氯化钴化学模拟低氧对肿瘤细胞增殖及凋亡的影响

Influence of chemical hypoxia induced by cobalt chloride on the proliferation and apoptosis of tumor cells

目的观察氯化钴(CoCl2)对体外培养的肿瘤细胞增殖和凋亡的影响,探讨合理的体外化学模拟低氧模式。方法(1)MTT方法和流式细胞术检测不同浓度CoCl2在相应时间内对A549和HeLa细胞活力以及增殖和凋亡的影响。(2)免疫印迹测定HIF-1α和凋亡蛋白的表达。结果(1)CoCl2(≤200μmol/L)在24h内对肿瘤细胞活力影响较轻,增加浓度或延长处理时间明显降低细胞活力。(2)CoCl2(200μmol/L)引起细胞早期凋亡。增加浓度(800μmol/L)导致晚期凋亡和坏死。(3)CoCl2(200μmol/L)诱导两种肿瘤细胞HIF-1α上调,24h达高峰(P<0.05),增加浓度或延长处理时间则下调。在A549细胞CoCl2引起BAX/BCL-2和P53表达上调;P53不参与CoCl2引起的HeLa细胞凋亡。结论CoCl2对肿瘤细胞的增殖凋亡的影响呈时间和浓度依赖性,进行化学模拟低氧时,要考虑CoCl2的作用浓度和时间。

Objective To investigate the effect of cobalt chloride on tumor cell proliferation and apoptosis in vitro, to explore the reasonable strategy of chemical hypoxia induced by CoCl2. Methods Two tumor cell lines （A549 and HeLa） were respectively exposed to CoCl2 （0. 05 - 2 mmol/L） for different time period （4 - 48 h） , cells viability,proliferation and apoptosis were maesured by MTT and FCM methods. HIF-1α and related apoptosis proteins expression were detected by Western blot. Results Cells viability was weakly changed in low concentration （≤200 μmol/L）of CoCl2 within 24 h. However, higher dose or prolonged challenge of CoCl2 significantly decreased cell survival rate （ P 〈 0. 05 ）. Most of the damaged cells were early apoptosis population after CoCl2 （200 μmol/L） incubation within 24 h, majority of the damaged cells were late apoptosis and necrosis part after CoCl2 （800 μmol/L） treatment. CoCl2 （200 μmol/L） exposure implicated in up-regulating the expression of HIF- 1 α, BAX, P53 and down-regulating the expression of BCL-2 in A549 cell within 24 h. However, higher concentration （ - 400 μmo]./L） or prolonged COCl2 challenge lead to HIF-1α expression decreasing. Similar results were observed in HeLa cell, except P53. Conclusion Effect of CoCl2 on the tumor cells proliferation and apoptosis is doseand time-dependent. But prolonged or higher dose of cobalt exposure effect （HIF-1α expression）. The concentration and duration of CoCl2 tion in chemical hypoxia. A549 cell; HeLa cell; HIF-1α; apoptosis was not positive to expected chemical hypoxia administration should be taken into consideration in chemical hypoxia.