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In order to investigate the effects of vector-based hairpin small interference RNA(shRNA) on the reversal of multi-drug resistance(mdr) of A2780/Taxol cells,a novel vector pEGFP-H1/mdr1 containing mdr1-shRNA targeting at position 2943-2963 of mdr1 was designed and synthesized.Subsequently,A2780/Taxol cells were transfected with pEGFP-H1/mdr1,and the expression of mdr1 mRNA and P-gp was detected by using RT-PCR and Western blot respectively.MTT was used to measure the 50% inhibition concentration(IC50) of Taxol to A2780/Taxol cells.The results showed that at the 24th and 48th h after transfection,the expression of mdr1 mRNA was decreased to(52.1±1.0)% and(0.01±1.7)%,and that of P-gp decreased to(88.3±2.1)% and 0%,respectively.At the 48th h after transfection,the relative reversal rate of A2780/Taxol cells to Taxol was 69.54%.In vivo,the nude mice xenografts were injected with pEGFP-H1/mdr1,and then administrated Taxol.The tumor volume in pEGFP-H1/mdr1-transfected group was significantly reduced as compared with that in blank control group or pEGFP-H1-transfected group(807.20±103.16 vs 1563.78±210.54 or 1480.78±241.24 mm3,both P<0.01).These results suggested that transfection of pEGFP-H1/mdr1 could efficiently down-regulate the expression of mdr1 mRNA and P-gp in A2780/Taxol cells,and effectively restore the sensitivity of A2780/Taxol cells to Taxol both in vitro and in vivo.
In order to investigate the effects of vector-based hairpin small interference RNA (shRNA) on the reversal of multi-drug resistance (mdr) of A2780 / Taxol cells, a novel vector pEGFP-H1 / mdr1 containing mdr1-shRNA targeting at 2943 -2963 of mdr1 was designed and synthesized.Subsequently, A2780 / Taxol cells were transfected with pEGFP-H1 / mdr1, and the expression of mdr1 mRNA and P-gp was detected by using RT-PCR and Western blot respectively. MTT was used to measure the 50% inhibition concentration (IC50) of Taxol to A2780 / Taxol cells. The results showed that at the 24th and 48th h after transfection, the expression of mdr1 mRNA was decreased to (52.1 ± 1.0)% and (0.01 ± 1.7) %, and that of P-gp decreased to (88.3 ± 2.1)% and 0%, respectively. At the 48th h after transfection, the relative reversal rate of A2780 / Taxol cells to Taxol was 69.54% .In vivo, the nude mice xenografts were injected with pEGFP-H1 / mdr1, and then administered Taxol. The tumor volume in pEGFP-H1 / mdr1-transfected group was significan tly reduced as compared with that in blank control group or pEGFP-H1-transfected group (807.20 ± 103.16 vs 1563.78 ± 210.54 or 1480.78 ± 241.24 mm3, both P <0.01) .sese results suggested that transfection of pEGFP-H1 / mdr1 down-regulate the expression of mdr1 mRNA and P-gp in A2780 / Taxol cells, and effectively restore the sensitivity of A2780 / Taxol cells to Taxol both in vitro and in vivo.