COMBINED EFFECT OF GLYCOSAMINOGLYCAN AND MECHANICAL STIMULATION ON THE IN VITRO BIOMECHANICS OF TISSUE ENGINEERED TENDON CONSTRUCTS

摘要

Tissue engineering offers an attractive alternative to direct repair or reconstruction of injuries to tendons, ligaments and capsular structures that represent almost 45% of the 32 million musculoskeletal injuries that occur each year in the United States [1]. Mesenchymal stem cell (MSC)-seeded collagen constructs are currently being used by our group to repair tendon injuries in the rabbit model [2, 3]. Although these cell-assisted repairs exhibit 50% greater maximum force and stiffness at 12 weeks compared to values for natural repair, tissues often lack the maximum force sufficient to resist the peak in vivo forces acting on the repair site [3]. Our laboratory has previously demonstrated that in vitro construct stiffness and repair stiffness at 12 weeks post surgery are positively correlated [4]. Therefore, in an effort to further improve the repair outcome using tissue engineering, we continue our investigation of scaffold materials to create stiffer MSC-collagen constructs. Our group has recently evaluated two scaffold materials, type I collagen sponges fabricated within the Engineered Skin Lab (ESL, Shriners Hospitals for Children) by a freezing and lyophilization process with and without glycosaminoglycan (chondroitin-6-sulfate; GAG) [5] and found the ESL sponges to significantly improve biomechanical properties of the constructs compared to sponges we currently use in the lab (P1076, Kensey Nash Corporation, Exton, PA). This study also demonstrated that GAG significantly upregulates collagen type I, decorin, and fibronectin gene expression (unpublished results). Another strategy to improve a construct's in vitro biomechanical properties is in vitro mechanical stimulation. In vitro mechanical stimulation has been reported to induce fibrillar orientation [6], cell alignment [7, 8], increased proliferation [7, 9], and increased secretion of growth factors (TGF-β, bFGF, and PDGF) [10] and collagen [7]. Mechanical stimulation of tenocytes on monolayer show similar results, exhibiting increased DNA synthesis [9] and expression of novel genes not typically seen in tendon cells [11]. The purpose of this study was to understand how the two new scaffold materials, ESL sponges with and without GAG, influence the response of MSCs to mechanical stimulation. As the addition of GAGs to the ESL sponge has been previously shown to upregulate genes associated with the extracellular matrix, we hypothesize that mechanical stimulation would further improve the in vitro biomechanical properties of MSC-seeded sponge with GAG due to the synergistic effects of GAG and mechanical stimulation.