miR-203靶向调控VEGFA对滋养细胞增殖、侵袭和转移的抑制作用

Inhibitory effect of miR-203 on proliferation,invasion and metastasis of trophoblast cells through targeting VEGFA

目的探讨miR-203对滋养层细胞增殖、侵袭及迁移的影响,阐明miR-203与血管内皮生长因子A(VEGFA基因的靶向关系。方法选取2019年8月—2020年8月莆田学院附属医院30例子痫前期(PE)患者胎盘组织及20名正常孕妇胎盘组织、人正常绒毛膜滋养细胞HTR-8/SVneo为研究对象。实验分为NC组(转染miR-NC空质粒),miR-203组(转染miR-203-mimics),si-NC组(转染si-NC)和si-VEGFA组(转染si-VEGFA),MTT检测细胞增殖能力;Transwell小室实验检测细胞迁移、侵袭能力;采用实时荧光定量PCR检测细胞、组织中miR-203、VEGFA mRNA表达水平;Western blot检测细胞VEGFA表达水平。生物信息预测miR-203与VEGFA的靶向作用关系,并使用荧光素酶报告实验验证。结果 PE组胎盘组织miR-203水平高于正常孕妇胎盘组织(P<0.01),PE组胎盘组织VEGFA-mRNA水平低于正常胎盘组织(P<0.01)。与NC组、si-VEGFA组相比,48 h和72 h时,miR-203组、si-VEGFA组细胞增殖能力明显降低(P<0.01)。与NC组、si-VEGFA组相比,miR-203组、si-VEGFA组细胞迁移和侵袭能力明显降低(P<0.01)。双荧光素酶报告基因检测结果提示VEGFA为miR-203的靶基因。结论 miR-203在PE患者胎盘组织中高表达,过表达miR-203可能通过抑制VEGFA抑制滋养层细胞的增殖、侵袭和迁移。

Objective To investigate the effect of miR-203 on the proliferation,invasion and migration of trophoblast cells,and clarify the targeting relationship between miR-203 and VEGFA.Methods A total of 30 placental tissue specimens frompatients with pre-eclampsia,20 placental tissue specimens fromnormal pregnant women and HTR8 SVneo cells were selected as the research objects.HTR8 SVneo cells were divided into NC group(transfected with miR-NC),miR-203 group(transfected with miR-203 mimics),si-NC group(transfected with si-NC) and si-VEGFA group(transfected with si-VEGFA).MTT was used to detect cell proliferation.Transwell chamber test was used to detect cell migration and invasion.The expression levels of miR-203 and VEGFA mRNA in cells and tissues were detected by real-time quantitative PCR.Western blot was used to detect the expression of VEGFA.The relationship between miR-203 and VEGFA was predicted by bioinformatics and verified by luciferase reporter assay.Results The level of miR-203 in placental tissue of PE group was higher than that of normal placenta(P<0.01).The level of VEGFA mRNA in PE group was lower than that in normal placenta(P<0.01).At 48 h and 72 h,compared with NC group,si-VEGFA group,the proliferation ability of miR-203 group and si-VEGFA group decreased significantly(P <0.01).Compared with NC group and si-VEGFA group,the migration and invasion ability of miR-203 and si-VEGFA groups were significantly decreased(P<0.01).Double luciferase reporter gene analysis showed that VEGFA was the target gene of miR-203.Conclusion miR-203 is highly expressed in placenta of PE patients.Overexpression of miR-203 may inhibit the proliferation,invasion and migration of trophoblast cells by inhibiting VEGFA.