Connective tissue remodeling provides mammals with a rapid mechanism to repair wounds after injury. Inappropriate activation of this reparative process leads to scarring and fibrosis. Here, we studied the effects of platelet-derived growth factor receptor-β blockade in vivo using the platelet-derived growth factor receptor (PDGFR)-β inhibitor imatinib mesylate on tissue repair. After 7 days, healing of wounds was delayed with significantly reduced wound closure and concomitant reduction in myofibroblast frequency, expression of fibronectin ED-A, and collagen type I. Using a collagen type I transgenic reporter mouse, we showed that inhibiting PDGFR-β activation restricted the distribution of collagen-synthesizing cells to wound margins and dramatically reduced cell proliferation in vivo. By 14 days, treated wounds were fully closed. Blocking PDGFR-β signaling did not prevent the differentiation of myofibroblasts in vitro but potently inhibited fibroblast proliferation and migration. In addition, PDGFR-β inhibition in vivo was accompanied by abnormal microvascular morphogenesis reminiscent of that observed in PDGFR-β^−/− mice with significantly reduced immunostaining of the pericyte marker NG2. Imatinib treatment also inhibited pericyte proliferation and migration in vitro. This study highlights the significance of PDGFR-β signaling for the recruitment, proliferation, and functional activities of fibro-blasts and pericytes during the early phases of wound healing.