Adaptation by corneal epithelial cells to chronic hypertonic stress depends on upregulation of Na : K : 2Cl cotransporter gene and protein expressionand ion transport activity

We examined the ability of SV40-immortalized human and rabbit corneal epith elial cells (HCEC and RCEC, respectively) to adapt to chronic hypertonic st ress. Under isotonic conditions, in the presence of 50 mu M bumetanide, pro liferation measured as H-3-thymidine incorporation declined in RCEC and HCE C by 8 and 35%, respectively. After 48 hr exposure to 375 mOsm medium, RCEC proliferation fell by 19% whereas in HCEC it declined by 45%. Light scatte ring behavior demonstrated that both cell lines mediate nearly complete reg ulatory volume increase (RVI) responses to an acute hypertonic (375 mOsm) c hallenge, which in part depend on bumetanide-sensitive Na-K-2Cl cotransport er (NKCC) activity. Following exposing RCEC for 48 hr to 375 mOsm medium, t heir RVI response to an acute hypertonic challenge was inhibited by 17%. Ho wever, in HCEC this response declined by 68%. During exposure to 375 mOsm m edium for up to 24 hr, only RCEC upregulated NKCC gene and protein expressi on as well as bumetanide-sensitive Rb-86 influx, These increases are consis tent with the smaller declines in RVI and proliferation capacity occurring during this period in RCEC than in HCEC. Therefore, adaptation by RCEC to c hronic hypertonic stress is dependent on stimulation of NKCC gene and prote in expression and functional activity. On the other hand, under isotonic co nditions, HCEC RVI and proliferation are more dependent on NKCC activity th an they ale in RCEC.