钙池操纵的Ca^(2+)通道对人肝癌细胞增殖的影响

Effect of store-operated Ca^(2+) channels on proliferative activity of hepatoma cells

目的:对比研究人肝癌细胞与人正常肝细胞的钙池操纵的Ca2+通道(store-operated Ca2+ channel,SOC)功能改变,探讨SOC对肝癌细胞增殖能力的影响.方法:培养人肝癌细胞(SMMC7721)与人正常肝细胞(HL-7702),应用膜片钳技术检测两组细胞的细胞膜SOC电流,应用激光共聚焦显微镜检测两组细胞的细胞内游离Ca2+离子浓度,应用MTT法检测两组细胞的增殖能力,应用流式细胞仪检测两组细胞的增殖周期.结果:人肝癌细胞的SOC电流密度为19.36pA/pF±4.9pA/pF,明显高于人正常肝细胞的电流密度8.90pA/pF±2.78pA/pF;人肝癌细胞的细胞内钙荧光强度增加31.81%±8.89%,明显高于人正常肝细胞的21.58%±6.01%;MTT生长曲线显示从第3天开始,人肝癌细胞的增殖能力明显高于人正常肝细胞,且时间越长,增殖能力差别越大;流式细胞仪研究提示人肝癌细胞的S期细胞比例(S-phase cells ratio,SPF)、增殖指数(proliferation index,PI)明显大于人正常肝细胞,从而提示人肝癌细胞的增殖能力明显高于人肝细胞.结论:与人正常肝细胞相比,人肝癌细胞的SOC电流密度增大、细胞内游离Ca2+浓度升高、增殖能力增强,提示人肝癌细胞增殖能力提高与其SOC功能增强有关.

AIM： To investigate the changes in the function of store-operated Ca2＋ channels （SOCs） between human hepatoma cells and human liver cells and to dicuss the effect of SOCs on the proliferative ability of hepatoma cells. METHODS： Cultured human hepatoma cells （SMMC7721） and human liver cells （HL7702） were used in this study. Membrant current of SOCs was detected using the patch-clamp technique. Intracellular free Ca2＋ concentration was determined using laser scanning confocal microscopy. Cell proliferation was assessed by MTT assay, and cell cycle progression was detected by flow cytometry. RESULTS： The SOC current density was signifi-cantly higher in human hepatoma cells than in human liver cells （19.36 pA/pF ± 4.99 pA/pF vs 8.90 pA/pF ± 2.78 pA/pF, P 0.05）. The increase in intracellular calcium fluorescence intensity was also significantly higher in human hepatoma cells than in human liver cells （31.81% ± 8.89% vs 21.58% ± 6.01%, P 0.05）. MTT growth curve showed that the proliferative ability of human hepatoma cells was significantly higher than that of human liver cells from the third day, and the difference was increasing with the prolongation of time. Flow cytometry analysis indicated that S-phase fraction （SPF） and proliferation index （PI） were significantly greater in human hepatoma cells than in human liver cells, suggesting that the proliferative ability of hepatoma cells is much high than that of human liver cells. CONCLUSION： Compared to human liver cells, human hepatoma cells have significantly increased SOC current density, intracellular free Ca2＋ concentration, and proliferative ability, indicating that the enhancement of proliferative ability of human hepatoma cells may be related to the enhancement of SOC function