Transcript‑activated matrices with optimized mRNA designs to direct chondroblastic lineage commitment

New strategies are required to modulate early chondroblastic differentiation in a controlled and transient manner, a key challenge in cartilage tissue engineering. Transcript‑Activated Matrices (TAMs), formed by embedding mRNA within biomaterial scaffolds, offer a promising platform for directing lineage specification. Here, we optimized TAM performance by engineering two key transcript features, directly transcribed poly(A) tails and chemically modified nucleotides, to enhance expression magnitude and persistence. Using SOX9 as a model transcription factor, we evaluated how these structural modifications influence chondroblastic programming. Template-encoded poly(A) tails markedly increased transcript output, and incorporation of chemically modified nucleotides further enhanced the magnitude and persistence of SOX9 expression. Optimized SOX9‑TAMs increased SOX9 expression by approximately 5 to 40-fold in 3D cultures and promoted early chondroblastic programming, as indicated by increased ACAN (∼20‑fold) and reduced hypertrophic signaling (e.g., decreased COL‑X). In a rabbit cartilage defect model, TAM implantation induced a consistent pro‑chondroblastic transcriptional signature (e.g., SOX9 ∼50‑fold, COL-II ∼20‑fold vs. controls), despite modest early histological changes. Together, these findings reveal that rational transcript design substantially improves TAM performance and provides precise control over early chondroblastic fate. This transcript‑engineered platform represents a promising approach for directing differentiation programs in tissue engineering applications.