低浓度活性氧逆转肾癌先天性多药耐药的实验研究

目的探讨低浓度活性氧对肾癌786-O细胞先天性多药耐药的逆转作用及相关机制。方法采用WST-1细胞增殖及细胞毒性检测试剂盒确定活性氧H2O2的非细胞毒性剂量,以及对786-O细胞药物敏感性的影响。罗丹明123实验检测细胞P糖蛋白（P-gp）功能,Western blot方法检测经典多药耐药基因（mdr1）产物P-gp的表达。结果在0.00001～0.1mmol/L浓度范围内,H2O2具有明显的促细胞生长作用,且显著增加了阿霉素及长春新碱的细胞毒性,0.02mmol/LH2O2孵育786-O细胞72h后其对阿霉素及长春新碱的药物敏感性分别增加至对照组的5.43及4.47倍,荧光染料罗丹明123的蓄积量显著增加,Western blot检测结果显示0.02mmol/LH2O2可抑制肾癌786-O细胞P-gp的表达。结论低浓度活性氧H2O2可部分逆转人肾癌786-O细胞先天性多药耐药对阿霉素的耐药性,其逆转机制与增加细胞内化疗药物浓度、抑制P-gp的表达有关。

Intrinsic MDR-reversing effect of lower concentration reactive oxygen species on human renal cell carci- noma cell line

Objective To investigate the reversal effect of lower concentration reactive oxygen species （ROS） on human renal cell carcinoma （RCC） cell line 786-0, a intrinsic-resistance cell line of human RCC, and its possible mechanisms. Methods Cytotoxicity of ROS and its sensibility to adriamycin （ADM） or vincristine （VCR） in 786-0 cell line were assessed by the WST-1 （tetrazolium salts） colorimetric assay using a Cell Counting Kit. The P-gp mediated efflux in RCC cells was deter- mined by measuring the retention of a fluorescent P-gp suhstrate, rhodamine 123 dye. P-gp produc- tion determined by Western blot. Results Lower concentrations of H202 （about 0. 000 01-0. 1 mmol/L） can enhanced cell growth and sensitized RCC cells to ADM and VCR, which dramatic in- creased the toxicity of ADM and VCR compared with single chemotherapeutic drugs treatments. Af- ter incubation with 0.02 mmol/L H202 for 72 h, the sensitivity of 786-0 to ADM or VCR was en- hanced by 5.43, 4.47 times, respectively （P〈0.01 vs control or drug alone）. The residual ratio of rhodamine 123 in 786-0 ceils was significantly increased. The expressions of P-gp in RCC cell line 786-0 incubate with lower concentration H2 02 were decreased when detected by Western blot. Conclusions Lower concentration ROS could partially reverse the multidrug resistance of RCC cell line 786-0 in vitro. Furthermore, the reversal effect may be due to the increase of intracellular accumulation of chemotherapeutic drugs and the decrease of the expressions of and P-gp.