Role of fibronectin deposition in branching morphogenesis of Madin-Darby canine kidney cells

Abstract

Background. Madin-Darby canine kidney (MDCK) epithelial cells grown in collagen gels in the presence of hepatocyte growth factor (HGF) form branching tubules. The tubule-lining epithelial cells are polarized with the basolateral surface in contact with the collagen gel and the apical surface facing the lumen. To delineate whether MDCK branching tubules construct the basal lamina, we characterized the composition of the extracellular matrix deposited by MDCK tubules. The tubule-lining cells produced an apparently incomplete basal lamina containing a discontinuous laminin substratum. In addition, a thick layer of fibronectin surrounded the basal cell surface of the branching tubule. In an attempt to delineate the role of fibronectin deposition in branching morphogenesis, we conducted this study. Methods. MDCK cells cultured in collagen gel were employed. We first used arginine-glycine- aspartate peptides containing disintegrin rhodostomin to disturb the interactions between fibronectin and cell surface integrins. Furthermore, we established several stable transfectants expressing fibronectin antisense RNA to examine the role of fibronectin in branching morphogenesis directly. Results. Rhodostomin inhibited the formation of branching tubules. The transfectants expressing fibronectin antisense RNA exhibited relatively lower levels of synthesized fibronectin and markedly diminished growth rates of branching tubules than the control clone. An inhibition of branching morphogenesis induced by the overexpression of fibronectin antisense RNA was manifested by the decrease in cell growth rates and cell migration. Conclusion. These results indicate that the deposition of fibronectin underlying the tubule-lining epithelium serves to enhance cell proliferation and migration, and hence facilitates the branching tubulogenesis of MDCK cells.