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目的:探讨抑制c-FLIP的表达对TRAIL诱导乳腺癌细胞MCF-7凋亡的影响。方法:重组腺病毒Ad-c-FLIP-siRNA和Ad-sTRAIL单独及联合感染对TRAIL耐药的乳腺癌细胞MCF-7,应用实时荧光定量聚合酶链反应(Real-time PCR)检测病毒感染后各组细胞内c-FLIP和TRAIL的mRNA表达变化;MTT法和结晶紫染色法检测MCF-7细胞活性,Hoechst 33258荧光染色检测各组细胞的凋亡情况。结果:与阴性对照组比较,c-FLIP-siRNA组和c-FLIP-siRNA+TRAIL组c-FLIP的mRNA相对表达量分别是(0.32±0.16)和(0.39±0.48)倍;TRAIL组和c-FLIP-siRNA+TRAIL组TRAIL的mRNA相对表达量分别是(96.21±1.54)和(87.33±1.66)倍;TRAIL组、c-FLIP-siRNA组及c-FLIP-siRNA+TRAIL组的抑制率(%)分别为(60.27±1.25)、(11.34±1.74)及(74.91±2.12)。对比阴性对照组的凋亡率(3.12±1.54),TRAIL组(12.79±2.46)和c-FLIP-siRNA+TRAIL组(25.50±3.17)组的凋亡率明显增高(P<0.05),c-FLIP-siRNA组(6.85±2.82)的凋亡率变化不明显,差异无统计学意义(P>0.05)。结论:siRNA抑制c-FLIP基因的表达能显著促进TRAIL对乳腺癌细胞MCF-7凋亡的诱导作用。
Objective: To investigate the effect of inhibiting the expression of c-FLIP on TRAIL-induced apoptosis of MCF-7 breast cancer cells. Methods: Single and combined infection of TRAIL-resistant breast cancer cells MCF-7 with recombinant adenovirus Ad-c-FLIP-siRNA and Ad-sTRAIL was detected by Real-time PCR after virus infection The expression of c-FLIP and TRAIL in each group were detected by MTT assay and crystal violet staining. The apoptosis of MCF-7 cells was detected by Hoechst 33258 staining. Results: Compared with the negative control group, the relative mRNA expression of c-FLIP in c-FLIP-siRNA group and c-FLIP-siRNA + TRAIL group was 0.32 ± 0.16 and 0.39 ± 0.48 times The relative mRNA expression levels of TRAIL in FLIP-siRNA + TRAIL group were (96.21 ± 1.54) and (87.33 ± 1.66) fold, respectively. The inhibition rates of TRAIL, c-FLIP-siRNA and c-FLIP- %) Were (60.27 ± 1.25), (11.34 ± 1.74) and (74.91 ± 2.12), respectively. Compared with negative control group, the apoptotic rate was significantly increased (3.12 ± 1.54), TRAIL group (12.79 ± 2.46) and c-FLIP-siRNA + TRAIL group (25.50 ± 3.17) The apoptosis rate of FLIP-siRNA group (6.85 ± 2.82) did not change significantly, the difference was not statistically significant (P> 0.05). CONCLUSION: siRNA inhibited the expression of c-FLIP gene and significantly promoted the apoptosis of MCF-7 breast cancer cells induced by TRAIL.