Stage-Specific Collagen V Knockdown Reveals Temporal Control of Fibrillogenesis, Remodeling, and Mechanics in Tendon Healing

Collagen V is a minor fibrillar collagen that regulates type I collagen fibrillogenesis; however, its time-dependent role during the stages of adult tendon healing remains unclear. We investigated the stage-specific effects of inducible Col5a1 knockdown during tendon repair in a murine injury model. Collagen V expression was transiently suppressed during either the late inflammatory phase (tamoxifen-induced knockdown at 5 days post-injury; TM5) or early remodeling phase (tamoxifen-induced knockdown at 21 days post-injury; TM21), with outcomes assessed using gene expression, ultrastructural, and mechanical analyses. Early knockdown at 5 days post-injury, TM5, was associated with increased fibril diameter, greater fibril heterogeneity, and reduced structural mechanical properties, including decreased stiffness and maximum load. In contrast, delayed knockdown at 21 days post-injury, TM21, imposed after initial fibril organization, resulted in minimal changes in quasi-static mechanics but altered viscoelastic behavior and late-stage gene expression. An allele dose-dependent response was observed, with complete Col5a1 knockdown producing greater structural disorganization and mechanical deficits. Transcriptional changes suggested time-dependent effects on extracellular matrix regulation, including proteoglycans, remodeling enzymes, and tenogenic markers. Collectively, these findings indicate that collagen V may function as a time- and dose-sensitive contributor to tendon healing, with a critical role during early fibril organization. Disruption during this time window has lasting effects on tendon structure and mechanics, informing stage-specific therapeutic strategies for tendon repair.