Protein-releasing polymeric scaffolds induce fibrochondrocytic differentiation of endogenous cells for knee meniscus regeneration in sheep

Sheep knee meniscus was regenerated by spatiotemporal delivery of two recombinant human proteins in 3D-printed bioresorbable scaffold. Bioprinting Cartilage Scaffolds Weak in the knees? You may need more cartilage. Because donor cartilage as well as synthetic products are scarce, Lee et al. devised a way to regenerate the knee meniscus, using the native tissue from sheep knees as inspiration. The authors three-dimensionally (3D) printed poly-ε-caprolactone (PCL) scaffolds based on the anatomically correct dimensions of the native meniscus. These scaffolds were loaded at specific locations with polymeric microspheres containing one of two growth factors, to be released sequentially. Connective tissue growth factor (CTGF) was released first, followed by transforming growth factor–β3 (TGFβ3). Control over both spatial and temporal protein release allowed for zone-specific matrix development: type I collagen in the outer zone and type II collagen in the inner zone, similar to the native meniscus. It is believed that such growth factor release induced endogenous stem cells to differentiate into fibrochondrocytes—the cells that make up the meniscus. In vivo in a large animal (sheep) model, the implanted scaffolds led to tissue regeneration with desired mechanical properties. If translated to humans, this acellular biomaterial would fulfill the major unmet need of repairing deteriorated cartilage in not only the knees but also tendon-bone junctions, the intervertebral discs of the spine, and the temporomandibular joint. Regeneration of complex tissues, such as kidney, liver, and cartilage, continues to be a scientific and translational challenge. Survival of ex vivo cultured, transplanted cells in tissue grafts is among one of the key barriers. Meniscus is a complex tissue consisting of collagen fibers and proteoglycans with gradient phenotypes of fibrocartilage and functions to provide congruence of the knee joint, without which the patient is likely to develop arthritis. Endogenous stem/progenitor cells regenerated the knee meniscus upon spatially released human connective tissue growth factor (CTGF) and transforming growth factor–β3 (TGFβ3) from a three-dimensional (3D)–printed biomaterial, enabling functional knee recovery. Sequentially applied CTGF and TGFβ3 were necessary and sufficient to propel mesenchymal stem/progenitor cells, as a heterogeneous population or as single-cell progenies, into fibrochondrocytes that concurrently synthesized procollagens I and IIα. When released from microchannels of 3D-printed, human meniscus scaffolds, CTGF and TGFβ3 induced endogenous stem/progenitor cells to differentiate and synthesize zone-specific type I and II collagens. We then replaced sheep meniscus with anatomically correct, 3D-printed scaffolds that incorporated spatially delivered CTGF and TGFβ3. Endogenous cells regenerated the meniscus with zone-specific matrix phenotypes: primarily type I collagen in the outer zone, and type II collagen in the inner zone, reminiscent of the native meniscus. Spatiotemporally delivered CTGF and TGFβ3 also restored inhomogeneous mechanical properties in the regenerated sheep meniscus. Survival and directed differentiation of endogenous cells in a tissue defect may have implications in the regeneration of complex (heterogeneous) tissues and organs.