CAJ | 학술논문

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Rh123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24 h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48 h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3 h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。
목적：통과대중화안경사독진행분리화각조분적초사，심조역전K562대아매소내약적활성성분KD-Ⅲ-1，위금후연구종류내약성전정공작기출。방법：통과응효분리득도적사독조분분별작용우K562대아매소내약주K562/A화민감주K562/S，사선유효활성조분；통과형광탐침Rh123측정P-gp단백활성화PI염색진일보학정해조분적역전K562/A적내약활성。결과：분별급여2μg/mL아매소( Adr)화각농도사독조분처리24 h후，가이발현사독조분대아매소민감주K562/S화내약주K562/A도유명현적억제작용，병정현출제량-효응관계。1μg/mL사독조분처리조여2μg/mL사독처리조억제작용명현；재약물지속작용48 h후대K562/A적활성잉유억제작용재0.5、1、2μg/mL적사독조분조표현적경가명현。통과Rho외배실험발현사독조독조평균형광강도(MFI)여음성대조조몰유명현차이，이2.5μg/mL사독조분조적MFI명현강저，여대조조상비구유통계학의의(P<0.05)。2.5μg/mL사독조독화분리조분분별대K562/S민감주화K562/A내약주작용3 h후，K562/S적PI염색양성솔명현승고，이K562/A적PI염색양성솔병몰유명현승고。결론：사독조분KD-Ⅲ-1대K562/A화K562/S세포균유명현억제적작용，억제작용가능여유도조망유관。
Objective:To explore the active ingredient, KD-Ⅲ-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 μg/mL cobra venom toxin yielded a marked inhibitory effect, which persisted for 48 hours on K562/A strains. This effect became more apparent when compared with the 0. 5 μg/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. 5μg/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.