Adenovirus-Based Gene Therapy for Bone Regeneration: A Comparative Analysis of In Vivo and Ex Vivo BMP2 Gene Delivery.

Adenovirus-mediated gene therapy is a promising tool in bone regenerative medicine. In this work, gene-activated matrices (GAMs) composed of (1) polylactide granules (PLA), which serve as a depot for genetic constructs or matrices for cell attachment, (2) a PRP-based fibrin clot, which is a source of growth factors and a binding gel, and (3) a BMP2 gene providing osteoinductive properties were studied. The study aims to compare the effectiveness of in vivo and ex vivo gene therapy based on adenoviral constructs with the BMP2 gene, PLA particles, and a fibrin clot for bone defect healing. GAMs with Ad-BMP2 and MSC(Ad-BMP2) show osteoinductive properties both in vitro and in vivo. However, MSCs incubated with GAMs containing transduced cells showed a more significant increase in osteopontin gene expression, protein production, Alpl activity, and matrix mineralization. Implantation of the studied matrices into critical-size calvarial defects after 56 days promotes the formation of young bone. The efficiency of neoosteogenesis and the volume fraction of newly formed bone tissue are higher with PLA/PRP-MSC(Ad-BMP2) implantation (33%) than PLA/PRP-Ad-BMP2 (28%). Thus, ex vivo adenoviral gene therapy with the BMP2 gene has proven to be a more effective approach than the in vivo delivery of gene constructs for bone regeneration.

Comparative Efficiency of Gene-Activated Matrices Based on Chitosan Hydrogel and PRP Impregnated with BMP2 Polyplexes for Bone Regeneration.

Gene therapy is one of the most promising approaches in regenerative medicine. Gene-activated matrices provide stable gene expression and the production of osteogenic proteins in situ to stimulate osteogenesis and bone repair. In this study, we developed new gene-activated matrices based on polylactide granules (PLA) impregnated with BMP2 polyplexes and included in chitosan hydrogel or PRP-based fibrin hydrogel. The matrices showed high biocompatibility both in vitro with mesenchymal stem cells and in vivo when implanted intramuscularly in rats. The use of porous PLA granules allowed the inclusion of a high concentration of polyplexes, and the introduction of the granules into hydrogel provided the gradual release of the plasmid constructs. All gene-activated matrices showed transfecting ability and ensured long-term gene expression and the production of target proteins in vitro. At the same time, the achieved concentration of BMP-2 was sufficient to induce osteogenic differentiation of MSCs. When implanted into critical-size calvarial defects in rats, all matrices with BMP2 polyplexes led to new bone formation. The most significant effect on osteoinduction was observed for the PLA/PRP matrices. Thus, the developed gene-activated matrices were shown to be safe and effective osteoplastic materials. PLA granules and PRP-based fibrin hydrogel containing BMP2 polyplexes were shown to be the most promising for future applications in bone regeneration.