Cells That Emerge from Embryonic Fibers of Type IV Collagen Explants Produce

Double immunofluorescence staining experiments designed to examine the synthesis and deposition of collagen types I and IV in cultured explants of embryonic mouse lung revealed the presence of connective tissue-like fibers that were immunoreactive with antitype IV collagen antibodies. This observation is contrary to the widely accepted belief that type IV collagen is found only in sheetlike arrangements beneath epithelia or as a sheathlike layer enveloping bundles of nerve or muscle cells. The extracellular matrix produced by cells that migrate from embryonic mouse lung rudiments in vitro was examined by double indirect immunofluorescence microscopy. Affinity-purified monospecific polyclonal antibodies were used to examine cells after growth on glass or native collagen substrata. The data show that embryonic mesenchymal cells can produce organized fibers of type IV collagen that are not contained within a basement membrane, and that embryonic epithelial cells deposit fibers and strands of type IV collagen beneath their basal surface when grown on glass; however, when grown on a rat tail collagen substratum the epithelial cells produce a fine meshwork. To our knowledge this work represents the first report that type IV collagen can be organized by cells into a fibrous extracellular matrix that is not a basement membrane. The basement membrane is a histological structure found subjacent to all epithelia and surrounding bundles of muscle fibers and nerve cells (Vracko, 1982). A hallmark of basement membranes is that they separate epithelia, muscle, or nerve from the adjacent connective tissue or mesenchyme. Type IV collagen is a major structural component of basement membranes (Timpl et al., 1982). The other general class of extracellular matrix is associated with mesenchymal connective tissue, which contains predominately collagen types I and III, although the latter is usually tess abundant (Eyre, 1980). These two collagen types form the scaffold of fibrous connective tissue found in bone, teeth, dermis, tendon, ligament, and ocular tissue. In addition, collagen types I and III form the fibrous stroma of all visceral organs, blood vessels, peripheral nerves, skeletal muscles, and the fibrous skeleton of the heart (Ham and Cormack, 1979; Eyre, 1980; Bornstein and Sage, 1980). These two general kinds of extracellular matrices are usually considered immutable. In the present study cells that emerge from embryonic mouse lung explants were examined by immunofluorescence microscopy after detergent permeabilization. The predominant fibrous collagen we observed under these conditions was immunoreactive type IV. THE JOURNAL OF CELL BIOLOGY . VOLUME 101 OCTOBER 1985 1175-I 181 © The Rockefeller University Press 0021-9525/85/1011175/07 $ 1.00 Examination of mesenchymal cell populations showed aligned fibers of type IV collagen. In fact, the heaviest concentrations of type IV collagen fibers were always observed in fields of axially aligned spindle shaped cells. The presence of aligned type IV collagen fibers in regions of mesenchymal outgrowth is consistent with the studies of Harris and his colleagues, who examined the interaction of fibroblasts with type I collagen fibers. The term tractional structuring was used by these workers to describe the process by which the cells aligned the collagen (Harris et al., 1981; Stopack and Harris, 1982). Our data suggest that tractional structuring of type IV collagen occurs as cells emerge from embryonic explants. The data also show that emergent embryonic epithelial cells produce type IV collagen fibers with a very wide variety of sizes beneath their basal surface when cultured on glass. Contrary to this, epithelial cells resting upon native rat tail collagen (RTC) ~ substrata produce a uniform coherent meshwork of type IV collagen that is probably analogous to its organization within basement membranes. To our knowledge the present work is the first report that JAbbreviations used in this paper: DIC, differential interference contrast; RTC, rat tail collagen. 1175 on A ril 0, 2017 D ow nladed fom Published October 1, 1985