The fetus responds to injury in a fashion fundamentally different from the adult: the process of mesenchymal regeneration proceeds in the absence of inflammation and scarring. We postulate that differences in the extracellular matrix, specifically the early appearance and prolonged maintenance of an environment rich in hyaluronic acid (HA), facilitates cell mobility, cell proliferation, and regeneration in fetal wounds. We propose to characterize the process of healing linear and open wounds in adult sheep and fetal lambs at various gestational ages. The fetal lamb is large enough to compare 1 cm linear incisions, 9 mm open wounds, and adjacent unwounded tissue, and to document the sequential appearance and disappearance of the cellular and extracellular components of repair using the following techniques: a) light microscopy including special stains, b) immunolocalization using fluorescent antibodies to identify cell markers and extracellular matrix components, c) electron microscopy, and d) localization of hyaluronic acid using a newly developed HA-binding protein stain. This model allows us to determine the effect of gestational age by comparing the healing process in adults and fetuses at 60, 100, and 130 days' gestation (term = 145 days). The fetal lamb model can be surgically manipulated to elucidate the effect of the fetal environment on healing: we will compare the process of healing skin and visceral wounds exposed to amniotic fluid to those excluded from amniotic fluid. We will also compare wounds made in adult (maternal) skin allografted onto the fetus at 60 days' gestation (the fetus is tolerant of skin grafts until 70 days) to wounds made on adjacent autografted fetal skin. Recent evidence suggests that prolonged maintenance of HA is central to fetal wound healing. We will compare HA deposition and metabolism in adult wounds and fetal wounds at various gestational ages. After documenting the morphology of HA deposition in our model, we will examine the biochemistry of HA deposition and metabolism by analyzing wound fluid (aspirated from wire mesh cylinders) as well as serum, urine and amniotic fluid using newly developed assays for HA synthesis, hyaluronidase activity, hyaluronidase inhibitors, and newly discovered HA stimulating and inhibiting factors. The devastating clinical problems associated with inflammation and scar formation may be alleviated by modulating the extracellular matrix components to provide a more fetal-like environment for adult tissue repair.
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