Effect of calcium channel antagonist diltiazem and calcium ionophore A23187 on cyclosporine A-induced apoptosis of renal tubular cells.
Calcium channel antagonists have been reported to have a favorable impact on cyclosporin A (CsA)-treated kidney transplant recipients. However, it is not clear whether this is because of their direct effect on antagonizing the toxicity of CsA to renal tubular cells. In this study, we have used Madin-Darby canine kidney tubular cells as a model to examine the effect of diltiazem, a calcium channel antagonist, on CsA-induced apoptosis. Moreover, to investigate the possible regulation of CsA cytotoxicity by intracellular calcium level, the effect of the calcium ionophore A23187 on CsA-induced apoptosis was also examined. We found that treatment of CsA (20 microM) alone caused 20-30% cell death, which was apparently (30-40%) enhanced by diltiazem at 100 microg/ml, accompanied by more severe DNA fragmentation, activation of caspases, and a decreased level of Bcl-2. The caspase inhibitor ZVAD-fmk or Bcl-2 overexpression was capable of suppressing apoptosis induced by the synergistic effect of diltiazem and CsA. Moreover, the survival rate of cells treated with CsA (30 microM) alone remained only 30%, however, it was markedly (approximately 40%) elevated by co-treatment with A23187 (75 ng/ml). The rescue of cells from CsA-induced apoptosis by A23187 was correlated with AKT activation, BAD phosphorylation, and caspase-3 inactivation. Taken together, our results suggest that the reported favorable impact of diltiazem on kidney grafts is likely not because of its direct protection on renal tubular cells. Instead, it enhances the toxicity of CsA to renal tubular cells. In addition, our findings raise a possibility that the intracellular calcium level and the AKT pathway may participate in the regulation of CsA cytotoxicity.
Cheng
Chi-Hung
CH
Section of Nephrology, Taichung Veterans General Hospital, Taichung, Taiwan. hcchen@nchu.edu.tw
Hsieh
Chin-Ling
CL
Shu
Kuo-Hsiung
KH
Chen
Yen-Ling
YL
Chen
Hong-Chen
HC
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D000001:Calcimycin / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D002478:Cells, Cultured; D016572:Cyclosporine / Q000633:toxicity; D053938:DNA Fragmentation / Q000187:drug effects; D004268:DNA-Binding Proteins / Q000378:metabolism; D004110:Diltiazem / Q000494:pharmacology*; D004285:Dogs; D004789:Enzyme Activation / Q000187:drug effects; D015870:Gene Expression; D019254:Genes, bcl-2; D007476:Ionophores / Q000494:pharmacology*; D007684:Kidney Tubules / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D014157:Transcription Factors / Q000378:metabolism
D002121:Calcium Channel Blockers; D004268:DNA-Binding Proteins; D007476:Ionophores; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D014157:Transcription Factors; D000001:Calcimycin; D016572:Cyclosporine; D053148:Caspase 3; D020169:Caspases; D004110:Diltiazem; D002118:Calcium