Demineralized bone matrix scaffold modified with mRNA derived from osteogenically pre-differentiated MSCs improves bone repair

Abstract

Gene therapy based on mRNA provides a promising approach for bone regeneration. Quick mRNA translation and controlled protein production could be earned by implantation of mRNA-activated scaffold in bone remodeling region. Furthermore, the expression levels of osteogenic-related mRNA in the cytoplasm of osteogenically pre-differentiated mesenchymal stem cells (MSCs) were high and the expression levels were different at different stages of osteogenically differentiated MSCs. This study intended to investigate the effect of osteoinductive-mRNAs (Oi-mRNAs), derived from osteogenically pre-differentiated MSCs at various stages (Day 1 (Oi1-mRNA), Day 3 (Oi3-mRNA), Day 7 (Oi7-mRNA), Day 14 (Oi14-mRNA) and Day 21 (Oi21-mRNA), respectively), on the osteogenic differentiation of MSCs. Further, the Oi-mRNAs combined with cationic polymer polyethylenimine (PEI) were loaded onto demineralized bone matrix (DBM) scaffold (Oi-mRNA/DBM). The results revealed that the Oi1-mRNA, Oi3-mRNA and Oi21-mRNA had no obvious effect on the osteogenic differentiation of MSCs, while the Oi7-mRNA increased the expression of alkaline phosphatase (ALP) and the Oi14-mRNA significantly promoted the expression of osteocalcin (OC) and osteopontin (OPN), and calcium deposition. In addition, the Oi14-mRNA/DBM scaffold could significantly enhance extracellular matrix (ECM) secretion and new collagen formation of MSCs. After being implanted into rat critical-sized cranium defect model, the Oi14-mRNA/DBM scaffold could promote the infiltration of cells and repair of bone defect in vivo. The DBM scaffold loaded with mRNA encoding osteoinductive protein may provide a powerful tool for bone defect repair.