Influence of fetal calf serum, fibroblast growth factors, and hepatocyte growth factor on three-dimensional cultures of human keratocytes in collagengel matrix

Background: We set out to evaluate the influence of fetal calf serum, acidi c fibroblast growth factor (aFGF), basic fibroblast growth factor (bFGF), a nd hepatocyte growth factor (HGF) on three-dimensional cultures of human ke ratocytes in type I collagen gel matrix. Methods: Polymerized gels were cul tured at 37 degrees C for 35 days. Gel contraction and integrated optical d ensity were assessed 3 times weekly for 5 weeks using an image analysis sys tem. Gels were studied at the end of the culture period by means of transmi ssion electron microscopy (TEM) and immunochemistry. Results: Serum signifi cantly increased gel contraction and decreased gel optical transmittance. K eratocyte density was significantly increased by serum and HGF. In TEM, col lagen density was higher with serum-supplemented media than with serum-free media, and higher with HGF-supplemented media than with HGF-free media. Im munoperoxidase staining of keratocyte-populated gels showed positive staini ng for vimentin, connexin 43, and type I, type V, and type VI human collage n, whereas no expression of desmin, alpha smooth muscle actin, and type IV collagen was observed. Expression of type I collagen was significantly incr eased by aFGF and HGF, expression of type VI collagen by serum and bFGF. Co nclusion: Serum and HGF improve ultrastructural and immunochemical features of human keratocyte-populated collagen gels.