INTRODUCTION: Adhesions after flexor tendon repair are due to overhealing or scarring. Early mobilization has clinically been shown to reduce adhesion formation and improve functional outcomes, but little is known about the cellular mechanisms mediating this preventative treatment. As tendons glides within their synovial-filled sheaths, the tendon cells may be experiencing fluid-induced mechanical shear stresses. To further elucidate how mechanical stresses may be contributing to the clinical efficacy of early mobilization, we examine in an in vitro model system the effects of fluid-induced shear stress on tenocyte expression patterns.
METHODS: Primary intrinsic tenocyte cultures were established from flexor tendons of Sprague-Dawley rats and sheared at 50 rpm (0.41 Pa) using a cone viscometer for 6 or 12 hours. Total RNA was harvested and compared to time-matched unsheared controls using cDNA microarrays containing 42,000 gene elements. Microarray data were validated for 5 genes by Northern blot analysis on biological replicates.
RESULTS: Microarray analysis demonstrated that mechanical shear stress induced an overall “anti-fibrotic” expression pattern with decreased transcription of Collagen Type I and Type III. Shear stress down-regulated pro-fibrotic molecules in the PDGF, IGF, and FGF signaling pathways and induced an overall decrease in TGF-b signaling with downregulation of TGF-b2, TGF-b3, TGF-RI, TGF-RII, and intercellular SMAD expression. Moreover, sheared tendon cells increased expression of MMPs and decreased expression of TIMPs, an expression pattern consistent with an anti-fibrotic increase in extracellular matrix degredation. Interestingly, several genes thought to play a role in tendon healing were increased with mechanical shear such as members of the BMP and VEGF cytokine families. In addition, novel genes were identified that may play important roles in the response of tendon cells to mechanical shear. Northern blot validation of our results for TGF-b1, TGF-b2, TGF-b3, Collagen Type I and Collagen Type III demonstrated excellent correlation with microarray data.
CONCLUSIONS: This study provides insight into the mechanisms by which early mobilization decreases adhesion formation without impaired tendon healing. Expression data from this unique model of decreased scar formation in combination with expression data from other models of decreased scarring studied in our laboratory may allow for the identification of novel gene-therapy targets aimed at controlling the scarring process in patients after flexor tendon surgery.