Fibrosis results when myofibroblasts invade the wound fibrin provisional matrix. Extracellular matrix receptors on the cell surface mediate cell adhesion, migration, and invasion. Recent work with transformed cells indicates that these cells use the cell surface matrix receptor CD44 for migration and invasion. In this study, we examine whether lung fibroblasts, isolated from patients dying with acute alveolar fibrosis, use CD44 to invade a fibrin matrix. Consistent with a role for CD44 in mediating fibroblast invasion and subsequent tissue fibrosis, immunohistochemical analysis of lung tissue from patients who died from acute alveolar fibrosis after lung injury reveals CD44-expressing mesenchymal cells throughout newly formed fibrotic tissue. PCR, Western, and immunoprecipitation analysis demonstrate that the 85-kD CD44 isoform is expressed by acute lung injury fibroblasts. Consistent with a role in mediating matrix adhesion and migration ultrastructurally, CD44 was found uniformly over the cell surface and was found densely labeling filopodia and lamellipodia, highly motile structures involved in cell migration. To determine if lung injury fibroblasts use CD44 to invade fibrin, a fibrin gel model of fibrosis was used. By blocking the function of CD44 with monoclonal antibodies, fibroblast invasion into a fibrin matrix was inhibited. To examine the mechanism by which CD44 mediates fibroblast invasion, the role of CD44 in fibroblast migration and adhesion was evaluated. Anti-CD44 antibody blocked fibroblast migration on the provisional matrix proteins fibronectin, fibrinogen, and hyaluronic acid. Additionally, fibroblast CD44 mediated adhesion to the provisional matrix proteins fibronectin, fibrin, and hyaluronic acid, but not to laminin, a component of the basement membrane. These findings support the hypothesis that fibroblast CD44 functions as an adhesion receptor for provisional matrix proteins and is capable of mediating fibroblast migration and invasion of the wound provisional matrix resulting in the formation of fibrotic tissue.