Gene‐mediated osteogenic differentiation of stem cells by bone morphogenetic proteins‐2 or ‐6

Abstract

Bone marrow-derived mesenchymal stem cells (BMDMSC) hold promise for targeted osteogenic differentiation and can be augmented by delivery of genes encoding bone morphogenetic proteins (BMP). The feasibility of promoting osteogenic differentiation of BMDMSC was investigated using two BMP genes in monolayer and three-dimensional alginate culture systems. Cultured BMDMSC were transduced with E1-deleted adenoviral vectors containing either human BMP2 or BMP6 coding sequence under cytomegalovirus (CMV) promoter control [17:1 multiplicities of infection (moi)] and either sustained in monolayer or suspended in 1 mL 1.2% alginate beads for 22 days. Adenovirus (Ad)-BMP-2 and Ad-BMP-6 transduction resulted in abundant BMP-2 and BMP-6 mRNA and protein expression in monolayer culture and BMP-2 protein expression in alginate cultures. Ad-BMP-2 and Ad-BMP-6 transduced BMDMSC in monolayer had earlier and robust alkaline phosphatase-positive staining and mineralization and were sustained for a longer duration with better morphology scores than untransduced or Ad-b-galactosidase-transduced cells. Ad-BMP-2-and, to a lesser degree, Ad-BMP-6-transduced BMDMSC suspended in alginate demonstrated greater mineralization than untransduced cells. Gene expression studies at day 2 confirmed an inflammatory response to the gene delivery process with upregulation of interleukin 8 and CXCL2. Upregulation of genes consistent with response to BMP exposure and osteogenic differentiation, specifically endochondral ossification and extracellular matrix proteins, occurred in BMP-transduced cells. These data support that transduction of BMDMSC with Ad-BMP-2 or Ad-BMP-6 can accelerate osteogenic differentiation and mineralization of stem cells in culture, including in three-dimensional culture. BMP-2-transduced stem cells suspended in alginate culture may be a practical carrier system to support bone formation in vivo. BMP-6 induced a less robust cellular response than BMP-2, particularly in alginate culture. ß

rhBMP-2 Modulation of Gene Expression in Infected Segmental Bone Defects

“…The fourth group received an ACS wetted only with 0.1 mL sterile water and 0.1 mL normal saline (carrier-only group). The carrier-only and rhBMP-2 only conditions were included as controls to verify bone growth would occur without the presence of infection, as would be expected [55]. Six animals from each of the four groups were euthanized at 1, 2, or 4 weeks after surgery.…”

Osteogenic gene regulation and relative acceleration of healing by adenoviral‐mediated transfer of human BMP‐2 or ‐6 in equine osteotomy and ostectomy models

“…9 BMP-6 genes showed more mature in vivo ectopic bone formation within murine muscle than BMP-2 genes, 10 but BMP-2 genes showed more robust in vitro osteogenesis in equine bone marrow stem cells than BMP-6 genes. 11 The effect of BMP-6 on bone formation has received recent research interest, but its in vivo osteoinductive efficacy at bone healing sites relative to the BMP-2 gene has not been reported. Additionally, despite the comprehensive comparisons of bone formation by endogenous rhBMPs, 12 the relative osteoinductivity among exogenous BMPs administered by gene delivery is only lightly studied.…”

Genetically Engineered Mesenchymal Stem Cells: The Ongoing Research for Bone Tissue Engineering

“…BMP-6 is characterized by maintaining joint integrity in adults. Interestingly, BMP-6-modified bone marrow MSCs (BM-MSCs) demonstrate accelerated osteogenic differentiation and mineralization in vitro, but the result is less robust than BMP-2-modified BM-MSCs (Zachos et al, 2006). BMP-9 only has a moderate osteoinductive capability (Celeste et al, 1990).…”

“…Adenovirus vector is one of the most popular vectors for MSC gene therapy aimed at BTE Dumont et al, 2002;Olmsted-Davis et al, 2002;Blum et al, 2003;Dayoub et al, 2003;Gugala et al, 2003;Li et al, 2003;Park et al, 2003;Riew et al, 2003;Sugiyama et al, 2003;Tsuda et al, 2003;Yang et al, 2003;Kumar et al, 2004;Zheng et al, 2004;Zhao et al, 2005;Egermann et al, 2006;Feeley et al, 2006;Zachos et al, 2006). Adenovirus vector is characterized by efficient transduction of quiescent cells and a relatively high capacity for transgene insertion.…”