The Inhibition Of Growth And Invasion Of Human Endometrial Carcinoma Cells By Silencing Ang-2 Gene Using RNA Interference Combined With Artesunate

Objective:To observe the inhibition effect of cellular growth and invasion on the human endometrial carcinoma cells(Ishikawa) in vitro after induced by inhibiting the expression of Ang-2 gene with RNA interference,artesunate(Art) and cooperation of the two former.Methods:Experiment groups were divided into five groups:blank control group,negative control group,pCMV-Ang-2 group,Art group and cooperation group.The designed recombinant shRNA targeting Ang-2 gene and scramble sequence were transfected into Ishikawa cells.The proliferation,morphology,apoptosis,cell cycle,invasion,expression of Ang-2 on mRNA and protein level were detected by MTT,fluorescence microscope,FCM,transwell, RT-PCR and Western blotting technology respectively.Results:(1)pRNAT-CMV3.2-Neo-Ang-2 and pRNAT-CMV3.2-Neo-neg were successfully constructed and transfected into Ishikawa cells.The expression of Ang-2 mRNA on Ishikawa cells was suppressed remarkably while the negative control had no obvious change.(2) After induced by the pCMV-Ang-2,the Art and the cooperation group,The cell proliferation all were significantly inhibited in time-dependent manners,the tendency of inhibition gradually increased with the time.But the inhibition effect of the cooperation group was markedly better than the others(P＜0.05).(3) After induced by the pCMV-Ang-2,the Art and the cooperation group,the apoptosis of Ishikawa cells were obvious,the apoptotic rates of the cooperation group were markedly better than the others(P＜0.05).At the same time,cell apoptosis was observed by fluorescence microscope,even resulting in apoptotic bodies,however,the apoptotic rates maintained low level in the control groups.(4) After induced by the pCMV-Ang-2,the Art and the cooperation group,the cell cycle of Ishikawa cells changed significentedly.In the pCMV-Ang-2 group and cooperation group,G1 phase cell proportion increased,while the S phase decreased;in the Art group,G2 phase cell proportion increased gradually,while the S phase decreased gradually.(5) The expression level of Ang-2 mRNA and protein decreased,the inhibition effect of the cooperation group was evidently better than the others(p＜0.05).(6) After induced by the pCMV-Ang-2,the Art and the cooperation group,the invasion obviously decreased.Conclusions:The pCMV-Ang-2,the Art and the cooperation could decrease the expression of Ang-2 gene on Ishikawa cells and the invasion of Ishikawa cells was inhibited.Accordingly,the proliferation of Ishikawa cells was inhibited,the cell cycle changed and the apoptotic rates increased.In a word,Ang-2 gene was related with invasion and metastasis of endometrial carcinoma,it could induce apoptotis and differentiation of endometrial carcinoma Ishikawa cells While the RNA interference targeting Ang-2 gene and the Art are likely to be effective clinical treatments of endometrial carcinoma,the cooperation of the two forbodes a more gorgeous prospect of treating tumors by integrated Chinese-western treatment.