRNA和蛋白水平显著升高（P<0.05）。蛋白尿、尿素氮、血清肌酐水平显著升高（P<0.05）。IL-6、IL-1β、TNF-α水平显著升高（P<0.05）。裂解的半胱天冬酶-3/半胱天冬酶-3、裂解的半胱天冬酶-9/半胱天冬酶-9和Bax/Bcl-2的比率显著升高（P<0.05）。细胞凋亡率和肾脏病理组织损伤显著增加（P<0.05）。与RI/R + LV组相比，RI/R + LV-HIF-1α组中HIF-1α的mRNA和蛋白水平显著升高（P<0.05）。蛋白尿、尿素氮、血清肌酐水平显著降低（P<0.05）。IL-6、IL-1β、TNF-α水平显著降低（P<0.05）。IL-10水平显著升高（P<0.05）。裂解的半胱天冬酶-3/半胱天冬酶-3、裂解的半胱天冬酶-9/半胱天冬酶-9和Bax/Bcl-2的比率显著降低（P<0.05），表明病理损伤程度和细胞凋亡率显著降低。
HIF-1α过表达通过改善病理损伤和免疫功能、减少炎症因子释放以及凋亡蛋白表达，对肾缺血再灌注大鼠具有保护作用。
