秋水仙碱对大鼠肾细胞NRK体外毒性机制初步探讨

Preliminary toxicological study of colchicine in rat renal cells NRK

目的：考察秋水仙碱体外肾毒性及其主要机制。方法：体外培养大鼠肾细胞NRK,通过MTT试验、细胞形态学改变、乳酸脱氢酶释放率、Hoechst/PI双染色法、流式细胞术考察秋水仙碱对大鼠肾细胞NRK是否具有毒性作用,进而阐明秋水仙碱肾毒性产生的可能机制。结果：秋水仙碱在0.1~10μmol/L浓度范围内,对NRK细胞的抑制率呈浓度依赖性。1μmol/L处理组细胞形态学发生明显改变,乳酸脱氢酶释放率随着给药浓度的增加也逐渐增加。Hoechst/PI双染荧光观察10μmol/L处理组细胞处于晚期凋亡及坏死状态,Annexin V-PI双染流式检测细胞凋亡率随着给药浓度的增加而升高。结论：秋水仙碱在0.1~10μmol/L的浓度范围内具有较强的体外肾毒性,其机制可能是秋水仙碱将细胞阻滞在G2/M期,影响细胞的分裂从而诱导细胞凋亡,对细胞产生毒性。

AIM： To determine whether colchicine causes nephrotoxicity and to investigate the possible toxicity mechanism. METHODS： The rat renal cells NRK were cultured in vitro and MTT assay,cellular morphology observation,lactate dehydrogenase release assay（LDH assay） were used to evaluate the nephrotoxicity that caused by colchicine,the Hoechst 33324/PI double staining and flow cytometry were used to evaluate the possible mechanism of colchicine toxicity.RESULTS：Results showed that colchicine at the concentrations of 0.1-10 μmol/L could inhibit NRK cells viability significantly in a concentration-dependent manner.Besides,the change of cell morphology were observed and the LDH release were significantly increased when cells were treated with 1 μmol/L of colchicine.Moreover,Hoechst 33342/PI double staining and flow cytometry analysis showed that cells were nearly in late apoptosis/necrosis state and they were blocked in G2/M phase after treatment with 10 μmol/L of clochicine.CONCLUSION： Colchicine at concentration of 0.1-10 μmol/L could cause serious nephrotoxicity.The mechanism of colchicine toxicity in NRK cells might be that colchicine blocked cell division in G2/M phase,which induces cells apoptosis,thereby,generates cytotoxicity.