Tao W, Liou G I, Wu X, Abney T O, Reinach P S
Division of Neoplastic Diseases, Mount Sinai Medical Center, New York, NY 10029, USA.
Invest Ophthalmol Vis Sci. 1995 Dec;36(13):2614-22.
To determine if there is a heterogeneous pattern of endothelin (ET) receptor subtype (i.e., ETA and ETB) gene expression in the bovine corneal epithelium (BCE). To determine if ET receptor subtype stimulation increases the effectiveness of epidermal growth factor (EGF) to accelerate wound closure in a primary culture of bovine corneal epithelial cells (BCEC).
In situ hybridization histochemistry was used to characterize ETA and ETB gene expression in the BCE. A wound closure assay evaluated wound healing rates in BCEC after 4 to 7 days in culture. [3H] thymidine incorporation and MTT assay measured proliferation.
ETA gene expression was appreciably higher in the basal cells than in the suprabasal cells, whereas the pattern for ETB was reversed. Epidermal growth factor (5 ng/ml) maximally increased wound closure by 145% above the control. With 5 ng/ml EGF, either 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 ng/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 through a selective increase in proliferation. Neither ET-1 nor s-6-c alone had any effect on proliferation or migration.
Both ETA and ETB genes are expressed in BCE. However, in BCEC only, ETB stimulation increases the effectiveness of EGF to stimulate wound closure. This response was caused by an increase in cell migration rather than proliferation because, after treatment with mitomycin C, neither ET-1 nor EGF stimulated wound closure.
确定牛角膜上皮（BCE）中内皮素（ET）受体亚型（即ETA和ETB）基因表达是否存在异质性模式。确定ET受体亚型刺激是否能提高表皮生长因子（EGF）在牛角膜上皮细胞（BCEC）原代培养中加速伤口闭合的效果。
采用原位杂交组织化学法对BCE中ETA和ETB基因表达进行表征。伤口闭合试验评估培养4至7天后BCEC的伤口愈合率。[3H]胸苷掺入法和MTT试验检测细胞增殖。
ETA基因在基底细胞中的表达明显高于基底上层细胞，而ETB的表达模式则相反。表皮生长因子（5 ng/ml）使伤口闭合最大程度地比对照提高了145%。在5 ng/ml EGF存在的情况下，10(-9) M ET-1或10(-8) M 6-羧基-sarafotoxin（s-6-c）使伤口闭合比单独使用5 ng/ml EGF时额外提高了39%（P < 0.001）。BQ123（10(-7) M）未改变ET-1或s-6-c的任何这些作用。表皮生长因子通过选择性增加细胞增殖来刺激伤口闭合。ET-1和s-6-c单独对细胞增殖或迁移均无任何影响。
ETA和ETB基因均在BCE中表达。然而，仅在BCEC中，ETB刺激可提高EGF刺激伤口闭合的效果。这种反应是由细胞迁移增加而非增殖增加引起的，因为在用丝裂霉素C处理后，ET-1和EGF均未刺激伤口闭合。
