Mitochondrial superoxide production during oxalate-mediated oxidative stress in renal epithelial cells.

Crystals of calcium oxalate monohydrate (COM) in the renal tubule form the basis of most kidney stones. Tubular dysfunction resulting from COM-cell interactions occurs by mechanism(s) that are incompletely understood. We examined the production of reactive oxygen intermediates (ROI) by proximal (LLC-PK1) and distal (MDCK) tubular epithelial cells after treatment with COM (25-250 microg/ml) to determine whether ROI, specifically superoxide (O(2)(*-)), production was activated, and whether it was sufficient to induce oxidative stress. Employing inhibitors of cytosolic and mitochondrial systems, the source of ROI production was investigated. In addition, intracellular glutathione (total and oxidized), energy status (ATP), and NADH were measured. COM treatment for 1-24 h increased O(2)(*-) production 3-6-fold as measured by both lucigenin chemiluminescence in permeabilized cells and dihydrorhodamine fluorescence in intact cells. Using selective inhibitors we found no evidence of cytosolic production. The use of mitochondrial probes, substrates, and inhibitors indicated that increased O(2)(*-) production originated from mitochondria. Treatment with COM decreased glutathione (total and redox state), indicating a sustained oxidative insult. An increase in NADH in COM-treated cells suggested this cofactor could be responsible for elevating O(2)(*-) generation. In conclusion, COM increased mitochondrial O(2)(*-) production by epithelial cells, with a subsequent depletion of antioxidant status. These changes may contribute to the reported cellular transformations during the development of renal calculi.

Simultaneous determination of oxalic and citric acids in urine by high-performance liquid chromatography.

A simple method for the simultaneous determination of oxalic and citric acids had been developed using reversed-phase HPLC. An aqueous solution containing potassium dihydrogen phosphate (0.25%) and tetrabutylammonium hydrogen sulphate (2.5 mmol) at pH 2.0 was used as mobile phase. Under these conditions both the components were well resolved and detected at 210 nm. The recovery for oxalic and citric acids was 97% and 102%, respectively. The method presented here was applied to urine specimens of a large number of urolithic patients and control subjects. Because of the simplicity of the method its application provides better means of monitoring the concentration of oxalic and citric acids in the formation of renal calculi.

Is hypocitraturia associated with phosphaturia--a potential cause of calcium urolithiasis in first-time stone formers.

The serum and 24 hour urinary excretion levels of various lithogenic and inhibitory substances were assessed in 24 male patients with calcium stone and no previous history of urolithiasis and in 19 age-matched controls. Two groups did not differ significantly (P < 0.01) except in the excretions of sodium, citric acid (being higher in normals) and inorganic phosphate (being higher in patients). Fifty percent patients had hyperphosphaturia, 29.2% hypocitraturia, 20.8% hyperoxaluria and 16.7% hypercalciuria. The present data suggests that hypocitraturia in association with phosphaturia might be one of the main risk factors responsible for calcium urolithiasis in this area.