Proprietary interest category: N. Reprint requests: Dr. Patrick R. Cammarata, Department of Anatomy and Cell Biology

Abstract

Purpose. Bovine lens epithelial cells (BLECs) accumulate osmotically active organic solutes (i.e., osmolytes) including myo-inositol when exposed to hypertonic stress (osmotic shock). In hypertonic medium, the increase in myo-inositol accumulation is attributed to an elevation in activity of Na + /myo-inositol cotransporter(s). The authors report the nature of the hypertonicity-induced enhancement of myo-inositol uptake in cultured BLECs by amplifying a 626 bp cDNA from lens cell RNA. Methods. A portion of cDNA encoding a Na + /myo-inositol cotransporter was isolated from cultured BLECs using PCR primers designed from an established myo-inositol transporter from Madin-Darby canine kidney (MDCK) cells. Using the reverse transcription-polymerase chain reaction, a 626 bp PCR product was amplified. Its nucleic acid sequence was determined by the dideoxynucleotide method using Sequenase kit. Na + /Myo-inositol cotransporter mRNA expression in the cultured cells was demonstrated under physiological and hypertonic conditions by northern analysis of poly (A) + RNA using the lens cell 626 bp cDNA as probe. Results. The BLEC cDNA sequence was 92% identical with the Na + /myo-inositol cotransporter of MDCK cells. Myo-inositol transporter mRNA was demonstrated in cultured BLECs and was significantly induced by hypertonic stress. Conclusions. These data suggest that cultured bovine lens epithelial cell adaptation to hypertonicity involves intracellular accumulation of small organic osmolytes (i.e., myo-inositol) through elevation of myo-inositol uptake activity resulting from the upregulation of transporter mRNA. Invest Ophthalmol Vis Sci. 1994; 35:1236-1242 .Little is known about the osmoregulatory role of organic osmolytes (including myo-inositol) under normal, hyperglycemic, and hypertonic conditions in lens cells. Many cells, including bovine lens epithelial cells, accumulate small organic osmolytes in adaptation to water stress in hypertonic environments