Abstract
Cyclosporine A (CsA) is known to have direct toxicity to renal tubular cells. Its toxicity may be mediated by intracellular calcium because CsA increases intracellular calcium concentration and enhances the activities of calcium‐dependent calpains and caspases. Calbindin‐D~28k~, a cytosolic calcium binding protein, has been used as an intracellular Ca^2+^ buffer to reduce calcium‐mediated cytotoxicity in non‐renal cells such as neuronal cells. We investigated the effects of gene transfer of calbindin‐D~28k~ cDNA on CsA cytotoxicity and intracellular calcium concentration ([Ca^2+^]~i~) in cultured murine proximal tubular (MCT) cells. A plasmid containing calbindin‐D~28k~ cDNA under the control of CMV promoter was transfected to MCT cells with liposomes. Cytotoxicity was assessed by LDH release and cell viability assay, and [Ca^2+^]~i~ was measured ratiometrically with fura‐2. Compared with MCT cells, cells transfected with calbindin‐D~28k~ cDNA showed a reduction in LDH release by 27, 30, 32, 33, and 19% (all P < 0.05), respectively, after 24 h exposure to 1, 2.5, 5, 10, and 25 μM CsA. Cell viability after CsA treatment was also significantly higher in CB cells. A mock transfection using plasmid without calbindin‐D~28k~ cDNA insert did not affect the LDH release or cell viability after CsA treatment. CsA treatment did not affect the protein and mRNA abundance of transfected calbindin‐D~28k~ cDNA. The expression of calbindin‐D~28k~ did not affect the baseline [Ca^2+^]~i~, but significantly suppressed CsA‐induced elevation in [Ca^2+^]~i~. The expression of calbindin‐D~28k~ in renal tubular cells provides cytoprotective effects against CsA toxicity, probably through its buffering effects on [Ca^2+^]~i~. © 2004 Wiley‐Liss, Inc.