5种生物碱胃癌多药耐药逆转剂的筛选及机制研究

Screening for five alkaloids as reversal agents against gastric cancer multiple drug resistance and study on their mechanism

目的研究骆驼蓬碱、去氢骆驼蓬碱、贝母素甲、贝母素乙和氧化苦参碱对肿瘤细胞多药耐药性(MDR)的逆转作用及机制。方法以胃癌亲本细胞系SGC-7901和MDR细胞系SGC-7901/VCR为细胞模型,采用MTT法检测上述5种生物碱的细胞毒活性及对MDR的逆转效果;用流式细胞仪检测MDR逆转效果最好的贝母素乙对肿瘤细胞内阿霉素(ADR)蓄积的影响;Western blotting法检测P-糖蛋白(P-gp)的表达;Hoechst荧光染色和细胞免疫荧光法检测贝母素乙诱导SGC-7901/VCR细胞凋亡情况。结果骆驼蓬碱、去氢骆驼蓬碱、贝母素甲、贝母素乙和氧化苦参碱均能不同程度地抑制SGC-7901和SGC-7901/VCR细胞的增殖,在非毒剂量下贝母素乙能够显著提高SGC-7901/VCR细胞对ADR的敏感性及细胞内ADR的浓度,降低P-gp表达。贝母素乙联合5-氟尿嘧啶(5-FU)给药可诱导SGC-7901/VCR细胞凋亡,凋亡细胞cleaved caspase-3呈高表达。结论贝母素乙具有作为胃癌MDR逆转剂的潜力,其逆转耐药的机制可能与下调P-gp表达和诱导细胞凋亡有关。

Objective To investigate five compounds,such as harmine（HM）,harmaline（HMI）,peimine（PM）,peiminine（PMI）,and oxymatrine（OMT）,for their reversal effect on multiple drug resistance（MDR） in SGC-7901 and SGC-7901/VCR cell lines and their relevant mechanism.Methods Taking SGC-7901 and SGC-7901/VCR cells as the models,the cytotoxicity and the reversal effects of the five compounds on MDR were examined by MTT assay.The effect of PMI on intracellular Adriamycin（ADR） accumulation and the expression of P-glucoprotein（P-gp） were respectively detected by flow cytometry and Western blotting.MDR cell apoptosis induced by PMI was analyzed by Hoechst fluorescence staining and immunofluorescence method.Results HM,HMI,PM,PMI,and OMT could inhibit the cell growth of SGC-7901 and SGC-7901/VCR.Furthermore,PMI could enhance the sensitivity of ADR,increase the intracellular ADR concentration,and decrease the expression of P-gp with nontoxic dosage in SGC-7901/VCR cell line.Fluorescence microscope and Hoechest assay suggested that PMI combined with 5-fluorouracil（5-FU） could induce SGC-7901/VCR cell apoptosis,and cleaved caspase-3 could be highly expressed in apoptotic cells.Conclusion PMI has the potential to be a promising as anti-MDR agent in gastric cancer by lowering P-gp expression and affecting cell apoptosis as well.