摘要
目的:探索miR-199b 对HaCaT细胞增殖、迁移及对VEGFA、JAG1、SET信号通路的影响。方法:转染miR-199b 上调与下调慢病毒到人永生化表皮细胞(HaCaT)。分为miR-199b 上调(up)、miR-199b 下调(dowm)、空载对照(NC组)三个组,进行CCK8、凋亡、周期试验,并采用RT-PCR技术检测VEGFA、JAG1、SET基因的表达,采用WB技术检测miR-199b 对下游蛋白变化(如VEGFA、JAG1等分子)的影响。结果:miR-199b up组OD450/fold值显著高于NC组,差异有统计学意义(P<0.05)、miR-199b down组OD450/fold值略低于NC组,差异无统计学意义(P>0.05)。与NC组相比,G1期miR-199b up组与miR-199b down组的细胞增多(P<0.05);在S期,miR-199b up组及miR-199b down组的细胞减少(P<0.05);在G2/M期,miR-199b up组及miR-199b down组的细胞减少,差异均具有统计学意义(P<0.05)。与NC组比较,miR-199b up组凋亡细胞百分比下降(P<0.05)、miR-199b down组凋亡细胞百分比上升(P<0.05);SET在miR-199b up组的表达明显低于NC组,在miR-199b down组表达明显高于NC组。从定量PCR结果可以看出,HaCaT细胞中,miR-199b up组JAG1表达丰度略高于NC组,差异无统计学意义(P>0.05),miR-199b down组JAG1表达丰度明显高于NC组(P<0.05);miR-199b up组VEGFA表达丰度明显低于NC组(P<0.05)、miR-199b down组VEGFA表达丰度明显高于NC组(P<0.05);miR-199b up组SET表达丰度小于NC组(P>0.05)、miR-199b down组SET表达丰度明显大于NC组(P<0.05)。结论:miR-199b 可抑制VEGF、JAG1、SET蛋白的表达,为其靶基因,可促进细胞的增殖,延长细胞周期,并抑制细胞凋亡。
Objective To explore the effects of miR-199b on proliferation and migration of HaCaT cells,as well as VEGFA,JAG1 and SET signaling pathways.Methods Transfection of miR-199b mimics and inhibitor lentivirus into human immortalized epidermal cells HaCaT was performed.The study was divided into three groups:miR-199b up,miR-199b down and no-load control.CCK8,cycle and scratch test were conducted,and the expression of VEGFA,JAG1 and SET genes were detected by RT-PCR,and the downstream protein changes(such as VEGFA,JAG1 and other molecules)were detected by WB technology.Results The OD450/fold value of miR-199b up group was significantly higher than that of NC group(P<0.05).The OD450/fold value of miR-199b down group was slightly lower than that of NC group(P>0.05).In the G1 phase,cells in the miR-199b up and down group were increased compared to the control group(P<0.05).At the S stage,compared with the control group,the cells in the miR-199b up and down group were reduced(P>0.05).In the G2/M phase,compared with the control group,the cells in the miR-199b up and down group were reduced(P>0.05).Compared with the NC group,the percentage of apoptotic cells in the miR-199b up group decreased(P<0.05)and the percentage of apoptotic cells in the miR-199b down group increased(P<0.05).The expression of SET was significantly lower in the miR-199b up group than in the NC group,and significantly higher in the miR-199b down group than in the NC group.According to the quantitative PCR results,the expression abundance of JAG1 in the miR-199b up group was slightly higher than that in the NC group(P>0.05).The expression abundance of JAG1 in the miR-199b down group was significantly higher than that in the NC group(P<0.05).The expression abundance of VEGFA in the miR-199b up group was significantly lower than that in the NC group(P<0.05).The expression abundance of VEGFA in the miR-199b down group was significantly higher than that in the NC group(P<0.05).SET expression abundance of miR-199b up group was smaller than that of NC group(P>0.05).The expression abundance of SET in the miR-199b down group was significantly greater than that in the NC group(P<0.05).Conclusion miR-199b is the target gene of VEGF,JAG1 and SET proteins by inhibiting their expression.It can promote cell proliferation,prolong cell cycle and inhibit cell apoptosis.
作者
邱尧
岳毅刚
邵家松
霍群
张敏
QIU Yao;YUE Yi-gang;SHAO Jia-song;HUO Qun;ZHANG Min(Department of Plastic Surgery,the Affiliated Hospital of Guilin Medical University,Guilin 541001,Guangxi,China)
出处
《中国美容医学》
CAS
2019年第1期89-92,共4页
Chinese Journal of Aesthetic Medicine
基金
广西自然科学基金项目(项目名称:IPS细胞改善慢性创面血液供应及其作用机制研究
编号:2015GXNSFAA139113)
关键词
微小RNA
增殖
凋亡
分化
信号通路
慢性创面
miRNA
proliferation
apoptosis
differentiation
signaling pathways
chronic wound