Osmotic regulation of aldose reductase protein synthesis in renal medullary cells

Renal medullary cells are normally exposed to high extracellular NaCl as part of the urinary concentrating mechanism. They react to this stress by accumulating sorbitol and other organic osmolytes. PAP-HT25, a line of epithelial cells derived from rabbit renal inner medulla, expresses this response. In hypertonic medium, these cells accumulate large amounts of sorbitol. There is a large increase in the amount of aldose reductase, which catalyzes production of sorbitol from glucose. The purpose of the present study was to investigate whether the aldose reductase protein increases because of faster synthesis or slower degradation. We measured the rate of synthesis and degradation of aldose reductase protein by pulse-chase with [35S]methionine, followed by immunoprecipitation with specific antiserum and autoradiography. The protein synthesis rate was 6 times greater in cells grown in hypertonic (500 mosmol/kg) medium, than in those grown in normal (300 mosmol/kg) medium. When control cells were switched to hypertonic medium, the synthesis rate increased 15-fold by 24 h, then decreased to 11-fold after 48 h. In contrast, synthesis rate continued to increase past 24 h when accumulation of sorbitol was prevented by inhibiting aldose reductase activity with Tolrestat. Thus, there is a feedback mechanism by which cellular sorbitol accumulation inhibits aldose reductase protein synthesis. Degradation of aldose reductase protein was slow (only about 25% in 3 days) and was not affected by osmolality. Thus, the osmoregulatory increase in aldose reductase protein is due to an increase in its synthesis rate and not to any change in its degradation.