Yang Chun, Deng Shao-Ping
Department of Gastroenterological Surgery, School of Medicine, UESTC&Sichuan Provincial People's Hospital, Chengdu 610072, China.
Sichuan Da Xue Xue Bao Yi Xue Ban. 2019 Jan;50(1):13-19.
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).
The cell transfection was used to transfect hsa-miR-302a-3p mimic into miR mimic group and transfect pc- into group, and the two genes were co-transfected into miR+ 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.
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 ( <0.05) in miR mimic group, and the number of cells was increased ( <0.05) as well as the invasion and migration rates in group. Compared with group, the number of cells, the invasion and migration rates were also decreased ( <0.05) in miR+ group. The protein expression level of E-cadherin was up-regulated ( <0.05) while the protein expression levels of Vimentin, N-cadherin and Snail were down-regulated ( <0.05), and the protein expression levels of p-P38, p-MAPKAPK and p-Hsp27 were also down-regulated ( <0.05).
hsa-miR-302a-3p overexpression can inhibit the proliferation and promote apoptosis of gastric cancer cell SGC-7901 by targeting negative regulation of expression.
探讨hsa-miR-302a-3p过表达通过靶向调控血管内皮生长因子A（VEGFA）抑制胃癌细胞SGC-7901增殖的作用机制。
采用细胞转染技术，将hsa-miR-302a-3p模拟物转染至miR模拟物组，将pc-转染至对照组，并将二者共转染至miR+组。通过RT-PCR和蛋白质印迹法检测转染效率。采用生物信息学靶向预测和荧光素酶检测验证二者的靶向关系。采用CCK-8法检测细胞增殖情况，采用Transwell法检测各组细胞的侵袭能力，采用划痕法检测各组细胞的迁移能力。在显微镜下观察细胞上皮-间质转化（EMT）的形态学变化。采用蛋白质印迹法检测生存相关蛋白Ki67和Caspase-3、EMT相关蛋白E-钙黏蛋白、波形蛋白、N-钙黏蛋白和Snail以及VEGFA下游靶基因p-P38、p-MAPKAPK和p-Hsp27的蛋白表达水平。
是miR-302a-3p的预测靶位点。与对照组相比，miR模拟物组细胞数量、侵袭率和迁移率均降低（<0.05），而对照组细胞数量增加（<0.05），侵袭率和迁移率也增加。与对照组相比，miR+组细胞数量、侵袭率和迁移率也降低（<0.05）。E-钙黏蛋白的蛋白表达水平上调（<0.05），而波形蛋白、N-钙黏蛋白和Snail的蛋白表达水平下调（<0.05），p-P38、p-MAPKAPK和p-Hsp27的蛋白表达水平也下调（<0.05）。
hsa-miR-302a-3p过表达可通过靶向负调控的表达抑制胃癌细胞SGC-7901的增殖并促进其凋亡。
