Abstract
Intracellular calcium plays an important role on the pathogenesis of hypoxia-induced cellular injury. Calbindin-D28k, a cytosolic vitamin D-dependent calcium binding protein, can serve as a buffer to limit a surge in intracellular Ca2+ concentration ([Ca2+]i) induced by various stimulations. To evaluate the possible cytoprotective effect of calbindin-D28k against hypoxic injury in proximal tubular cells, a plasmid containing calbindin-D28k cDNA under the control of CMV immediate-early gene promoter was transfected into the murine proximal tubular epithelial (MCT) cells. The expression of calbindin-D28k in the transfected cells was verified with Northern blot analysis, Western blot analysis, and immunofluorescent staining. The non-transfected and transfected MCT cells were subjected to chemical hypoxia induced by antimycin A (10 μM) and glucose deprivation for 30-120 min. The transfection of calbindin-D28k reduced lactate dehydrogenase (LDH) release by 41%, 41%, 24%, and 24%, respectively, at 30, 60, 90 and 120 min after hypoxia when compared to the non-transfected cells (all p < 0.05). Cell viability after hypoxic injury was also significantly higher in transfected cells than non-transfected cells. Transfection with the plasmid without calbindin-D28k cDNA did not affect LDH release or cell viability after chemical hypoxic injury. [Ca+2]i was measured ratiometrically with fura-2 after exposure to chemical hypoxia. The rate of initial rise in [Ca2+]i and final [Ca+2]i at 30-120 min were significantly lowered in transfected cells. In conclusion, this study demonstrated that transfection of calbindin-D28k gene into MCT cells provide protective effects against chemical hypoxic injury probably through its buffering effects on [Ca+2]i.
Original language | English (US) |
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Pages (from-to) | 559-569 |
Number of pages | 11 |
Journal | Life Sciences |
Volume | 71 |
Issue number | 5 |
DOIs | |
State | Published - Jun 21 2002 |
Keywords
- Intracellular calcium
- Lactate dehydrogenase
- Renal cells
- Transfection
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)
- Pharmacology, Toxicology and Pharmaceutics(all)