miR-145抑制VSMC增殖的作用及其相关信号通路研究

Antiproliferative Effect of miR-145 on Vascular Smooth Muscle Cells and the Signal Transduction Pathways

目的探讨miR-145对PDGF-BB诱导的大鼠原代血管平滑肌细胞(VSMC)的作用及丝裂原激活的蛋白激酶(MAPK)信号转导途径的作用。方法体外培养大鼠原代VSMC,再将细胞分为空白对照组、miR-NC组、PDGF-BB+miR-145组及PDGF-BB组。CCK-8法检测各组细胞的增殖情况;实时RT-PCR方法检测PCNA、c-Jun及SM22a的表达水平;Western印迹方法检测ERK1/2和p-ERK1/2的表达、JNK和p-JNK的表达以及p38MAPK和p-p38MAPK的表达。结果 miR-145过表达后能够抑制PDGF诱导的大鼠原代VSMC增殖,并下调VSMC增殖相关基因PCNA、c-Jun的表达、上调分化相关基因SM22a的表达;PDGF-BB诱导VSMC后,ERK、JNK、p38MAPK的磷酸化水平均明显上调,而转染miR-145慢病毒后再加PDGF刺激,ERK、JNK、p38MAPK的磷酸化水平均明显下调。结论 miR-145能够抑制去分化型VSMC中的MAPK信号通路,进而抑制VSMC的增殖。

Objective To explore the effects of miR - 145 on proliferation of primary cultured vascular smooth muscle cells （VSMC） stimulated by platelet - derived growth factor（PDGF）- BB and mitrogen - activated protein kinase （MAPK） activity in VSMCs. Methods Primary cultured VSMCs were divided into four groups： Control group, miR - NC group, PDGF - BB＋ miR - 145 group,and PDGF - BB group. VSMC proliferation was measured by CCK - 8. The mRNA expression of C - Jun, PCNA and SM22a was detected by real - time PCR. The protein expression level of ERK1/2 and p - ERK1/2 ,JNK and p- JNK,p38MAPK and p - p38MAPK were assessed by western blot. Results Overexpression of miR - 145 inhibited the proliferation of primary cultured VSMCs and could upgraded the expression of VSMC proliferation related genes and downgraded VSMC dedifferentiated related genes. The protein expression of phosphor- ERK1/2, phosphor- JNK, and phosphor - p38MAPK were increased in PDGF - BB group, but it reduced in PDGF- BB＋miR - 145 group. Conclusion This study demonstrated that miR - 145 could inhibit PDGF - induced proliferation of VSMC,which might be mediated by the inhibition of MAPK.