蛇毒组分对K562阿霉素敏感株和耐药株的抑制作用

目的：通过对中华眼镜蛇毒进行分离和各组分的初筛，寻找逆转K562对阿霉素耐药的活性成分KD-Ⅲ-1．为今后研究肿瘤耐药性奠定工作基础。方法：通过凝胶分离得到的蛇毒组分分别作用于K562对阿霉素耐药株K562／A和敏感株K562／S，筛选有效活性组分；通过荧光探针Rhl23测定P-gP蛋白活性和PI染色进一步确定该组分的逆转K562／A的耐药活性。结果：分别给予2μg／mL阿霉素（Adr）和各浓度蛇毒组分处理24h后，可以发现蛇毒组分对阿霉素敏感株K562／S和耐药株K562／A都有明显的抑制作用，并呈现出剂量-效应关系。1μg／mL蛇毒组分处理组与2μg/mL蛇毒处理组抑制作用明显；在药物持续作用48h后对K562／A的活性仍有抑制作用在0．5、1、2μg／mL的蛇毒组分组表现的更加明显。通过Rho外排实验发现蛇毒粗毒组平均荧光强度（MFI）与阴性对照组没有明显差异，而2．5μg／mL蛇毒组分组的MFI明显降低，与对照组相比具有统计学意义（P〈0．05）。2．5μg／mL蛇毒粗毒和分离组分分别对K562／S敏感株和K562／A耐药株作用3h后。K562／S的PI染色阳性率明显升高，而K562／A的PI染色阳性率并没有明显升高。结论：蛇毒组分KD-Ⅲ-1对K562／A和K562／S细胞均有明显抑制的作用，抑制作用可能与诱导凋亡有关。

Snake venom components inhibits growth of K562 adriamycin-sensitive and- resistant strains

Objective： To explore the active ingredient, KD-m-1, for reversal of K562 resistance to adriamycin （Adr） via separation and primary screening of individual components of cobra venom, thus offering the sound basis for future study on tumor drug resistance. Methods： The K562 Adr-resistant strains, K562/A, and-sensitive strains, K562/S, were incubated with isolated snake venom components derived from gel separation, for exploration of the effective component. The P-gp protein activity was measured by the fluorescent probe Rh123, and the PI staining was applied to determine the resistance reversal capacity of the components of K562/A. Results ： Following incubation with 2 μg/mL Adr and cobra venom toxin of various concentrations, we found that the latter dose-dependently inhibited the growth of K562/A and K562/S strains at hour 24. The 1μg/ mL and 2 p,g/mL cobra venom toxin yielded a marked inhibitm effect, which persisted for 48 hours on K562/A strains. This effect became more apparent when compared with the 0.5μg/ml cobra venom toxin group. Rho efflux experiments failed to show significantly different mean fluorescence intensity （ MFI ） ratios between the venom crude toxin group and negative control group. The MFI of 2.5μg/mL cobra venom group was significantly lower than that in control group （ P 〈 0.05 ）. Incubation with 2.5 μg/mL crude toxin and isolated toxin yielded a higher rate of PI-positive staining K562/S, but not K562/A, strains. Conclusion： The venom toxin component, KD-Ⅲ-1, significantly suppresses the growth of K562/A and K562/S cell lines possibly via induction of apoptosis.