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J.E. Jumblatt, L.B. Lanceta, T. Jiang, M.M. Jumblatt; Role of Calcium in 15(S)-HETE-Mediated Secretion of MUC1 Mucin by Corneal Epithelial Cells . Invest. Ophthalmol. Vis. Sci. 2003;44(13):1338.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: MUC1 mucin is prominently expressed at the ocular surface, both as a transmembrane protein on conjunctival and corneal epithelial cells and as a secreted glycoprotein in aqueous tears. The eicosanoid 15(S)-HETE has been shown to stimulate MUC1 secretion from human conjunctiva and to thicken the mucin layer that coats the rabbit corneal surface, although the underlying biochemical mechanisms are unknown. The present study was undertaken to evaluate the role of Ca(2+)in 15(S)-HETE-induced MUC1 secretion by CEPI cells, an SV40 virus-immortalized human corneal epithelial cell line. Methods: Expression of MUC1 mRNA and protein by CEPI cells was demonstrated by RT-PCR and Western blot analysis, respectively. Secretion of MUC1 was measured by dot blot immunoassay using DF3 anti-MUC1 antibody and purified Ca15-3 antigen as the standard. In general, cells were pre-incubated with pharmacological agents for 30 min followed by a 60 min incubation in medium ± 15(S)-HETE. Results: 15(S)-HETE (1 µM) induced a >2-fold increase in MUC1 release relative to vehicle controls. The response to 15(S)-HETE was mimicked by the Ca(2+) ionophore ionomycin (10 µM) and absent in Ca(2+)-free medium supplemented with EGTA (1 mM). In medium containing 1.5 mM Ca(2+), 15(S)-HETE-induced MUC1 release was blocked by the intracellular Ca(2+) chelator BAPTA/AM (100 µM) but was unaffected by the Ca(2+) channel antagonist verapamil (10 µM) or depletion of intracellular Ca(2+) stores by the Ca(2+) pump inhibitor cyclopiazonic acid (15 µM). Conclusion: The results suggest that 15(S)-HETE-induced MUC1 release from CEPI cells is a Ca(2+)-dependent process, possibly involving the influx of Ca(2+) via store-operated Ca(2+) channels similar to those described in epidermal keratinocytes. Whether the cellular mechanism of MUC1 release involves Ca(2+)-mediated exocytosis or proteolytic shedding remains to be determined. Support: NEI Grant EY10736, Alcon Research, Ltd., Ky. Lions Eye Foundation, RPB.
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