A latent TGF-β conjugated scaffold improves neocartilage development

TGF-β is conventionally supplemented in culture medium at supraphysiologic doses to accelerate neocartilage development. While enhancing extracellular matrix (ECM) biosynthesis, supraphysiologic TGF-β further promotes non-hyaline cartilage features, including hyperplasia, hypertrophy, and ECM heterogeneities. In native cartilage, TGF-β is present in a latent complex (LTGF-β), which undergoes cell-mediated activation, leading to moderated, physiologic dosing regimens that avoid detrimental features associated with TGF-β excesses. Here, we explore a bio-inspired strategy, consisting of LTGF-β-conjugated scaffolds, providing TGF-β exposure regimens that are moderated and uniformly administered throughout the construct. We evaluate the performance of LTGF-β scaffolds with bovine chondrocyte-seeded agarose constructs compared to outcomes from active TGF-β media supplementation (MS) at a physiologic 0.3 ng/mL dose (MS-0.3), supraphysiologic 10 ng/mL dose (MS-10), or TGF-β free. LTGF-β scaffolds achieve native-matched mechanical properties and sGAG content, while providing a cell morphology and collagen distribution more reminiscent of hyaline cartilage. LTGF-β scaffolds further afford an optimal chondrogenic phenotype, marked by an up to 28-fold reduction of COL-I and 17-fold reduction of COL-X expression relative to TGF-β-free and MS-10, respectively. Further, LTGF-β scaffolds significantly reduce mechanical and biochemical heterogeneities relative to MS-0.3 and MS-10. Overall, LTGF-β scaffolds improve the composition, structure, material properties, and cell phenotype of neocartilage. STATEMENT OF SIGNIFICANCE: Inspired by native regulatory mechanisms, we introduce a latent TGF-β conjugated scaffold that enables localized, cell-mediated activation of TGF-β at physiologic levels. This approach yields neocartilage with native-matched composition and mechanical properties, while maintaining a hyaline-cartilage-like cell phenotype and morphology, thus mitigating the adverse developmental features associated with conventional active TGF-β dosing. This bio-inspired platform offers a compelling solution to current TGF-β delivery challenges to improve cartilage regeneration outcomes.