基础医学与临床 ›› 2017, Vol. 37 ›› Issue (8): 1098-1102.

• 研究论文 • 上一篇    下一篇

miR-328对高糖诱导HUVECs间质转分化的调控及信号机制

王亮,陈云肖,吴均芳,姜红,万欢,黄江燕,黄俊   

  1. 南昌大学第一附属医院
  • 收稿日期:2016-06-20 修回日期:2016-09-25 出版日期:2017-08-05 发布日期:2017-07-17
  • 通讯作者: 黄俊 E-mail:junhuang918@163.com
  • 基金资助:
    糖尿病心肌微血管内皮细胞间质转分化中相关 miRNA 的筛选及功能研究

Regulation of miR-328 on human umbilical vein endothelial cell induced by high glucose and its signaling mechanism in endothelial-mesenchymal transition

  • Received:2016-06-20 Revised:2016-09-25 Online:2017-08-05 Published:2017-07-17

摘要: 目的 探讨miR-328对高糖诱导人脐静脉内皮细胞(HUVECs)间质转分化的调控作用及机制。方法 培养HUVECs,构建携带miR-328基因的重组慢病毒,转染HUVECs。细胞分7组:正常葡萄糖;甘露醇;高浓度葡萄糖;miR-328;miR-328病毒阴性对照;高浓度葡萄糖+U0126;miR-328+U0126。免疫荧光双染色鉴定HUVECs间质转分化;RT-qPCR检测miR-328表达;Western blot检测Ⅰ、Ⅲ型胶原蛋白,MEK1/2、P-MEK1/2、ERK1/2、P-ERK1/2表达。结果1).经高糖处理HUVECs呈CD31、α-SMA染色双阳性;2). 与对照组比较,高糖组miR-328表达增高(p<0.05);与高糖及miR-328组比较,U0126处理后miR-328表达降低(p<0.05);3). 与对照组比较,高糖及miR-328组Ⅰ、Ⅲ型胶原蛋白表达增多(p<0.05);与高糖及miR-328组比较,U0126处理后Ⅰ、Ⅲ型胶原蛋白表达减少(p<0.05);4).与对照组比较,高糖及miR-328处理后p-MEK1/2、p-ERK1/2表达增高(p<0.05);而U0126处理则能抑制这一现象(p<0.05)。结论 高糖可诱导HUVECs间质转分化,同时miR-328表达增高;miR-328可诱导HUVECs间质转分化;HUVECs间质转分化与MEK1/2-ERK1/2信号通路有关。

关键词: 人脐静脉内皮细胞, miR-328, 内皮细胞间质转分化, MEK1/2-ERK1/2

Abstract: Objective The study is to investigate the rolel of miR-328 in endothelial mesenchymal transition (EndMT) induced by high glucose in human umbilical vein endothelial cells (HUVECs) and its signaling mechanism. Methods HUVECs were cultured in high glucose environment to induce EndMT; The recombinant lentiviruses were produced by miR-328 and antagomiR- 328 transfection of HUVECs. The experiment was divided into seven groups: normal glucose; mannitol group; high glucose; miR-328; miR-328 virus negative control; high glucose + U0126; miR-328 + U0126. Double immunofluorescent staining was used to determine expression of EndMT markers; Changes in miR-328 expression is examined by RT-qPCR; The expressions of typeⅠⅢ collagen, p-MEK1/2 and p-ERK1/2 are examined by Western blot. Results 1). Immunofluorescent staining showed that the HUVECs showed positive staining for CD31 and α-SMA in high glucose group. 2). Compared with the control group, the expression of miR-328 was up-regulated(p <0.05) in HUVECs treated by high glucose or miR-328. Compared with high glucose group or miR-328 group, miR-328 expression were less pronounced after treatment with U0126. 3). The expressions of typeⅠ/Ⅲ collagen increased in HUVECs treated by high glucose or miR-328 when compared with control group (p <0.05). Compared with high glucose group or miR-328 group, typeⅠ/Ⅲ collagen expressions were less pronounced after treatment with U0126. 4). The expressions of p-MEK1 / 2 and p-ERK1 / 2 were increased in HUVECs treated by high glucose or miR-328 in comparison to the control group (p <0.05); a lower expression of p-MEK1 / 2 and p-ERK1 / 2 were observed in U0126 group than in high glucose group or miR-328 group. Conclusion The phenomenon of EndMT in HUVECs is induced by high glucose, and the expression of miR-328 is increased at the same time; overexpression of miR-328 induced EndMT in HUVECs; miR-328 induced EndMT is related with MEK1 / 2-ERK1 / 2 signaling pathway.

Key words: HUVEC, miR-328, endothelial–mesenchymaltransition(EndMT), MEK1/2-ERK1/2