柴胡皂苷-d对大鼠肝癌细胞H4-ⅡE细胞内Ca2+的影响研究

摘要

Objective To investigate the influences of saikosaponin-d (SS- D) on the free cytoplasmie concentration of Ca2+ in H4-HE cells of rat models. Methods Proper concentration of chemicals used in the study was assessed by thiazol blue ( MTT) method. Changes in free cytoplasmie Ca2+ concentration ( [ Ca2+ ] cyt) and intracellular stores Ca2+ were measured by Fluo - 3/AM load technique. Results MTT test showed that cell survival rates were all over 90% after treatment of SS - D at the concentration of 10, 30, 50, 70, 100 and 120μmol/L, respectively. The negative control group showed a increase of [ Ca ] cyt with mean fluorescence intensity ( MFI) of 35. 76 ± 1.37 , and the increase was even more obvious in the positive control group, which showed MFI of 74. 74 ± 1. 26. For other groups, the MFI of [Ca2+ ] cyt was 59.95 ± 1. 02,73. 89 ± 1. 35,80. 30 ±0. 04,93. 30 ±0. 83, respectively, showing an SS - D dose - dependent increase. MFI were significantly higher in positive control group and the treatment group than that in the negative control group (P＜0. 05). There was also significant differences in the MFI between treatment group 1,3,4 and that in the positive control group ( P＜0. 05). SS-D, LCA and UDCA can induce the release of Ca2+ from intracellular stores. Conclusion SS-D may regulate physiological activities of intracellular Ca2+ by increasing [ Ca2+] cyt level since it can induce Ca2+ influx.%目的 探讨柴胡皂苷-d(saikosaponin -D,SS - D)诱导大鼠肝癌细胞H4-ⅡE细胞内钙离子浓度([Ca2+ ]cyt)变化的影响研究.方法 通过噻唑蓝(MTT)比色实验选择合适的药物浓度、荧光指示剂Fluo - 3/AM检测[Ca2+] cyt及钙库中Ca2+的变化.结果 MTT比色结果显示,10、30、50、70、100μmol/L SS -D对细胞的存活率都在90％以上.第一大组中阴性对照组可引起细胞内钙离子浓度升高,其平均荧光强度(MFI)为35.76±1.37；阳性对照组可使其大幅度升高,其MFI值为74.74±1.26.不同浓度的SS -D也可使其不同程度升高,并且与SS -D呈浓度依赖性,其平均荧光强度(MFI)分别为59.95±1.02、73.89±1.35、80.30±0.04、93.30±0.83.阳性对照组及处理组的MFI值明显高于阴性对照组(P＜0.05)；处理组1、处理组3及处理组4 MFI值与阳性对照组比较差异有统计学意义(P＜0.05)；石胆酸(LCA)及SS -D、熊脱氧胆酸(UDCA)都能诱导细胞钙库中Ca2+释放.结论 SS -D能诱导细胞内Ca2+内流,增加[ Ca2+] cyt,调控细胞内Ca2+的生理活动.