OXALATE TOXICITY IN CULTURED MOUSE INNER MEDULLARY COLLECTING DUCT CELLS

Abstract

Purpose: Oxalate, a metabolic end product and a major constituent of the majority of renal stones, has been shown to be toxic to renal epithelial cells of cortical origin. However, to our knowledge it is unknown whether inner medullary collecting duct (IMCD) cells, which are physiologically exposed to higher concentrations of oxalate, also behave in a similar manner. Materials and Methods: A line of IMCD cells was exposed to oxalate (0.2 to 10 mM) for various time points. Trypan blue, and hematoxylin and eosin stains were used to assess cell morphology and membrane integrity. The production of reactive oxidative species was determined using the nitro blue tetrazolium reaction and crystal violet staining was used to measure cell density. Results: Exposure of IMCD cells to oxalate produced time and concentration dependent changes in the light microscopic appearance of the cells. Long-term exposure to oxalate resulted in alterations in cell viability with net cell loss following exposure to concentrations of 2 mM and greater. Free radical production was time and concentration dependent. Crystal formation occurred in less than 1 hour and cells in proximity to crystals lost membrane integrity. Compared to IMCD cells LLC-PK1 and HK2 cells showed significant toxicity starting at lower oxalate concentrations (0.4 mM and above). Conclusions: To our knowledge the results provide the first direct demonstration of toxic effects of oxalate in IMCD cells, a line of renal epithelial cells of the inner medullary collecting duct, and suggest that cells lining the collecting duct are relatively resistant to oxalate toxicity.