Expression Of MiR-203a-5p And MiR-205-5p In Mycosis Fungoides And Their Biological Functions In HUT-78 Cell

Objective: To study the expression differences of miR-203a-5p and miR-205-5p in mycosis fungoides,paramycosis fungoides,eczema and normal prepuce tissues,and to verify the target genes by clinical samples,and to further explore the regulatory mechanism of miRNA in Hut-78 cells.Methods:(1)from January 2016 to may 2017 in the Department of Dermatology,people’s Hospital of Xinjiang Uygur Autonomous Region,patients with MF of erythema stage and normal skin tissue,chronic eczema skin tissue and normal skin tissue were divided into four groups.The expression levels of miR-203a-5p and miR-205-5p in the four groups were detected by gene detection.(2)Mi RNA and target gene annotation,mirwalk2.0,miRBase and other public human micro RNA gene database and literature reports were used to screen micro RNA target genes.The differences of gene and protein expression of E2F1,E2F3,ZEB2 and ccng1 in the four groups were detected by gene detection and protein detection.(3)Hut-78 cells were cultured in vitro.Mi R-203a-5p mimics,miR-203a-5p mimics NC,miR-205-5p mimics,miR-205-5p mimics NC,miR-203a-5p inhibitor,miR-203a-5p inhibitor NC,miR-205-5p inhibitor and miR-205-5p inhibitor NC were transfected into Hut-78 cells.In this study,MTT method was used to detect the cell growth status,flow cytometry was used to detect the changes of cell apoptosis and cycle,JC-1 method was used to detect the early apoptosis of cells,and edu method was used to detect the changes of cell proliferation rate in the process of DNA replication.(4)The expression of mir-203a-5p and miR-205-5p was detected by real-time PCR,and the protein expression of p53,E2F1,E2F3 and MDM2 was detected by protein detection.SPSS20.0 software was used to evaluate the final data of the experiment,P<0.05 was statistically significant.Results:(1)the expression of miR-205-5p and miR-203a-5p was low in the lesions of MF,which was significantly different from that in the adjacent tissues of MF,eczema and normal prepuce tissues.The expression of miR-203a-5p was higher in eczema than in MF and normal prepuce.(2)E2F1,E2F3,ZEB2 and ccng1 were highly expressed in the lesions of MF,which were significantly different from the adjacent tissues of MF,eczema and normal prepuce tissues.(3)Mi R-203a-5p mimics and miR-205-5p mimics significantly reduced the proliferation activity of Hut-78 cells.Mi R-203a-5p inhibitor and miR-205-5p inhibitor significantly increased the proliferation activity of Hut-78 cells in a time-dependent manner;the apoptotic rate of Hut-78 cells in miR-203a-5p mimics group and miR-205-5p mimics group was significantly increased,and the apoptotic rate of Hut-78 cells in miR-203a-5p inhibitor group and miR-205-5p inhibitor group was significantly decreased;the apoptotic rate of Hut-78 cells in miR-203a-5p mimics group and miR-205-5p mimics group was significantly decreased Mi R-203a-5p inhibitor group and miR-205-5p inhibitor group had no effect on the cell cycle of Hut-78 cells;the prophase apoptosis rate of Hut-78 cells in miR-203a-5p mimics group and miR-205-5p mimics group was significantly increased,and the prophase apoptosis rate of Hut-78 cells in miR-203a-5p inhibitor group and miR-205-5p inhibitor group was significantly decreased;miR-203a-5p inhibitor group had no effect on the cell cycle of Hut-78 cells Mi R-203a-5p inhibitor group and miR-205-5p inhibitor group had no effect;miR-203a-5p and miR-205-5p had a certain regulatory effect on Hut-78 cells,and the effect of mimics group was better than that of inhibitor group;miR-203a-5p had a higher regulatory effect on Hut-78 cells than miR-205-5p.The regulation of miR-203a-5p and miR-205-5p on Hut-78 cells can promote cell proliferation and induce cell apoptosis,and the apoptosis rate is higher than the proliferation rate.(4)Mi R-203a-5p and miR-205-5p mimics groups showed high expression in Hut-78 cells,while miR-203a-5p and miR-205-5p mimics groups showed low expression in Hut-78 cells;miRNA mimics group inhibited the protein expression of E2F1,E2F3 and p53,and promoted the expression of MDM2;miRNA inhibitor group promoted the expression of E2F1,E2F3 and p53,and inhibited the expression of MDM2;miRNA mimics group promoted the expression of E2F1,E2F3 and p53,and inhibited the expression of MDM2 In mimics group,the proliferation of Hut-78 cells was promoted by mediating p53 signaling pathway,and the high expression of miRNA could promote p53 induced apoptosis.Conclusion:(1)miR-203a-5p and miR-205-5p are differentially expressed in mycosis fungoides,paramycosis fungoides,eczema and normal prepuce.The target genes of these two micro RNAs are E2F1,E2F3,ZEB2 and ccng1.(2)There was a negative correlation between miR-203a-5p and miR-205-5p and target genes.(3)Mi R-203a-5p7 plays an important role in the regulation of cell proliferation and apoptosis.(4)Mi R-203a-5p and miR-205-5p can regulate the biological function of Hut-78 cells by regulating the activities of upstream and downstream factors of p53 signaling pathway.(5)The overexpression of miR-203a-5p and miR-205-5p may be the key miRNA in the regulation of mycosis fungoides,which can be used to control the pathogenesis of mycosis fungoides to provide a theoretical basis for the pathogenesis and early diagnosis of MF.