3D bioprinted bilayer GelMA/HAp hydrogel scaffold functionalized with miR-140-5p–modified MSCs for enhanced osteochondral regeneration

BACKGROUND: Osteochondral defects remain a major clinical challenge due to the limited intrinsic healing capacity of articular cartilage and the complex structural integration required between cartilage and subchondral bone. Tissue-engineered scaffolds offer a promising strategy for improving repair outcomes. METHODS: We developed a 3D bioprinted bilayer GelMA/hydroxyapatite (HAp) hydrogel scaffold incorporating miR-140-5p-modified mesenchymal stem cells (MSCs). The upper chondral-oriented layer consisted of 5% GelMA60, while the lower relatively stiffer HAp-containing layer comprised 5% GelMA90 with 1% HAp. Mechanical properties, cell viability, migration, and differentiation were evaluated in vitro. Osteochondral repair efficacy was further assessed in a rabbit femoral condyle defect model using macroscopic scoring, Micro-CT, and histological analysis. RESULTS: The bilayer scaffold demonstrated enhanced compressive strength and maintained favorable swelling characteristics. MSC viability remained high in both scaffold groups and exceeded 90% by day 5 after bioprinting. miR-140-5p modification significantly promoted MSC migration and upregulated chondrogenic markers. In vivo, the functionalized scaffold markedly improved cartilage surface integrity, subchondral bone reconstruction, and ICRS scores compared with controls. CONCLUSION: The 3D bioprinted bilayer GelMA/HAp scaffold combined with miR-140-5p-modified MSCs significantly enhances osteochondral repair in a rabbit model and represents a promising strategy for clinical translation.