The purpose of this study was to characterize the time course of matrix metalloprotease-3 (MMP-3) and tissue inhibitor of metalloprotease-1 (TIMP-1) expression in mouse tibialis anterior (TA) muscle post-injury. Mice were anesthetized, the TA muscle exposed, and injury induced by applying a cold steel probe (-79 degrees C) to the muscle for 10 s. Muscle was collected from uninjured and injured legs at 3, 10, 24, 48, and 72 h post-injury. qRT-PCR, immunoblotting, and immunohistochemistry were used to quantify/localize MMP-3 and TIMP-1. MMP-3 transcripts increased 19- and 12-fold, 10 and 24 h post-injury (p < 0.01), respectively. TIMP-1 transcript levels increased 9-, 34-, and 60-fold, 10, 24, and 48 h post-injury (p = 0.01), respectively, with a subsequent decrease 72 h post-injury (p < 0.01). Protein levels of the pro-form of MMP-3 increased within 3 h post-injury and remained elevated (p < 0.05). Active MMP-3 decreased over time, reaching a 72% decrease 72 h post-injury (p < 0.05). TIMP-1 protein decreased 75% within 3 h post-injury, returning to baseline by 72 h post-injury. In response to injury, injured skeletal muscle preferentially produces increased levels of the latent form of the MMP-3 protein with a concomitant decrease in the active form, and a significant decrease in TIMP-1 expression. The altered pattern of MMP-3/TIMP-1 expression may be due to alterations in post-transcriptional mechanisms that are responsible for specific regulation of the MMP-3/TIMP-1 system. These data suggest that there is a disproportionate regulation of the MMP-3/TIMP-1 system following traumatic injury and this response may contribute to impaired extracellular matrix remodeling.