(574b) Stem Cell Response to Strain and Structural Variation Across CG-Cgcap Instructive Biomaterials

The tendon bone junction (TBJ) is a unique anatomical zone which transmits high tensile loads from aligned, fibrous tendon to stiff bone. Injuries to the rotator cuff account for 4.5 million physician visits per year and an annual 250,000 surgeries in the United States alone. However, current surgical techniques do not provide regeneration at the TBJ and the re-failure rate is extremely high (>90%). We are developing a porous collagen-glycosaminoglycan biomaterial containing structural anisotropy and mineralization to promote TBJ regeneration. Structurally graded scaffolds were created by lyophilizing a layered suspension of type I collagen and chondroitin sulfate (CG) with a mineralized version (CGCaP) of the same suspension. Structural alignment was introduced into scaffold variants (osteotendinous scaffolds) by using a previously established directional solidification technique. Layered scaffold variants were created by placing a divider within a mold with the mineralized suspension on one side and the non-mineralized suspension on the other, then removing the divider shortly before lyophilization. We characterized pore structure and examined cellular response after tensile strain across two multi-compartment scaffold variants, one containing only a mineral gradient (layered scaffold) and the other incorporating microstructural alignment characteristic of the native osteotendinous interface (osteotendinous scaffold). Human mesenchymal stem cells (hMSCs) were then cultured under strain overnight on each scaffold variant. We found that layered scaffolds, which did not contain structural alignment cues, induced very little change in nuclear elongation (aspect ratio) or nuclear orientation of hMSCs under strain, while the osteotendinous scaffolds induced an increase in nuclear aspect ratio and alignment with strain. Most notably, cell nuclei and actin fibers were significantly more aligned and aspect ratio was significantly increased without strain in the non-mineralized compartment of the osteotendinous scaffold. This suggests that pore architecture alone was responsible for the cellular response. Overall, these results suggest that structural cues presented within a biomaterial influence human mesenchymal stem cell behavior to a greater extent than static strain.