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杨 春, 邓绍平.hsa-miR-302a-3p靶向VEGFA抑制胃癌细胞增殖的机制研究.四川大学学报(医学版),2019,50(1):13-19
hsa-miR-302a-3p靶向VEGFA抑制胃癌细胞增殖的机制研究
Mechanism of hsa-miR-302a-3p-targeted VEGFA in the Inhibition of Proliferation of Gastric Cancer Cell
  
中文关键词:  hsa-miR-302a-3p VEGFA 胃癌细胞
英文关键词:hsa-miR-302a-3p VEGFA Gastric cancer cell
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中文摘要:
      目的 探讨过表达hsa-miR-302a-3p下调血管内皮生长因子A(VEGFA)对胃癌细胞SGC-7901存活、运动和上皮间质转化的调节作用。方法 采用细胞转染,将hsa-miR-302a-3p mimic转染于miR组细胞,pc-VEGFA转染于VEGFA组细胞,同时将两个基因共转染于miR+VEGFA组。采用RT-PCR和Western blot检测转染效率,生物信息学靶向预测和荧光素实验验证两个基因靶向关系,采用CCK-8法检测各组细胞增殖,Transwell检测各组细胞侵袭能力,划痕实验检测各组细胞迁移能力,显微镜下观察细胞的形态是否发生上皮间质转化(EMT),采用Western blot检测各组细胞存活相关蛋白Ki67和Caspase-3、EMT相关蛋白E-cadherin、Vimentin、N-cadherin和Snail以及VEGFA下游靶基因p-P38、p-MAPKAPK和p-Hsp27蛋白表达水平。结果 VEGFA是miR-302a-3p的预测靶位点;与对照组相比,miR组细胞数量、侵袭和迁移率均下降( P<0.05),VEGFA组细胞数量、侵袭和迁移率均升高( P<0.05);与VEGFA组相比,miR+VEGFA组细胞数量、侵袭和迁移率均下降( P<0.05),E-cadherin蛋白表达水平上调( P<0.05),Vimentin、N-cadherin和Snail蛋白表达水平下调( P<0.05),p-P38、p-MAPKAPK和p-Hsp27蛋白表达水平下调( P<0.05)。结论 hsa-miR-302a-3p通过抑制VEGFA的表达,抑制胃癌细胞SGC-7901的增殖、侵袭和迁移。
英文摘要:
      Objective To explore the role mechanism of hsa-miR-302a-3p overexpression in the inhibition of proliferation of gastric cancer cell SGC-7901 by targeted-regulating vascular endothelial growth factor A (VEGFA). Methods The cell transfection was used to transfect hsa-miR-302a-3p mimic into miR mimic group and transfect pc-VEGFA into VEGFA group, and the two genes were co-transfected into miR+VEGFA group. The transfection efficiency was detected by RT-PCR and Western blot. The bioinformatics targeting prediction and fluorescein assay were used to verify the targeting relationship between the two genes. Cell proliferation was detected by CCK-8 test, and Transwell assay was used to detect the invasion ability of each group, and scratch assay was used to detect the migration ability of each group. The morphology changes of epithelial-mesenchymal transition (EMT) in cells were observed under microscope. Western blot was used to detect the protein expression levels of survival-related proteins Ki67 and Caspase-3, EMT-related proteins E-cadherin, Vimentin, N-cadherin and Snail and VEGFA downstream target genes p-P38, p-MAPKAPK and p-Hsp27. Results VEGFA was the predicted target site of miR-302a-3p. Compared with control group, the number of cells, the invasion and migration rates were also reduced ( P<0.05) in miR mimic group, and the number of cells was increased ( P<0.05) as well as the invasion and migration rates in VEGFA group. Compared with VEGFA group, the number of cells, the invasion and migration rates were also decreased ( P<0.05) in miR+VEGFA group. The protein expression level of E-cadherin was up-regulated ( P<0.05) while the protein expression levels of Vimentin, N-cadherin and Snail were down-regulated ( P<0.05), and the protein expression levels of p-P38, p-MAPKAPK and p-Hsp27 were also down-regulated ( P<0.05). Conclusion hsa-miR-302a-3p overexpression can inhibit the proliferation and promote apoptosis of gastric cancer cell SGC-7901 by targeting negative regulation of VEGFA expression.
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