The cytoskeleton and substratum adhesion in chick embryonic corneal epithelial cells.

The adhesions and cytoskeleton of 15-day chick embryonic comeal epithelial cells have been studied using interference reflection microscopy (IRM), electron microscopy and fluorescent staining with specific antibodies against actin, tubulin, prekeratin, fibronectin and laminin In vivo, desmosomes were the most frequent intercellular junctions and hemidesmosomes were prominent at the basal surface of the epithelium in contact with the basement membrane. The intact epithelium stained for actin, tubulin, and prekeratin and laminin, but not fibronectin, and the basement membrane for fibronectin and laminin. In monolayer culture intercellular desmosomes were reformed after dissociation with trypsin. However, we have concentrated on cell-substratum adhesions, because the relationship between substratum adhesions and the cytoskeleton has not been thoroughly investigated for epithelial cells. Most of our results apply to contacts with gelatin substrata but the situation was similar on glass or tissue-culture plastic. By IRM, focal contacts similar to those of fibroblasts were present, mainly beneath the leading lamellae of the peripheral cells of monolayered islands. Fluorescent antibody staining revealed that each focal contact was positioned at the end of an actin microfilament bundle. However, there was no correspondence between focal contacts and either prekeratin filaments or microtubules. Fibronectin fibrils were found principally beneath peripheral cells, but evidence is presented suggesting that fibronectin is not directly involved in cell-substratum adhesion of these cells. We suggest that the fibrillar fibronectin pattern arises because cells physically reorganize fibronectin which adsorbs or binds to the substratum from the medium. Electron microscopy suggested that these cells formed two types of contacts with the substratum : small dense plaque-like structures, probably hemidesmosomes, were present in addition to focal contacts. The former were associated with tonofilaments and were the same size (about o-i fim in diameter) as the desmosomes and hemidesmosomes found in vivo. They were much smaller than focal contacts which were from one to several micrometers in length. These structures were not as well formed as the hemidesmosomes found in vivo, possibly because the substratum was not entirely appropriate. The significance of the two types of cell-substratum adhesions is discussed.