MYOFIBROBLAST TRANSFORMATION OF CAT CORNEAL ENDOTHELIUM BY TRANSFORMING GROWTH-FACTOR-BETA(1), GROWTH-FACTOR-BETA(2), AND GROWTH-FACTOR-BETA(3)

PURPOSE. Under certain pathophysiologic conditions, the corneal endoth elium can produce an abnormal posterior collagenous layer (PCL) that r educes light transmission. Previous studies suggest that formation of PCLs can result from transformation of endothelial cells to a prolifer ative myofibroblast phenotype. The purpose of this study was to determ ine the potential role of transforming growth factor (TGF)-beta on cor neal endothelial transformation. METHODS. Three corneal buttons (6-mm diameter) were obtained from each cornea of 28 adult cats. After a 2-m m diameter mechanical scrape injury was made, each button was cultured for 24, 48, or 72 hours in serum-free medium (SFM) or SFM supplemente d with 10% fetal calf serum, TGF-beta(1), TGF-beta(2), TGF-beta(3), ba sic fibroblast growth factor (bFGF), or TGF-beta(1) and bFGF. Buttons were single and double labeled using phalloidin and antibodies to ZO-1 , Ki67, fibronectin, cu-smooth muscle (SM) actin, and vinculin. Counts of Ki67-positive cells were used as a measure of endothelial prolifer ation. RESULTS. Organ culture in TGF-beta(1), beta(2), or beta(3) indu ced myofibroblast transformation of corneal endothelial cells, with fo rmation of stress fibers containing alpha-SM actin, loss of normal per icellular ZO-1 organization, development of extracellular fibronectin fibrils, and formation of focal contacts as indicated by punctate vinc ulin staining. However, TGF-beta did not stimulate endothelial prolife ration above that in serum-free control samples. Serum and bFGF each s timulated proliferation significantly, without inducing myofibroblast transformation. A combination of TGF-beta(1) and bFGF resulted in both myofibroblast transformation and increased proliferation. CONCLUSIONS . These results suggest that TGF-beta plays a key role in the loss of normal endothelial differentiation, abnormal extracellular matrix synt hesis, and myofibroblast transformation, which can induce development of PCLs. However, other factors such as bFGF seem to be required to st imulate concomitant proliferation of corneal endothelium.