Morphology and cellular origins of substrate-attached material from mouse fibroblasts

Abstract

Scanning electron microscopy (SEM) has been used to determine the pattern of reorganization of substrate-bound cell surface material during attachment of Swiss 3T3 mouse fibroblasts to fresh serum-coated glass substrates after prior detachment with the Ca 2+-specific chelator EGTA (ethylene- bis-(oxyethylenenitrilo)-tetraacetic acid). A limited number of filopodia make contact with the substrate surface and branch into finger-like extensions. A thin web-like membrane organizes between these filopodial extensions to eventually form mature footpads which appear to have a limited size. Footpads appear to be the principal morphological entities mediating cell substrate adhesion. The mechanism of EGTA-mediated detachment of cells and the morphology of the substrate attached material (SAM) left adherent to the substrate after complete removal of cells have also been examined. EGTA treatment results in cell Founding, with minimal effects on the morphology of the adhesive footpads which remain linked to the cell body via stressed retraction fibers. These stressed fibers eventually break, leaving footpads on the substrate as SAM and liberating the cell body into suspension. This evidence suggests that EGTA treatment minimally affects adhesive materials in the footpad site but disorganizes the cytoskeleton of the cell with resultant shearing of the cell body away from the footpads. The size, morphology, and pattern of SAM left on the substrate establish that it is essentially footpad material. SAM from highly-spread cells appears to be virtually-intact footpads, while SAM from attaching cells appears to be disrupted footpad material. These results are discussed in terms of the morphological and biochemical organization of the footpad adhesion sites of cells.