Title: Interface Scaffold Design Principles for Integrative Cartilage Regeneration
Description: Osteoarthritis is a degenerative joint disease characterized by painful, progressive articular cartilage lesions that deteriorate joint function. It remains leading cause of disability in the United States, affecting nearly 30 million Americans with increasing prevalence in the aging population, which has resulted in an annual economic burden of $128 billion. Symptomatic, full thickness cartilage injuries often require surgical intervention, because the tissue is predominantly avascular and thus has a limited self-healing capacity. However, clinical management strategies including matrix-induced autologous chondrocyte implantation and osteochondral grafting are inadequate in the long-term due to poor integration of cartilage grafts with surrounding host cartilage and subchondral bone. In addition to physical congruence between graft and host cartilage, a structural or chemically functional barrier that limits osseous invasion into the cartilage compartment is critical in order to maintain the integrity of repaired cartilage.

Given these significant clinical challenges, the objective of this thesis is to establish design principles for homotypic and heterotypic tissue integration via a cup-shaped fibrous scaffold system that encapsulates cartilage grafts (autologous or engineered), and integrates them simultaneously with host cartilage and bone at their respective interfaces. Additionally, to facilitate clinical translation of the scaffold cup, an innovative “green electrospinning” method is developed using FDA Q3C Class 3 solvents with minimal manufacturing impact on the environment. It is hypothesized that, to fuse cartilage grafts with host cartilage, the walls of the envisioned cup can direct cell migration directly to the graft-host cartilage interface via chemotactic agent delivery, where scaffold electroactivity will encourage cells to deposit a structurally contiguous neocartilage matrix. At the boundary between the graft and underlying bone, the scaffold cup base will mimic the topography and ceramic chemistry of the native osteochondral interface while preventing bone vasculature from growing upwards into the cartilage, guided by the hypothesis that this will enable the formation of a calcified cartilage interface layer that merges the graft and subchondral bone.

To test these hypotheses, this thesis began with green electrospinning the scaffold cup walls incorporated with insulin-like growth factor 1 (IGF-1), a well-established chondrocyte chemoattractant that induced cell migration from cartilage autografts towards resulting fibers. Additionally, the walls contained an optimized dose of graphite nanoparticles to impart electroactivity to the fibers. Mimicking the fixed charge density of cartilage in this way promoted chondrocyte proliferation and biosynthesis of a hyaline cartilage-like matrix in vitro, with selective regulation of proteoglycans (biglycan and decorin) and downregulation of collagen type I compared to a graphite-free fiber control. Moreover, the graphite fibers sequestered IGF-1, sustaining release of the growth factor and improving functional graft-cartilage shear integration strength in vitro. In a full thickness defect osteochondral construct repaired with the scaffold cup and implanted subcutaneously in rat dorsi, localized IGF-1 delivery promoted graft-host cartilage interface matrix elaboration with significantly greater integration strength measured with graphite in the cup walls.

For integration with subchondral bone, design criteria for the scaffold cup base were set by quantitatively mapping the compositional and morphometric characteristics of healthy and osteoarthritic human osteochondral tissues, and evaluating FEBio simulations of calcified cartilage and polymer-ceramic composite fibers in silico. These analyses established the need for an interdigitating mesh topography and ceramic particle incorporation, which minimize shear and distribute loading across the fibers, respectively, recapitulating the osteochondral interface’s force gradient from cartilage to bone in order to functionally integrate the tissues. Thus, the dose of calcium deficient apatite (CDA) nanoparticles, which capture the high calcium-phosphate ratio and semi-crystalline atomic structure of native bone mineral, was optimized to promote deep zone chondrocyte growth and biosynthesis of a calcified cartilage matrix in vitro. Moreover, CDA enhanced remodeling of the interface in vivo, with undulating fibers preventing osseous upgrowth.

Taken together, these findings delineate the importance of strategic biomimicry in scaffold design, specifically with regards to interface regeneration and cartilage integration. The proposed approach is unique in that it utilized cell homing and an electroactive substrate to mimic properties of the cartilage matrix, with a strategy for simultaneous graft integration with host cartilage and bone. Moreover, the cup design is readily adaptable to current cartilage repair techniques including press-fit autografting and cell-based graft implantation, as well as emerging tissue engineered grafting strategies. Beyond cartilage repair, the scaffold design criteria established in this thesis are broadly applicable to integrating other complex tissue systems, and may inform the regeneration of critical soft-soft (muscle-tendon) and soft-hard (tendon- or ligament-bone) interfaces in the musculoskeletal system.