MicroRNA-195 Functions As A Tumor Suppressor By Targeting VEGFA In Gastric Adenocarcinoma

Background and purpose:Gastric cancer(GC)is the second leading cause of cancer-related death and the fifth most common cancer worldwide.MicroRNAs(miRNAs)are a group of short,noncoding RNAs that negatively regulate gene expression post transcriptionally by binding to the 3'-untranslated regions(3'-UTR)of their target gene mRNAs.Angiogenesis is essential for cancer development and progression since adequate blood supply is necessary for cancer cell growth and metastasis.Without an adequate vascular supply,solid tumors can grow only to a critical size of 1-2 mm(or about 106 cells),primarily due to lack of oxygen and nutrients.Vascular endothelial growth factor(VEGF)(also known as VEGFA),a secreted dimeric glycoprotein,is the most important regulator of tumor angiogenesis.VEGFA can specifically bind to vascular endothelial cell receptor(VEGFR)on endothelial cells,and then induce the proliferation,survival and migration of endothelial cells.In GC,several microRNAs have found to affect the proliferation,migration and tube formation of vascular endothelial cells through paracrine pathways by targeting VEGFA.These microRNAs might be attractive targets for GC therapy.Therefore,the aim of our study was to screen and investigate the dysfunction miRNAs in GC and further to investigate the function and mechanism of miR-195 in GC.Methods:The difference expression level of miR-195 in GC tissues compared with the normal tissues was calculated by T test.The CCK-8 proliferation assay,scratch assay and Transwell assay were performed to detect the influence of miR-195 on the proliferation ability,migration ability and invasive ability of GC cell lines.The CCK-8 proliferation assay,scratch assay and tube formation assay were performed to detect the influence of GC cell lines paracrine pathway on the proliferation ability,migration ability and tube formation ability of HUVEC.Double luciferase reporter gene assay was used to evaluate whether miR-195 could directly target VEGFA gene.Results:1)A number of 124 differentially miRNAs profiles were found between GC tissues and normal tissues in TCGA database.Of these,75 were significantly up-regulated and 49 were significantly down-regulated in GC tissues compared with normal tissues.Bioinformatics analysis suggested that miR-195 may target VEGFA in gastric adenocarcinoma.2)The expression level of miR-195 in GEO datasets and clinical GC tissues was significantly down-regulated compared with that in normal gastric tissues.Moreover,the expression level of miR-195 in GC tissues was correlated with lymph node metastasis,TNM stage and prognosis of gastric cancer patients.3)The proliferation ability,migration ability and invasive ability of GC cell lines were inhibited after transfection of miR-195 mimics.Similarly,the proliferation ability,migration ability and tube formation ability of HUVEC were inhibited after cultured with the conditioned medium which was collected after transfection of miR-195 mimics in GC cell lines.4)Bioinformatics analysis combined with luciferase assays revealed that miR-195 directly target the 3'UTR of VEGF mRNA.MiR-195 regulates angiogenesis through regulaing the secretion of VEGFA in GC cell lines which was.Moreover,the expression of miR-195 was negatively correlated with the expression of VEGFA.Similarly,the expression of miR-195 was negatively correlated with MVD.Conclusions:1)The expression level of miR-195 in GC tumor tissues was significantly lower than that in normal gastric tissues.The expression level of miR-195 in GC tumor tissues was correlated with the lymph node metastasis,TNM stage and prognosis of GC patients.2)MiR-195 can inhibit the proliferation ability,migration and invasion ability of GC cell lines.MiR-195 can inhibit the tumor angiogenesis by regulating the secretion of VEGFA in GC cell lines.Therefore,our findings highlight the importance of miR-195 dysfunction in GC and implicate miR-195 as a potential therapeutic target for GC.