The debilitating effects of rotator cuff tears and the high incidence of failure associated with current grafts underscore the clinical demand for functional solutions for tendon repair and augmentation. To address this challenge, we have designed a poly(lactide-co-glycolide) (PLGA) nanofiber-based scaffold for rotator cuff tendon tissue engineering. In addition to scaffold design and characterization, the objective of this study was to evaluate the attachment, alignment, gene expression, and matrix elaboration of human rotator cuff fibroblasts on aligned and unaligned PLGA nanofiber scaffolds. Additionally, the effects of in vitro culture on scaffold mechanical properties were determined over time. It has been hypothesized that nanofiber organization regulates cellular response and scaffold properties. It was observed that rotator cuff fibroblasts cultured on the aligned scaffolds attached along the nanofiber long axis, whereas the cells on the unaligned scaffold were polygonal and randomly oriented. Moreover, distinct integrin expression profiles on these two substrates were observed. Quantitative analysis revealed that cell alignment, distribution, and matrix deposition conformed to nanofiber organization and that the observed differences were maintained over time. Mechanical properties of the aligned nanofiber scaffolds were significantly higher than those of the unaligned, and although the scaffolds degraded in vitro, physiologically relevant mechanical properties were maintained. These observations demonstrate the potential of the PLGA nanofiber-based scaffold system for functional rotator cuff repair. Moreover, nanofiber organization has a profound effect on cellular response and matrix properties, and it is a critical parameter for scaffold design.

Biological fixation of orthopedic soft tissue grafts to bone poses a significant clinical challenge. The clinical success of soft tissue-based grafts for anterior cruciate ligament (ACL) reconstruction is limited by the lack of functional graft integration with subchondral bone. Soft tissues such as the ACL connect to subchondral bone via a complex interface whereby three distinct tissue regions (ligament, fibrocartilage, and bone) work in concert to facilitate load transfer from soft to hard tissue while minimizing stress concentration at the interface. Although a fibrovascular tissue forms at the graft-to-bone interface following surgery, this tissue is nonphysiologic and represents a weak link between the graft and bone. We propose that the re-establishment of the native multi-tissue interface is essential for biological graft fixation. In vivo observations and our in vitro monolayer co-culture results suggest that osteoblast-fibroblast interaction is important for interface regeneration. This study focuses on the design of a triphasic scaffold system mimicking the multi-tissue organization of the native ACL-to-bone interface and the evaluation of osteoblast-fibroblast interactions during three-dimensional co-culture on the triphasic scaffold. We found that the triphasic scaffold supported cell proliferation, migration and phenotypic matrix production while maintaining distinct cellular regions and phase-specific extracellular matrix deposition over time. This triphasic scaffold is designed to guide the eventual reestablishment of an anatomically oriented and mechanically functional fibrocartilage interfacial region directly on biological and synthetic soft tissue grafts. The results of this study demonstrate the feasibility of multi-tissue regeneration on a single scaffold, and the potential of interface tissue engineering to enable the biological fixation of soft tissue grafts to bone.

Achieving functional graft integration with subchondral bone poses a significant challenge for orthopaedic soft tissue repair and reconstruction. Soft tissues such as the anterior cruciate ligament (ACL) integrate with bone through a fibrocartilage interface, which minimizes stress concentrations and mediates load transfer between soft and hard tissues. We propose that biological fixation can be achieved by regenerating this fibrocartilage interface on biological or synthetic ACL grafts. This study focuses on the in vivo evaluation of a stratified scaffold predesigned to mimic the multitissue transition found at the ACL-to-bone interface. Specifically, the scaffold consists of three distinct yet continuous phases: Phase A for ligament formation, Phase B for the interface, and Phase C for the bone region. Interface-relevant cell types, specifically fibroblasts, chondrocytes, and osteoblasts, will be tri-cultured on this scaffold, and the formation of cell type- and phase-specific matrix heterogeneity as well as fibrocartilage formation will be evaluated over 8 weeks in a subcutaneous athymic rat model. Acellular scaffolds as well as scaffolds co-cultured with fibroblasts and osteoblasts will serve as controls. It was found that the triphasic scaffold supported multilineage cellular interactions as well as tissue infiltration and abundant matrix production in vivo. In addition, controlled phase-specific matrix heterogeneity was induced on the scaffold, with distinct mineral and fibrocartilage-like tissue regions formed in the tri-cultured group. Cell seeding had a positive effect on both host infiltration and matrix elaboration, which also translated into increased mechanical properties in the seeded groups compared to the acellular controls. In summary, the biomimetic and multiphasic design coupled with spatial control of cell distribution enables multitissue regeneration on the stratified scaffold, and demonstrates the potential for regenerating the interface between soft tissue grafts and bone.