Precisely regulated physically-crosslinked carriers enable synergetic release of bioactive factors for MSC-mediated cartilage regeneration

Articular cartilage has limited capacity for self-repair due to its avascular nature. Successful cartilage repair requires the harmonious integration of sufficient stem cell recruitment, an optimal local microenvironment and a sustained repair timeframe. Here, we present a biocompatible, physically crosslinked silk fibroin platform with tunable β-sheet content (5-50 %) via freeze-assembly. This platform enables flexible and precise tuning of drug release kinetics without chemical cross-linkers. This system allows controlled drug release durations ranging from 1 to 35 days, suitable for both hydrophilic (MSC affinity peptide, MAP, serving a pro-recruiting role) and hydrophobic (kartogenin, KGN, pro-differentiating role) drugs. In a rat cartilage defect model, a sustained 21-day MAP release profile was identified as optimal, achieving an unprecedented high density of MSC recruitment (∼2.34 × 10⁴ cells/mm³) within a differentiating-friendly timeframe. Synchronized with KGN delivery, the co-delivery system further promoted robust hyaline cartilage regeneration. This outcome may be attributed to the effect of <i>Cdh2</i> genes involved in cell adhesion and p38 MAPK pathways. This work provides a structurally programmable, scalable strategy to achieve coordinated, high-density MSC recruitment and timed differentiation, advancing the paradigm of precise biomaterial design for tissue repair.