Study On The Effect Of Mir-145-5p Targeting DUSP6 On Proliferation,Migration,Invasion And Apoptosis Of Endometrial Carcinoma Ishikawa Cells

Objective To study the effects of micro Ribonucleic Acid-145-5p(micro RNA-145-5p,mi R-145-5p)on the proliferation,migration,invasion and apoptosis of endometrial carcinoma,and to investigate its effect on the expression of dual specific phosphatase 6(DUSP6).To lay a foundation for the study of mi R-145-5p targeted therapy of endometrial carcinoma.Methods1.The Ishikawa cell line of endometrial carcinoma was routinely cultured,and Lipofectamine 2000 method was used to transfect mimic,inhibitor and negative controls into Ishikawa cells,which interfered with mi R-145-5p expression.The experiment was divided into 5 groups: un-transfected group,mimic negative control group,mimic group,inhibitor negative control group and inhibitor group.q RT-PCR was used to detect the expression level of mi R-145-5p after transfection.2.MTT assay was used to detect the effect of mi R-145-5p on the proliferation of Ishikawa cells.3.Effect of mi R-145-5p on the migration ability of Ishikawa cells was observed by scratch assay.4.Transwell assay was used to detect the invasive ability of mi R-145-5p on Ishikawa cells.5.The apoptosis of Ishikawa on mi R-145-5p was assessed by flow cytometry.6.Screening for candidate target genes of mi R-145-5p by four bioinformatics online sites of mi RTar Base,Target Scan,Target Miner and mi RDB.7.q RT-PCR and Western Blot assay were used to detect the m RNA and protein expression of DUSP6 after endometrial carcinoma Ishikawa cell line transfection,and to verify the potential targets of mi R-145-5p.Western Blot method to detect the expression level of phosphorylated extracellular signal-regulated kinase1/2(p ERK1/2)downstream of DUSP6.Results1.The results of q RT-PCR showed that the expression level of mi R-145-5p in mimic group was significantly higher than that in un-transfected group and mimic negative control group(P<0.05).The expression of mi R-145-5p in inhibitor group was significantly lower than that in un-transfected group and inhibitor negative control group(P<0.05).This study was successfully up-regulated and down-regulated the expression of mi R-145-5p in endometrial carcinoma Ishikawa cell line.2.MTT assay confirmed that the cell proliferation ability of mimic group was lower than that of un-transfected group and mimic negative control group.Overexpression of mi R-145-5p inhibited Ishikawa cell proliferation(P<0.05).The cell proliferation ability of inhibitor group was higher than that of un-transfected group and inhibitor negative control group.The low expression of mi R-145-5p promoted Ishikawa cell proliferation(P<0.05).3.Scratch assay showed that the cell scratch area of mimic group was significantly larger than that of un-transfected group and mimic negative control group after 24 hours.Overexpression of mi R-145-5p inhibited the migration of Ishikawa cells(P<0.05).The cell scratch area of inhibitor group was significantly smaller than that of un-transfected group and inhibitor negative control group after 24 hours.The low expression of mi R-145-5p promoted Ishikawa cell migration(P<0.05).4.Transwell experiments showed that overexpression of mi R-145-5p,the number of invasion of Ishikawa cell in mimic group was significantly less than that in un-transfected group and mimic negative control group.Overexpression of mi R-145-5p could inhibit cell invasion(P<0.05).The low expression of mi R-145-5p,the number of invasion of Ishikawa cell in inhibitor group was significantly greater than that in un-transfected group and inhibitor negative control group.The low expression of mi R-145-5p expression could promote cell invasion(P<0.05).5.Flow cytometry experiments showed that the apoptosis rate in mimic group was significantly higher than that in un-transfected group and mimic negative control group.Overexpression of mi R-145-5p promoted apoptosis of endometrial carcinoma Ishikawa cells(P<0.05).The apoptosis rate in inhibitor group was significantly lower than that in un-transfected group and inhibitor negative control group.The low expression of mi R-145-5p inhibited apoptosis of Ishikawa cells(P<0.05).6.Bioinformatics online software suggests that DUSP6 is one of the target genes of mi R-145-5p.7.q RT-PCR and Western blotting showed that the expression of DUSP6 m RNA and protein in mimic group was significantly lower than that in un-transfected group and mimic negative control group.Overexpression of mi R-145-5p significantly inhibited the m RNA and protein expression levels of DUSP6 in Ishikawa cells(P<0.05).The expression of DUSP6 m RNA and protein in inhibitor group was significantly higher than that in un-transfected group and inhibitor negative control group.The low expression of mi R-145-5p promoted the m RNA and protein expression levels of DUSP6 in Ishikawa cells(P<0.05).Western Blot experiments showed that the protein expressions of DUSP6 and p ERK1/2 showed an opposite trend(P<0.05).Conclusions mi R-145-5p plays a tumor suppressor role in endometrial carcinoma Ishikawa cells.Overexpression of mi R-145-5p could inhibit Ishikawa cell proliferation,migration,invasion,and promote Ishikawa cell apoptosis.The low expression mi R-145-5p expression could promote Ishikawa cell proliferation,migration,invasion,and inhibit Ishikawa cell apoptosis.mi R-145-5p affect the development and progression of endometrial carcinoma by targeting regulates DUSP6 and indirectly regulates p ERK1 / 2 protein expressions.This study provides a new idea in the fields of proliferation,migration,invasion,apoptosis and prevention of endometrial carcinoma.mi R-145-5p may be a potential target for the treatment of endometrial carcinoma.