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目的:氯化钴(CoCl2)作用于人胰腺癌耐药细胞株Patu8988/5-FU致其化学性缺氧,观察其线粒体损伤及耐药性的影响。方法:采用梯度浓度CoCl2(0、6.25、12.50、25.00、50.00、100.00、200.00、400.00μmol/L)培养人胰腺癌耐药细胞株Patu8988/5-FU,MTT法测定Patu8988/5-FU细胞增殖活性和耐药性;JC-1荧光染色流式细胞仪分析线粒体膜电位变化;RT-PCR检测HIF-1α及多药耐药基因MDR1的表达情况;罗丹明外排实验检测Patu8988/5-FU细胞膜上的P-gp泵功能。结果:随着CoCl2浓度升高或以100μmol/L作用时间延长,Patu8988/5-FU增殖活性下降,线粒体膜电位去极化增加。常氧和缺氧时其对5-氟脲嘧啶(5-Fu)的耐药指数分别达到28.11±3.19、52.08±3.53(P<0.01);HIF-1α和MDR1 mRNA的表达亦明显高于常氧对照组(P<0.05)。罗丹明在缺氧细胞株内的积聚量低于常氧对照组(P<0.01)。结论:利用CoCl2模拟Patu8988/5-FU化学性缺氧存在浓度和时间依赖性;缺氧使Patu8988/5-FU获得性耐药性增加可能是通过引起HIF-1α表达增加和上调多药耐药基因MDR1的表达引起的。
AIM: To investigate the effects of cobalt chloride (CoCl2) on the chemosensitivity of human pancreatic cancer cell line Patu8988 / 5-FU induced by chemical hypoxia and to observe its mitochondrial damage and drug resistance. Methods: Human pancreatic cancer cell line Patu8988 / 5-FU was cultured with gradient concentrations of CoCl2 (0, 6.25,12.50,25.00,50.00,100.00,200.00,400.00μmol / L). The proliferation of Patu8988 / 5-FU cells The expression of HIF-1α and multidrug resistance gene MDR1 was detected by RT-PCR. The rhodamine efflux assay was used to detect the expression of Patu8988 / 5-FU P-gp pump function on the cell membrane. Results: The proliferation of Patu8988 / 5-FU decreased and the depolarization of mitochondrial membrane potential increased with the increase of the concentration of CoCl2 or the prolonged time of 100μmol / L. The resistance index of 5-Fu to normoxia and hypoxia reached 28.11 ± 3.19 and 52.08 ± 3.53 respectively (P <0.01), and the expressions of HIF-1α and MDR1 mRNA were also significantly higher than those of normal Oxygen control group (P <0.05). Rhodamine accumulation in hypoxic cell line was lower than that in normoxia control group (P <0.01). Conclusions: The concentration-dependent and time-dependent effects of CoCl2 on the chemosensitivity of Patu8988 / 5-FU cells to hypoxia can be attributed to the increased expression of HIF-1α and the upregulation of multidrug resistance Gene expression caused by MDR1.