In vivo co-distribution of fibronectin and actin fibers in granulation tissue: immunofluorescence and electron microscope studies of the fibronexus at the myofibroblast surface.

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Abstract
The fibronexus (FNX), a very close transmembrane association of individual extracellular fibronectin fibers and actin microfilaments, was found previously at the substrate-binding surface of fibroblasts in tissue culture. To determine whether the fibronexus might be involved in fibroblast adhesion during wound healing in vivo, colocalization of actin and fibronectin in granulation tissue formed within full-thickness guinea pig skin wounds was sought. At 7-9 days, most of the actin fibers were observed to be coincident with congruent fibronectin fibers using double-label immunofluorescence microscopy. These fibronectin and actin fibers were colocalized at the myofibroblast surface surrounding the nucleus, and along attenuated myofibroblast processes which extended deeply into the extracellular matrix. This conspicuous codistribution of fibronectin and actin fibers prompted a search for fibronexuses at the myofibroblast surface with EM. Three kinds of FNX were observed as follows: tandem associations between the termini of individual extracellular fibronectin fibers and actin microfilament bundles at the tips of elongate myofibroblast processes, plaque-like and, track-like FNX, in which parallel fibronectin and actin fibers were connected by peripendicular transmembranous fibrils. Goniometric studies on the external and internal components of these cross-linking fibrils showed that their membrane-associated ends are probably co-axial. Using immunoelectron microscopy on ultrathin cryosections, the densely staining external portion of these various FNX does indeed contain fibronectin. The finding that these FNX appear to connect collagen fibers to intracellular bundles of actin microfilaments is particularly significant. Evidently, the fibronexus is an important in vivo cell surface adhesion site functioning in wound repair, and perhaps within fibronectin-rich tissues during embryogenesis, tumor growth and inflammation.