Assembly of epithelial cell fibrillins

Abstract

Fibrillins are large structural macromolecules that are components of connective tissue microfibrils. Fibrillin microfibrils have been found in association with basement membranes, where microfibrils appear to insert directly into the lamina densa. It is unknown whether fibrillins are limited to these sites of microfibril insertion or are present throughout the lamina densa. In this study, electron microscopic immunolocalization demonstrated the presence of fibrillin-1 throughout the lamina densa in the dermal- epidermal junction. In order to investigate whether fibrillin microfibrils might be present in the lamina densa, epithelial cell cultures (WISH, HaCaT, and primary keratinocytes) were analyzed by immunofluorescence, immunoblotting, and extraction of microfibrils followed by rotary shadowing electron microscopy and compared to mesenchymal cell cultures (dermal fibroblasts and MG63 osteosarcoma). In contrast to mesenchymal cells, which elaborate a fibrillin fibril network, epithelial cells primarily deposit fibrillin into the extracellular matrix in a non-fibrillar form. Coculture experiments using human epithelial cells and mouse fibroblasts implicated the cells themselves in the assembly of fibrillin. The importance of the cell in this process was further underscored by novel data demonstrating that keratinocytes selectively secrete fibrillin-1 into the matrix and not into the medium and can differentiate between fibrillin-1 and fibrillin-2.