Variation in sorbitol accumulation and polyol-pathway activity in cultured human proximal tubule cells.

The polyol pathway is present in tissues of several organs where its activation may participate in the development of diabetic complications. We measured the accumulation of polyol-pathway intermediates in HPT cells isolated from 21 different human kidneys from nondiabetic individuals. When exposed to 27.5 mM glucose in the growth media, cells isolated from approximately 75% of individuals (accumulators) accumulated sorbitol within 1-4 days, whereas 25% (nonaccumulators) accumulated only negligible amounts, even when the period of exposure was extended to 2 wk. Surprisingly, measurement of the activities of the polyol-pathway enzymes showed no difference in the levels of either AR or SDH between accumulators and nonaccumulators, even when the conversion of galactose to galactitol was used to measure AR activity in intact cells independently of SDH. Measurement of sorbitol in the growth media indicated that nonaccumulators were not releasing sorbitol into the growth media. Fructose levels in the conditioned growth media were 4 times higher in the sorbitol-accumulating cells. Together, these results indicate that the tendency of cells from an individual to accumulate significant amounts of sorbitol may reflect the cells' ability to metabolize sorbitol in steps subsequent to the polyol pathway.

An electrophysiological freeze fracture assessment of cadmium nephrotoxicity in vitro.

Human proximal tubule cell cultures exposed to doses of cadmium chloride (CdCl2) between 0.05 microgram/ml and 0.5 microgram/ml exhibited alterations in cell membrane structure and transport function. At these Cd concentrations, cell numbers were not significantly altered from control values in either nonreplicating confluent, or actively replicating subconfluent cultures. Transmission electron microscopy revealed few alterations in cultures treated with 0.05 microgram/ml Cd. Tight junctions were intact; organelles and myeloid body formation appeared normal. Freeze fracture analysis confirmed the integrity of the tight junctions as well as increased numbers of vesicles or pits along the lateral cell membrane, indicating increased endocytotic activity. Cells exposed to 0.1 microgram/ml Cd were characterized by decreased numbers of microvilli and inhibited myeloid body formation. Cd doses of 0.5 micrograms/ml elicited nuclear chromatin condensation, fragmented sealing strands in 5 to 10% of the tight junction profiles, sparse microvilli, and inhibited myeloid body formation. Electrophysiologic assessments of transport function by Ussing chamber analysis revealed decreases in transepithelial potentials for all three concentrations, with significant differences at Cd concentrations of 0.5 to 0.1 micrograms/ml. Cells treated with 0.5 micrograms/ml Cd also exhibited slight decreases in electrical resistance, consistent with the minimal fragmentation of sealing strands observed in freeze fracture replicas. Resistance in cultures treated with 0.1 or 0.05 micrograms/ml Cd remained within control values and indicated that drops in potential difference and short circuit current in these cells reflected true alterations in ion transport.