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目的:检测不同模式加热联合甲基莲心碱(neferine,Nef)对耐药乳腺癌MCF-7/Adr细胞的增殖抑制及γH2AX和mdr-1/P-gp表达的影响。方法:采用MTT法检测金属模块加热、培养基加热、烤箱加热及水浴加热等不同模式加热联合10μg/mL Nef对经阿霉素培养的MCF-7/Adr细胞的增殖抑制,实时定量PCR检测mdr-1mRNA的表达,Western印迹法检测γH2AX及P糖蛋白(P-glycoprotein,P-gp)的表达。结果:42℃及45℃不同模式加热组较37℃培养组MCF-7/Adr细胞吸光度值明显下降,mdr-1/P-gp表达下降,γH2AX表达上调(均P<0.01)。加热联合10μg/mLNef组较单纯热疗组及单纯10μg/mLNef组MCF-7/Adr细胞吸光度值明显下降(均P<0.01),mdr-1/P-gp表达下降,γH2AX表达上调(均P<0.05)。42℃及45℃不同模式加热组比较,水浴加热组P-gp表达下降及γH2AX表达上调最为明显(P<0.05)。结论:加热能增加阿霉素对MCF-7/Adr细胞的细胞毒性反应,能明显增加MCF-7/Adr细胞DNA损伤且随温度升高而加剧;MCF-7/Adr细胞对不同模式的加热反应可能不同;热疗联合Nef能增敏阿霉素的化学治疗效果。
OBJECTIVE: To investigate the effects of different modes of heating combined with neferine (Nef) on the proliferation and the expression of γH2AX and mdr-1 / P-gp in MCF-7 / Adr cells. Methods: The proliferation of MCF-7 / Adr cells cultured with doxorubicin (10μg / mL Nef) was detected by MTT assay with metal mold heating, medium heating, oven heating and heating in a water bath. MTT assay was used to detect mdr -1 mRNA expression was detected by Western blot. The expression of γ-Hax2AX and P-glycoprotein (P-gp) were detected by Western blotting. Results: Compared with the control group, the absorbance of MCF-7 / Adr cells in 42 ℃ and 45 ℃ heat-treated groups were significantly decreased, while the expression of mdr-1 / P-gp was decreased and γH2AX was up-regulated. The absorbance of MCF-7 / Adr cells in heating combined 10μg / mLNef group was significantly lower than that of hyperthermia group and 10μg / mLNef group (all P <0.01), while the expression of mdr-1 / P-gp was decreased <0.05). Compared with heating at 42 ℃ and 45 ℃, P-gp expression and γH2AX up-regulation were the most significant (P <0.05) in water-heating group. CONCLUSION: Heating can increase cytotoxicity of doxorubicin to MCF-7 / Adr cells, increase DNA damage of MCF-7 / Adr cells and increase with increasing temperature. Heating of MCF-7 / Adr cells to different modes The reaction may be different; hyperthermia combined with Nef can enhance the chemotherapeutic effect of doxorubicin.