ET(B) AND EPIDERMAL GROWTH-FACTOR RECEPTOR STIMULATION OF WOUND CLOSURE IN BOVINE CORNEAL EPITHELIAL-CELLS

Purpose, To determine if there is a heterogeneous pattern of endotheli n (ET) receptor subtype (i.e., ET(A) and ET(B)) gene expression in the bovine corneal epithelium (BCE). To determine if ET receptor subtype stimulation increases the effectiveness of epidermal growth factor (EG F) to accelerate wound closure in a primary culture of bovine corneal epithelial cells (BCEC). Methods, In situ hybridization histochemistry was used to characterize ET(A) and ET(B) gene expression in the BCE. A wound closure assay evaluated wound healing rates in BCEC after 4 to 7 days in culture. [H-3] thymidine incorporation and MTT assay measur ed proliferation. Results, ET(A) gene expression was appreciably highe r in the basal cells than in the suprabasal cells, whereas the pattern for ET(B) was reversed. Epidermal growth factor (5 ng/ml) maximally i ncreased wound closure by 145% above the control. With 5 ng/ml EGF, ei ther 10(-9) M ET-1 or 10(-8) M sarafotoxin-6-c (s-6-c) increased wound closure by an additional 39% (P < 0.001) above that measured with 5 n g/ml EGF alone. BQ123 (10(-7) M) did not alter any of these effects of ET-1 or s-6-c. Epidermal growth factor stimulated wound closure throu gh a selective increase in proliferation. Neither ET-1 nor s-6-c alone had any effect on proliferation or migration. Conclusions. Both ET(A) and ET(B) genes are expressed in BCE. However, in BCEC only, ET(B) st imulation increases the effectiveness of EGF to stimulate wound closur e. This response was caused by an increase in cell migration rather th an proliferation because, after treatment with mitomycin C, neither ET -1 nor EGF stimulated wound closure.