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JOURNAL ARTICLE
RESEARCH SUPPORT, NON-U.S. GOV'T
[Expression of microRNA 27a and its correlation with drug resistance in human ovarian cancer A2780/Taxol cells].
Zhonghua Fu Chan Ke za Zhi 2010 May
OBJECTIVE: To investigate the expression of microRNA 27a (miR-27a) and relationship with drug resistance in human ovarian cancer A2780/Taxol cells.
METHODS: A stem-loop-mediated real-time PCR was used to detect miR-27a expression in A2780 and A2780/Taxol cells. The cells were transfected with the mimics or inhibitors of miR-27a or negative control RNA (NC) by lipofectamine 2000. The expressions of MDR1 gene, P-glycoprotein (P-gp) and homeodomain-interacting protein kinase 2 (HIPK2) protein levels were measured by real-time PCR and western blot respectively. Methyl thiazolyl tetrazolium (MTT) assay was used to analyze drug sensitivity. Apoptosis analysis was measured by fluorescence activated cell sorter (FACS).
RESULTS: (1) miR-27a was an average of 2.2-fold higher expression level in A2780/Taxol cells than that in A2780 cells, with a significant difference between the two groups (P<0.05). (2) A2780/Taxol cells transfection with inhibitors of miR-27a showed that the levels of MDR1 mRNA was decreased by 39%, P-gp protein level [(26+/-5)%] decreased than that in the NC group [(43+/-7)%], HIPK2 protein level [(30+/-6)%] increased than that in the NC group [(19+/-4)%], the 50% inhibition concentration (0.5 micromol/L) was less than that in the NC group (6.8 micromol/L), apoptosis rate [(32.5+/-3.6)%] was higher than that in the NC group [(5.6+/-2.1)%], and there were significant differences between two groups (all P<0.05). (3) Transfection of A2780 cells with mimics of miR-27a led to increase MDR1 mRNA expression by 121% as compared with one transfection with NC (P<0.05).
CONCLUSION: The expression of miR-27a is upregulated in A2780/Taxol cells, which may regulate MDR1 and P-gp expression by targeting HIPK2.
METHODS: A stem-loop-mediated real-time PCR was used to detect miR-27a expression in A2780 and A2780/Taxol cells. The cells were transfected with the mimics or inhibitors of miR-27a or negative control RNA (NC) by lipofectamine 2000. The expressions of MDR1 gene, P-glycoprotein (P-gp) and homeodomain-interacting protein kinase 2 (HIPK2) protein levels were measured by real-time PCR and western blot respectively. Methyl thiazolyl tetrazolium (MTT) assay was used to analyze drug sensitivity. Apoptosis analysis was measured by fluorescence activated cell sorter (FACS).
RESULTS: (1) miR-27a was an average of 2.2-fold higher expression level in A2780/Taxol cells than that in A2780 cells, with a significant difference between the two groups (P<0.05). (2) A2780/Taxol cells transfection with inhibitors of miR-27a showed that the levels of MDR1 mRNA was decreased by 39%, P-gp protein level [(26+/-5)%] decreased than that in the NC group [(43+/-7)%], HIPK2 protein level [(30+/-6)%] increased than that in the NC group [(19+/-4)%], the 50% inhibition concentration (0.5 micromol/L) was less than that in the NC group (6.8 micromol/L), apoptosis rate [(32.5+/-3.6)%] was higher than that in the NC group [(5.6+/-2.1)%], and there were significant differences between two groups (all P<0.05). (3) Transfection of A2780 cells with mimics of miR-27a led to increase MDR1 mRNA expression by 121% as compared with one transfection with NC (P<0.05).
CONCLUSION: The expression of miR-27a is upregulated in A2780/Taxol cells, which may regulate MDR1 and P-gp expression by targeting HIPK2.
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