The Study Of MiRNA 101 In Regulation Of CRMP4 Gene Promoter Methylation And Inhibition Of Invasion And Metastasis Of Prostate Cancer PC3 Cells

Objectives:Explore the correlation among MicroRNA-101,CRMP4 gene and CRMP4 gene promoter methylation status,make sure whether Micro RNA-101 can regulate CRMP4 gene promoter methylation ? To clarify that whether up-regulation of CRMP4 by miR-101 could prohibit metastasis and invasion competence in vitro?Methods:Constructed using the plasmid of pre-miRNA-101 and siRNA EZH2?Prostate cancer PC3 cell were transfected with pre-miRNA-101 mimic ? pre-mico RNA negative control,siRNA EZH2,siRNA negative control by using liposome Lipofectamine 2000?To deal with prostate cancer PC3 cells by using DZNEP drug,and to detect CRMP4 gene promoter methylation in prostate cancer PC3 cells by using Methylation-specific PCR(MSP)?Cell scratch test and Transwell were used to assess the changes of PC3 cancer cell Migration and invasion ability after transfection.Results:Constructed using the plasmid of pre-miRNA-101 and siRNA EZH2 and transfected into PC3 cells using Lipofectamine 2000.Transfection efficiency was more than 80%?After 48 hours,Pre-miRNA-101 group and siRNA EZH2 group results show that by using real-time RT-PCR :Compared Experimental group and control group in prostate cancer cells,the expression level of mRNA CRMP4 was significantly increased(P?0.05),and the expression level of mRNA EZH2 was significantly decreased(P?0.05)?And to the negative control group and blank control group,the expression of EZH2 and CRMP4 have No difference(P>0.05)?Methylation specific PCR(MSP)show that :transfection of pre-miRNA-101 and EZH2 siRNA plasmid and PC3 cells treated with DZNEP drug,The methylation of CRMP4 gene promoter region was significantly decreased compared with the control group(P?0.05),but the degree of methylation was not significantly different(P>0.05)?Cell scratch test and Transwell showed that after transfection of PC3 cells migration and invasion ability significantly lowered.(P?0.05)Conclusion:1 there have similar functions Between Pre-miRNA-101 and siRNA EZH2,they can reduce the expression of EZH2 mRNA,which in turn increases CRMP4 mRNA expression?Finally,inhibition of metastasis of prostate cancer.2 The methylation level of CRMP4 gene promoter was higher in metastatic prostate cancer;3 MiR-101 can regulate the CRMP4 methylation status of CRMP4 gene promoter,and then affect the expression of DNA gene.4 CRMP4 gene promoter methylation is closely related to the metastasis of prostate cancer,or it is helpful for the early diagnosis and prognosis evaluation of the patients.