Effects of cadmium chloride on mitochondrial functions of adrenocortical cells

Abstract

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. Key words: Cadmium chloride; Adrenocortical cells; Flow cytometry; Mitochondrial membrane potential; Cytotoxicity