LncRNA HIF1A-AS2通过抑制miR-138-5p促进滋养层细胞的侵袭和上皮间充质转化

LncRNA HIF1A-AS2 promotes trophoblast invasion and epithelial-mesenchymal transformation by inhibiting miR-138-5p

目的研究长链非编码RNA(long non-coding RNA,lncRNA)缺氧诱导因子-1α-反义链2(hypoxia-inducible factor 1 alpha antisense RNA 2,HIF1A-AS2)对滋养层细胞侵袭和上皮间充质转化(epithelial-mesenchymal transition,EMT)的作用和机制。方法比较20个子痫前期(preeclampsia,PE)胎盘(PE组)和20个正常胎盘(对照组)组织中HIF1A-AS2及其预测的靶基因miR-138-5p的表达水平。选用人绒毛膜滋养层细胞系HTR-8/Svneo,在转染HIF1A-AS2过表达质粒、miR-138-5p拟似物和抑制物及各自阴性对照的HTR-8/SVneo细胞中,通过CCK-8检测细胞增殖、Transwell实验检测侵袭、Western blot检测细胞EMT标记分子和转录因子表达来研究HIF1A-AS2和miR-138-5p对滋养层细胞的作用和机制。通过荧光素酶基因报告实验验证HIF1A-AS2和miR-138-5p的结合关系。结果 PE胎盘中HIF1A-AS2显著降低(P<0.05),而miR-138-5p的表达显著增高(P<0.05)。过表达HIF1A-AS2能够抑制miR-138-5p的表达(P<0.05),促进HTR-8/SVneo细胞的侵袭能力(P<0.05),上调EMT标记分子Vimentin和N-cadherin以及EMT转录因子Twist1的表达水平(P<0.05)。此外,miR-138-5p抑制物与过表达HIF1A-AS2对HTR-8/SVneo细胞的作用相同,而miR-138-5p拟似物能够减弱过表达HIF1A-AS2对该细胞的作用。另外,HIF1A-AS2和miR-138-5p在HTR-8/Svneo细胞中可以直接结合。结论 LncRNA HIF1A-AS2能够促进滋养层细胞的侵袭和上皮间充质转化,其作用可能通过抑制miR-138-5p水平来实现。

Objective To investigate the effect and underlying mechanism of long non-coding RNA(lncRNA)hypoxia-inducible factor 1 alpha antisense RNA 2(HIF1 A-AS2)on trophoblast invasion and epithelial-mesenchymal transition(EMT).Methods The mRNA levels of HIF1 A-AS2 and its predicted targering miR-138-5 p in the placental tissues of preeclampsia(PE)patients(PE group,n=20)and normal placental tissues(control group,n=20)were detected with qRT-PCR.Human chorionic trophoblast cell line HTR-8/SVneo(epithelial-like cells with adherent growth)were employed to be transfected with HIF1 A-AS2 overexpression plasmid,and/or miR-138-5 p mimic,miR-138-5 p inhibitor as well as theirs corresponding negative controls.Then the cell proliferation,cell invasion and protein expression of EMT-associated molecules(Vimentin and N-cadherin)and EMT-transcription factor(Twist1)were measured by CCK-8 assay,Transwell assay,and Western blotting,respectively.The binding relationship between HIF1 A-AS2 and miR-138-5 p was verified by luciferase gene reporter assay.Results The mRNA level of HIF1 A-AS2 was significantly decreased,while that of miR-138-5 p was markedly increased in the PE placental tissues,compared with the normal placental tissues(P<0.05).Over-expression of HIF1 A-AS2 obviously inhibited the expression of miR-138-5 p,promoted cell invasion and enhanced the expression of Vimentin,N-cadherin,and Twist1 in the HTR-8/SVneo cells(P<0.05).The effects of miR-138-5 inhibitor showed similar effects on HTR-8/Svneo cells that of over-expression of HIF1 A-AS2,but these effects of the overexpression on HTR-8/Svneo cells could be weakened when miR-138-5 p mimic was introduced.Furthermore,the direct bonding between HIF1 A-AS2 and miR-138-5 p was identified in HTR-8/SVneo cells.Conclusion LncRNA HIF1 A-AS2 promotes trophoblast invasion and EMT,which may be fulfilled by its inhibition on the expression of miR-138-5 p.