The enhancement of bone regeneration by gene activated matrix encoding for platelet derived growth factor

Abstract

Gene therapy using non-viral vectors that are safe and efficient in transfecting target cells is an effective approach to overcome the shortcomings of protein delivery of growth factors. The objective of this study was to develop and test a non-viral gene delivery system for bone regeneration utilizing a collagen scaffold to deliver polyethylenimine (PEI)-plasmid DNA (pDNA) [encoding platelet derived growth factor-B (PDGF-B)] complexes. The PEI-pPDGF-B complexes were fabricated at amine (N) to phosphate (P) ratio of 10 and characterized for size, surface charge, and in vitro cytotoxicity and transfection efficacy in human bone marrow stromal cells (BMSCs). The influence of the complex-loaded collagen scaffold on cellular attachment and recruitment was evaluated in vitro using microscopic techniques. The in vivo regenerative capacity of the gene delivery system was assessed in 5 mm diameter critical-sized calvarial defects in Fisher 344 rats. The complexes were ~100 nm in size with a positive surface charge. Complexes prepared at an N/P ratio of 10 displayed low cytotoxicity as assessed by a cell viability assay. Confocal microscopy revealed significant proliferation of BMSCs on complex-loaded collagen scaffolds compared to empty scaffolds. In vivo studies showed significantly higher new bone volume/total volume (BV/TV) % in calvarial defects treated with the complex-activated scaffolds following 4 weeks of implantation (14- and 44-fold higher) when compared to empty defects or empty scaffolds, respectively. Together, these findings suggest that non-viral PDGF-B gene-activated scaffolds are effective for bone regeneration and are an attractive gene delivery system with significant potential for clinical translation.

3D Bioprinting of Developmentally Inspired Templates for Whole Bone Organ Engineering

“…The approach could also be adapted to other biofabrication methods such as inkjet printing and electrospinning where cellular and extra cellular material can be arranged in complex patterns [31] . Future work will look at recapitulating biomolecule gradients that occur during skeletal developmental processes using bioprinted patterns of plasmid DNA encoding for vascular, chondrogenic and osteogenic factors such as VEGF, PDGF, TGF-β3 and BMP-2 [32][33][34][35] . We will also explore the spatial and temporal control of these and other factors [3,36] to help engineer the micro-environment of developing bones.…”

RECENT RESEARCH ON THE GROWTH PLATE: Mechanisms for growth plate injury repair and potential cell-based therapies for regeneration

“…PDGFs in particular have been well documented to be involved in stimulating cell migration and proliferation as well as promoting angiogenesis during injury repair. In bone fracture repair, the chemotactic and mitogenic properties of PDGFs for mesenchymal cells and osteoblasts are essential for the initiation of correct bone fracture repair (Bordei 2011, Korsak et al 2013, Elangovan et al 2014. Similarly, in rats with growth plate injury, the inhibition of PDGF signalling caused a significant reduction in the amount of mesenchymal infiltrate, decreased amounts of bony and/or cartilage repair tissues, and thus an overall delay in bony repair 14 days post-injury ), highlighting the importance of PDGF expression during growth plate injury repair in regulating the fibrogenic phase and downstream tissue repair processes.…”

Intramembranous bone regeneration and implant placement using mechanical femoral marrow ablation: rodent models

“…49,50 Gene expression Gene-expression profiling assesses the relative activity of genes within the tissue being studied and can give insights into molecular mechanisms. 4,51 Tissues should be harvested and immediately placed in RNAlater or an equivalent reagent to optimally preserve the RNA. Tissue is left in RNAlater at room temperature for 24 h and then frozen at À 20 1C to undergo RNA isolation and for microarray or next-generation sequencing.…”