Fibronectins (FN) comprise a family of adhesive glycoproteins that are prominent components of wound healing. These proteins arise by alternative splicing of a single gene transcript at three sites, termed EIIIA, EIIIB, and V. Extravasated plasma FN, which lacks the EIIIA and EIIIB domains, along with fibrin, comprise the" provisional" matrix that forms within minutes of tissue injury. By 2 days after cutaneous excisional wounding in rats, total FN messenger RNA (mRNA) expression is increased locally and dramatically within the surrounding dermis, in the subjacent muscle (panniculus carnosus) and, notably, at the wound margins. Moreover, in contrast to normal skin, 2-day wounds express EIIIA-and EIIIB-containing" embryonic" FN mRNAs. To identify the cells responsible for synthesizing the various FN isoforms, we performed in situ hybridization with probes for the various FN mRNAs. Collagen and lysozyme probes were employed to distinguish fibroblasts from macrophages. At early intervals (2 days) after wounding, macrophages were the principal cells that expressed FN mRNA. Moreover, many of these cells expressed embryonic FN mRNAs. At 7 to 10 days, when the wound defect was maturing, fibroblasts were the major cells synthesizing these embryonic FNs. It is widely accepted that wound macrophages phagocytose debris and provide degradative enzymes and cytokines essential for early stages of tissue repair. Our findings suggest an additional function for wound macrophages--synthesis of embryonic FNs providing an extracellular matrix that facilitates wound repair, perhaps by promoting cell migration.