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目的探讨霍山石斛提取物(Water extracts from Dendrobium huoshanense,WEDH)对人喉癌细胞株Hep-2增殖和凋亡的影响及其机制。方法分别用3.75、7.50、15.00、30.00和60.00 mg/ml的WEDH处理Hep-2细胞,采用MTT法检测各组细胞的增殖水平;倒置相差显微镜观察细胞的形态学变化;Annexin V-FITC/PI双染色激光扫描共聚焦显微镜观察细胞凋亡;Fluo-3 AM标记激光扫描共聚焦显微镜观察细胞内游离Ca2+浓度([Ca2+])i;分光光度法检测细胞的Caspase-3活性。结果 WEDH可明显抑制Hep-2细胞增殖,且呈时间和剂量依赖性,作用24、48和72 h的IC50值分别为48.54、26.67和14.93 mg/ml;经WEDH作用48 h的细胞形态均发生显著改变,呈现典型的细胞凋亡特征,且呈浓度依赖性,细胞内[Ca2+]i浓度和Caspase-3活性均显著高于阴性对照组(P<0.05)。结论 WEDH对Hep-2细胞具有明显的抑制增殖和诱导凋亡作用,其机制可能与升高细胞内[Ca2+]i浓度,激活Caspase-3有关。
Objective To investigate the effects of Dendrobium huoshanense (WEDH) on proliferation and apoptosis of human laryngeal carcinoma cell line Hep-2 and its mechanism. Methods Hep-2 cells were treated with WEDH at doses of 3.75, 7.50, 15.00, 30.00 and 60.00 mg / ml respectively. The proliferation of the cells was detected by MTT assay. Morphological changes were observed by inverted phase contrast microscope. Annexin V-FITC / PI Cell apoptosis was observed by double-stained laser scanning confocal microscope. The intracellular free Ca2 + concentration ([Ca2 +] i) was measured by Fluo-3 AM laser scanning confocal microscope. Caspase-3 activity was detected by spectrophotometry. Results WEDH significantly inhibited the proliferation of Hep-2 cells in a time and dose-dependent manner. The IC50 values of WEDH at 48, 44, 72 and 72 h were 48.54, 26.67 and 14.93 mg / ml, respectively. (P <0.05). The concentration of [Ca2 +] i and the activity of Caspase-3 were significantly higher than that of the negative control group (P <0.05). Conclusion WEDH can significantly inhibit the proliferation and induce apoptosis of Hep-2 cells. The mechanism may be related to the increase of intracellular [Ca2 +] i concentration and the activation of Caspase-3.