Spatially patterned microribbon-based hydrogels induce zonally-organized cartilage regeneration by stem cells in 3D

Regenerating cartilage with biomimetic zonal organization, which is critical for tissue function, remains a great challenge. The objective of this study was to evaluate the potential of spatially-patterned, multi-compositional, macroporous, extracellular matrix-based microribbon (µRB) µRB scaffolds to regenerate cartilage with biochemical, mechanical, and morphological zonal organization by mesenchymal stem cells (MSCs) compared to conventional multi-layer nanoporous hydrogels. MSCs were seeded in either trilayer microribbon (µRB) or hydrogel (HG) scaffolds that were composed of layered biomaterial compositions that had been chosen for their ability to differentiate MSCs into chondrocytes with zonal properties. To mimic the aligned collagen morphology in the superficial layer of native cartilage, an additional experimental group added MSC-laden aligned µRBs to the surface of the superficial layer of a µRB trilayer. Tuning µRB alignment and compositions in different zones led to zonal-specific responses of MSCs to create neocartilage with zonal biochemical, morphological, and mechanical properties, while trilayer HGs led to minimal cartilaginous deposition overall. Trilayer µRBs created neocartilage exhibiting significant increases in compressive modulus (up to 456 kPa) and > 4-fold increase in sGAG production from superficial to deep zones. Aligned gelatin µRBs in the superficial zone further enhanced biomimetic mimicry of the produced neocartilage by leading to robust collagen deposition and superficial zone protein production. STATEMENT OF SIGNIFICANCE: Regenerating cartilage with zonal organization using mesenchymal stem cells (MSCs) remains a great challenge. We developed a spatially-patterned, gradient, macroporous, trilayer microribbon (µRB) scaffold that we used to engineer MSC-based neocartilage with zonal trends that match native cartilage in many aspects, including collagen, sGAG, superficial zone protein, and compressive moduli. This is in direct contrast to conventional trilayer nanoporous hydrogels which led to minimal cartilage deposition and weak mechanical properties. It took only 21 days for MSC-seeded trilayer µRB scaffolds to reach cartilage-mimicking compressive moduli without requiring high cell seeding density, which has never been reported before. While this paper focuses on cartilage zonal organization, gradient µRB scaffolds can be used to repair other tissue interfaces such as osteochondral defects.