Under normal growth conditions, in vitro dermal microvascular endothelial cells (HDMEC) retain an epithelioid morphology and do not synthesize matrix proteins found increased in scar tissue. When injured by a standard scratch, cells at the wound edge and within the culture transform into spindle-shaped, myofibroblast-like cells. To determine if the transformed cells synthesize matrix proteins, expression of type I collagen and alpha smooth muscle actin (α-SMA) was investigated by immunohistochemistry and quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). Twelve hours following injury, a major upregulation in expression of α-SMA and type I collagen was observed both in cells proximal and distal to the wound edge. Cells with the typical morphology of myofibroblasts and displaying intracellular α-SMA positive fibrils were observed in HDMEC throughout the culture. In contrast, type IV collagen, a basement membrane protein, was not detected in migrating cells. Following completion of wound repair (24–36 h), type I collagen was no longer expressed and type IV collagen synthesis increased to prewound levels. Quantitative RT-PCR confirmed the changes in gene expression for both type I collagen and α-SMA at each time point during repair. These results demonstrate that normal skin microvascular endothelial cells retain an ability to transform into myofibroblast-like cells when injured and to synthesize matrix proteins not expressed in noninjured cells. The synthesis of matrix proteins by injured endothelial cells suggests a direct role for the endothelium in the pathology of scar formation.