Bone regeneration using stem cells and growth factors has disadvantages while needing to use supraphysiological growth factors concentrations. Gene therapy has been proposed as alternative, but also has limitation. Messenger RNA (mRNA)-based transcript therapy is a novel approach that may solve plasmid DNA-based gene therapy limitations. Although much more efficient in delivering genes into the cell, mRNA is unfortunately unstable and immunogenic. However, recent reports indicated that chemical modifications of the mRNA molecule can improve stability and toxicity. In this study, we have combined biomaterials and chemically modified mRNA (cmRNA) encoding Metridia luciferase, eGFP, and bone morphogenetic protein (BMP)-2 to develop transcript-activated matrices (TAMs) for gene transfer to stem cells. BMP-2 cmRNA was produced to evaluate its feasibility in stimulating osteogenic differentiation. Fibrin gel and micro-macro biphasic calcium phosphate (MBCP) granules were used as biomaterials. A sustained release of hBMP-2 cmRNA from both biomaterials was observed during 7 days. This occurred significantly faster from the MBCP granules compared to fibrin gels (92% from MBCP and 43% from fibrin after 7 days). Stem cells cultured in hBMP-2 cmRNA/fibrin or on hBMP-2 cmRNA/MBCP were transfected and able to secrete significant amounts of hBMP-2. Furthermore, transfected cells expressed osteogenic markers in vitro. Interestingly, although both TAMs promoted gene expression at the same level, hBMP-2 cmRNA/MBCP granules induced significantly higher collagen I and osteocalcin gene expression. This matrix also induced more mineral deposition. Overall, our results demonstrated the feasibility of developing efficient TAMs for bone regeneration by combining biomaterials and cmRNAs. MBCP synergistically enhances the hBMP-2 cmRNA-induced osteogenic pathway.
Keywords: BMP-2; biomaterials; biphasic calcium phosphate; chemically modified mRNA; fibrin; mRNA delivery; transcript-activated matrices.