Immobilization management of acute phase increases healing ligament strength

This study examined how acute-phase knee immobilization affects conservative healing strength of the anterior cruciate ligament (ACL) in a rat model. Male Wistar rats underwent ACL transection followed by a controlled anterior tibial instability (CATI) procedure to promote spontaneous ligament continuity. Animals were assigned to CATI-only controls or to 1-week or 2-week postoperative immobilization (external fixation with Kirschner wires). Ligament healing was evaluated with range-of-motion tracking, histology (HE; healing scores and cell counts), immunohistochemistry for collagen types I and III, and acute-phase transcriptomics (RNA-seq at 2 weeks). At 8 weeks, ACL mechanical testing demonstrated that 2-week immobilization significantly increased maximum breaking strength and stiffness versus CATI-only and 1-week immobilization groups. Collagen I and III staining intensity was also higher in the 2-week immobilization group at 8 weeks. In vitro, cyclic stretch applied to ACL-derived fibroblasts in an IL-1β inflammatory environment suppressed ECM-related gene expression (including Col1a1/Col3a1), supporting a mechanism where limiting early mechanical strain during inflammation promotes matrix synthesis and organized regeneration. RNA-seq identified differential gene expression and enrichment of pathways including PI3K-Akt and MAPK signaling in immobilized ligaments.