缺氧诱导因子-1α通过miR-126/JAK2-STAT3轴调控慢性肾病小鼠肾纤维化

Hypoxia-inducible factor-1αregulates renal fibrosis in mice with chronic kidney disease by inducing miR-126/JAK2-STAT3 axis

目的:探究缺氧诱导因子-1α(HIF-1α)通过miR-126/JAK2-STAT3轴促进慢性肾病小鼠肾纤维化的分子机制。方法:选取8周龄雄性C57/BL6小鼠,分为假手术组、模型组、HIF-1αmimic组、miR-126 mimic组、HIF-1α+miR mimic组、WP1066组、miR-126 mimic+WP1066组;用全自动生化仪检测小鼠肾24 h尿蛋白、血尿素氮(BUN)、血肌酐(SCr)水平,qRT-PCR检测肾组织内miR-126相对表达量,免疫印迹检测肾组织内HIF-α,肾纤维化关键蛋白fibronectin、collagen-Ⅰ、α-SMA,JAK2/STAT3信号通路关键蛋白表达;Masson染色分析小鼠肾病理形态及间质纤维化程度。结果:模型组模型建立后miR-126表达受到抑制,差异有统计学意义。与假手术组相比,模型组小鼠24 h尿蛋白、BUN、SCr水平均上调,肾组织肾小管、间质结构紊乱,肾纤维化明显,fibronectin、collagen-Ⅰ、α-SMA、p-JAK2、p-STAT3表达上调;HIF-1α过表达进一步上调24 h尿蛋白、BUN、SCr、fibronectin、collagen-Ⅰ、α-SMA、p-JAK2、p-STAT3表达;miR-126过表达下调24 h尿蛋白、BUN、SCr、fibronectin、collagen-Ⅰ、α-SMA、p-JAK2、p-STAT3表达;HIF-1α过表达可以逆转miR-126过表达抑制肾纤维化的趋势,差异有统计学意义。与模型组相比,JAK2/STAT3信号通路被阻断后,p-JAK2、p-STAT3、fibronectin、collagen-Ⅰ、α-SMA表达明显下调,miR-126的过表达可以进一步下调小鼠肾组织内p-JAK2、p-STAT3、fibronectin、collagen-Ⅰ、α-SMA表达,差异有统计学意义。结论:小鼠肾纤维化促进肾组织内HIF-1α高表达,HIF-1α通过抑制miR-126提升JAK2-STAT3信号通路活性,最终促进肾纤维化进展。

Objective:To investigate the molecular mechanism of hypoxia-inducible factor-1α(HIF-1α)promoting renal fibrosis in mice with chronic kidney disease by inducing miR-126/JAK2-STAT3 axis.Methods:Male C57/BL6 mice at 8 weeks old were divided into sham operation,model,HIF-1αmimic,miR-126 mimic,HIF-1α+miR mimic,WP1066,and miR-126 mimic+WP1066 groups.Using a fully automated biochemical analyzer,24-hour urinary protein,blood urea nitrogen(BUN),and serum creatinine(SCr)levels were measured in the mouse kidneys.MiR-126 expression in kidney tissues was assessed via qRT-PCR,and HIF-αexpression,along with renal fibrosis-related proteins(fibronectin,collagen-Ⅰ,α-SMA),and key proteins in the JAK2/STAT3 signaling pathway were detected through immunoblotting.Masson staining aided in the analysis of renal pathology and interstitial fibrosis in mice.Results:The expression of miR-126 was inhibited after UUO model was established,and the difference was statistically significant.Compared with the sham operation group,mice in the model group exhibited elevated levels of 24-hour urinary protein,BUN,and SCr.There was evident disruption in the structure of renal tubules and interstitium,accompanied by marked renal fibrosis.Expression levels of fibronectin,collagen-Ⅰ,α-SMA,p-JAK2,and p-STAT3 were up-regulated.Furthermore,overexpression of HIF-1αfurther elevated the expression of 24-hour urinary protein,BUN,SCr,fibronectin,collagen-Ⅰ,α-SMA,p-JAK2,and p-STAT3.Conversely,overexpression of miR-126 led to downregulation of these parameters.Additionally,overexpression of HIF-1αcould reverse the inhibitory effect of miR-126 overexpression on renal fibrosis,with statistical significance.Compared with the model group,blockade of the JAK2/STAT3 signaling pathway resulted in significant downregulation of p-JAK2,p-STAT3,fibronectin,collagen-Ⅰ,andα-SMA expression.Conclusions:Mouse renal fibrosis promotes high expression of HIF-1αin renal tissue,and HIF-1αenhances the activity of JAK2-STAT3 signaling pathway by inhibiting miR-126,ultimately promoting the progression of renal fibrosis.