镉对肾上腺皮质细胞线粒体功能的影响

目的 研究镉对肾上腺皮质细胞线粒体功能的影响，为探讨其内分泌毒作用机制提供依据。方法 原代分离培养豚鼠肾上腺皮质细胞，用氯化镉（ＣｄＣｌ２）０、６．２５、１２．５０、２５．００、５０．００、１００．００、２００．００ｕｍｏｌ／ｌ处理细胞１ｈ，以５０ｕｍｏｌ／Ｌ ＣｄＣｌ２处理细胞０、１５、３０、６０、１２０、２４０ｍｉｎ，观察ＣｄＣｌ２对肾上腺皮质细胞线粒体功能毒作用的剂量－效应及时间－效应关系。应用四甲基偶氮唑蓝（ＭＴＴ）试验检测线粒体酶活力，用流式细胞仪结合罗丹明１２３（Ｒｈ１２３）和碘化丙啶（ＰＩ）双标记法检测线粒体膜电位（ＭＭＰ）和细胞存活状态。结果 ＣｄＣｌ２引起细胞线粒体酶活力及ＭＭＰ降低：以６．２５～２００．００ｕｍｏｌ／Ｌ ＣｄＣｌ２染毒细胞１ｈ，对照组ＭＴＴ的平均吸光度（Ａ５７０ｎｍ）值为０．

Effects of cadmium chloride on mitochondrial functions of adrenocortical cells

Objective To sutdy the effects of cadmium chloride on mitochondrial functions of adrenocortical cells so as to explore the possible mechanism of its cytotoxicity. Methods Adrenocortical cells were dispersed and primarily cultured from adult male guinea pigs. The cells were then treated with cadmium chloride(CdC12) of 0,6.25,12.50,25.00,50.00,100.00,200.00 μmol/L for 1 h in dose-effect studying and treated with 50 μmol/L CdCl2 for 0,15,30,60,120,240 minutes in time-course studying. The mitochondrial enzyme activity was evaluated with MTT colorimetric assay.The mitochondrial membrane potential (MMP,△ ψm) and cell viability were assessed in flow cytometry(FCM) by dual-labelling with rhodamine 123 (Rh123) and propidium iodide(PI). Results Cd-Cl2 could induce MTT reduction and △ψm decreasing with the early remarkable changes from the lowest dosage(6.25μmol/L) and the shortest period ( 15 min) ( P < 0.05 ). The average absorption( A ) of MTT at 1 h and 6.25～200.00 μnol/L CdCl2 exposure vs that of control were 0. 138～0.067 vs 0. 164 .The average fluorescent intensity of Rh123 were 2.44～ 1.02 vs 2.63.Both MTT value and △ψm were decreased in different period(15～240main) of 50 μmol/L CdCl2 exposure. The former was 0.093～0.049 vs 0.180, MPP was 1.37～0.93 vs 1.85. IC50 of mitochondrial enzyme activity inhibited by CdCl2 was 61.29 μmol/L. However, the percentages of dead cells were obviously increased after treated with 50μmol/L CdCl2 for 60 min. It could be seen that MTT reduction and △ψm decreasing happened earlier than the increasing of dead cell percentage. All those indices were changed in dose and time-dependent manner(dosc-efect studying: rMTT = - 0. 816, P < 0.01; rMMp = - 0. 775, P < 0.01; r dead cells =0.967, P < 0.01; time-course: rMTT = - 0. 706, P < 0.01; rMMP = - 0. 406, P < 0.05; r dead cells = 0.980, P <0.01 ). Conclusion CdCl2 may induce damage of adrenocorortical cells. The damage of mitochondrial functions may be a critical early event in adrenocortical cytotoxicity of cadmium.