ABSTRACT
Objective To explore the impacts of over-expression of microRNA-7 (miRNA-7) on the sensitivity of cis-platin in esophageal carcinoma cell line TE-1, and the possible mechanism thereof. Methods Lipofectmin 2000 method was used to transient transfect with miRNA-7 mimic into esophageal cancer cell line TE-1, which was taken as transfection group, mimic negative control was taken as transfection conrtol group. The expressions of miRNA-7 and epidermal growth factor receptor (EGFR) mRNA were detected by RT-PCR in the above two groups and normal control group. The total EGFR and EGFR in cytoplasmic and nucleus were detected with Western blot assay in transfection group and transfection control group. CCK-8 was used to detect IC50 of cisplatin in transfection group and transfection control group. The expression of EGFR was observed with immunofluorescence confocal microscope in two groups. Results The miRNA-7 expression was signifi-cantly increased in transfection group than that of transfection conrtol group and control group. The expression of EGFR mRNA was significantly reduced in transfection group (P<0.001). The total EGFR was significantly decreased in transfec-tion group than that of transfection conrtol group. The level of nuclear EGFR was significantly increased ( P<0.01),and cyto-plasm EGFR expression was significantly decreased in transfection group than that of transfection control group ( P<0.05). CCK-8 results showed that after the over expression of miRNA-7 in TE-1, the IC50 of cisplatin (48 h) increased in transfec-tion group than that of control group (P<0.01). Immunofluorescence results showed that EGFG in nuclear was higher in transfection group than that of transfection control group but its expressions reduced in cell membrane and cytoplasm. Con-clusion The over-expressed miRNA-7 in esophageal cancer cells TE-1 can reduce cisplatin sensitivity by the increased EGFR in nuclear translocation.
ABSTRACT
Objective:To investigate the influence of miR-7 knock down on the development of CD 4+SP cells in the thymus in mice,and preliminary explore its possible mechanism.Methods:The changes of volume ,weight and total cell counts of thymus in miR-7 knock down (miR-7KD) mice were observed compared with Wild-type(WT)mice;the pathological changes of thymus were observed by HE staining.FACS analysis was performed on the proportion ,as well as the expression level of CD44 and CD62L,of thymus CD4+single positive (SP) cells.Meanwhile,the proliferation percentage of CD4+SP cells was measured by Ki-67 staining.The apoptosis percentage of CD4+SP cells was analyzed by FACS.The changes on the transduction of ERK 1/2 pathways were determined by Western blot.Results:Compared with WT mice ,the size,weight and total cell number of thymus were marked reduced in miR-7KD mice( P<0.05 );moreover ,pathological change also was presented.The proportion and total cell number of thymus CD 4+SP cells were marked decreased ( P<0.05 ).Furthermore ,the expression level of CD 44 and proliferation percentage ,as well as apoptosis percentage ,of CD4+SP cells were obviously increased (P<0.05),however,the expression level of CD62L of CD4+SP cells were decreased (P<0.05). Finally,the level of total ERK1/2 and phosphor-ERK1/2 was decreased obviously ( P<0.05 ).Conclusion: miR-7 knock down can affect the development of CD 4+SP cells in the thymus , which might be closely related to the cell activation state and altered the transduction of ERK1/2 pathways.