Calcitriol Directly Sensitizes Renal Tubular Cells to ATP-Depletion- and Iron-Mediated Attack

摘要

Vitamin Ds have been reported to have diverse effects on cell homeostasis, leading to suggestions that they have therapeutic applications extending beyond their traditional actions on the Ca2+/parathyroid/bone axis. As some of these potential indications carry an inherent risk of acute renal failure (ARF; eg, cancer chemotherapy and organ transplantation), the goal of this study was to assess whether vitamin Ds directly affect renal tubule injury responses. Cultured human proximal tubular (HK-2) cells were exposed to physiological or pharmacological doses of either calcitriol (D₃) or a synthetic vitamin D₂ analogue (19-nor) for 3 to 48 hours. Their impact on cell integrity (percent lactate dehydrogenase (LDH) release and tetrazolium dye MTT uptake) under basal conditions and during superimposed injuries (ATP depletion/Ca2+ ionophore or iron-mediated oxidant stress) were determined. As vitamin Ds can be anti-proliferative, cell outgrowth ([3H]thymidine uptake and crystal violet staining) was also tested. Finally, the action of D₃ on in vivo ARF (glycerol-induced myoglobinuria) and isolated proximal tubule injury responses were assessed. D₃ induced a rapid, dose-dependent increase in HK-2 susceptibility to both ATP-depletion/Ca2+-ionophore- and Fe-mediated attack without independently affecting cell integrity or proliferative responses. In contrast, D₂ negatively affected only Fe toxicity and only after relatively prolonged exposure (48 hours). D₃ dramatically potentiated in vivo ARF (two- to threefold increase in azotemia), suggesting potential in vivo relevance of the above HK-2 cell results. Proximal tubules, isolated from these glycerol-exposed mice, suggested that D₃ can worsen tubule injury despite a parodoxic suppression of H₂O₂ production. In contrast, D₃ had a mild negative impact on cellular energetics (depressed ATP/ADP ratios), and it accentuated plasma membrane phospholipid breakdown. The latter was observed in both glycerol-treated and control tubules, suggesting a primary role in the injury- potentiation effect of D₃. Vitamins D(s) may directly, and differentially, increase proximal tubule cell susceptibility to superimposed attack. This property should be considered as new uses for these agents are defined.