piRNA-823通过TGF-β1/Smad2/3信号通路抑制高糖诱导的人脐静脉内皮细胞内皮-间充质转化和血管生成

piRNA-823 inhibits high glucose induced endothelial-mesenchymal transition and angiogenesis in human umbilical vein endothelial cells through TGF-β1/Smad2/3 signaling pathway

摘要:

目的]探讨piRNA-823在高糖处理的人脐静脉内皮细胞(HUVEC)表型转化中的生物学功能及其分子机制。 [方法]将HUVEC置于33.3 mmol/L高糖培养基中孵育72 h。采用RT-qPCR检测piRNA-823的相对表达水平,采用Western blot检测内皮细胞标志物、间充质细胞标志物和转化生长因子β1(TGF-β1)信号通路相关蛋白的表达变化,利用划痕实验和Transwell实验检测细胞迁移能力的变化,通过血管生成实验评估新生血管形成情况。将piRNA-823 mimic(piRNA-823过表达)转染至HUVEC中,分析其对高糖诱导的内皮-间充质转化(EndMT)及血管生成的影响。进一步采用TGF-β1激活剂(SRI011381)和抑制剂(SB525334)干预,以验证piRNA-823是否通过调控TGF-β1/Smad2/3信号通路发挥作用。 [结果]piRNA-823 mimic显著抑制高糖诱导的HUVEC活力、增殖和迁移能力以及血管生成。piRNA-823 mimic能够抑制高糖诱导的HUVEC EndMT,具体表现为上调内皮细胞相关标志物的表达,同时下调间充质细胞标志物的表达。划痕实验、Transwell实验和血管生成实验进一步证实piRNA-823 mimic可有效逆转高糖诱导的HUVEC增殖、迁移能力的增强以及新生血管数量的增加。机制研究发现,TGF-β1激活剂可部分逆转piRNA-823 mimic的保护效应,TGF-β1抑制剂则能增强其作用,表明piRNA-823通过抑制TGF-β1/Smad2/3信号通路的激活发挥调控作用。 [结论]piRNA-823通过抑制TGF-β1/Smad2/3信号通路的激活,显著抑制高糖诱导的HUVEC EndMT、细胞增殖和迁移能力以及血管生成。

Abstract:

Aim To investigate the biological function and molecular mechanisms of piRNA-823 in the phenotypic transformation of human umbilical vein endothelial cells (HUVEC) induced by high glucose. Methods HUVEC were incubated in high glucose (33.3 mmol/L) culture medium for 72 h. The relative expression levels of piRNA-823 were detected by RT-qPCR, the expression changes of endothelial cell markers, mesenchymal cell markers and proteins related to transforming growth factor-β1 (TGF-β1) signaling pathway were detected by Western blot, the changes of cell migration ability were evaluated by scratch and Transwell assays, the formation of new angiogenesis were assessed through angiogenesis experiments. piRNA-823 mimic (overexpression of piRNA-823) were transfected into HUVEC to analyze their effects on high glucose induced endothelial-mesenchymal transition (EndMT) and angiogenesis. Further intervention was performed using TGF-β1 activator (SRI011381) and inhibitor (SB525334) to verify whether piRNA-823 exerts its effect by regulating the TGF-β1/Smad2/3 signaling pathway. Results piRNA-823 mimic significantly inhibited the viability, proliferation, migration and angiogenesis of HUVEC induced by high glucose. The piRNA-823 mimic inhibited high glucose induced EndMT in HUVEC, characterized by upregulation of endothelial cell markers and downregulation of mesenchymal cell markers. Scratch experiments, Transwell experiments and angiogenesis experiments further confirmed that piRNA-823 mimic could effectively reverse high glucose induced HUVEC proliferation, migration ability enhancement, and increase in the number of new angiogenesis. Mechanistic studies revealed that the TGF-β1 activator partially reversed the protective effect of piRNA-823 mimic, whereas the TGF-β1 inhibitor enhanced its effect, suggesting that piRNA-823 exerts its regulatory role by suppressing the activation of the TGF-β1/Smad2/3 signaling pathway. Conclusion piRNA-823 significantly inhibits high glucose induced EndMT, proliferation, migration and angiogenesis in HUVEC by suppressing the activation of TGF-β1/Smad2/3 signaling pathway.