Promising Efficacy ofEscherichia coliRecombinant Human Bone Morphogenetic Protein-2 in Collagen Sponge for Ectopic and Orthotopic Bone Formation and Comparison with Mammalian Cell Recombinant Human Bone Morphogenetic Protein-2

Abstract

Nonglycosylated recombinant human bone morphogenetic protein (rhBMP)-2 prepared in Escherichia coli (E. coli rhBMP-2) has recently been considered as an alternative to mammalian cell rhBMP-2. However, its clinical use is still limited owing to lack of evidence for osteogenic activity comparable with that of mammalian cell rhBMP-2 via microcomputed tomography-based analysis. Therefore, this study aimed to evaluate the ability of E. coli rhBMP-2 in absorbable collagen sponge to form ectopic and orthotopic bone and to compare it to that of mammalian rhBMP-2. In vitro investigation was performed to study osteoblast differentiation of human mesenchymal stromal cells. Both types of rhBMP-2 enhanced proliferation, alkaline phosphatase activity, and matrix mineralization of human mesenchymal stromal cells at similar levels. Similar tendencies were observed in microcomputed tomography analysis, which determined bone volume, fractional bone volume, trabecular thickness, trabecular separation, bone mineral density, and other characteristics. Histology from an in vivo osteoinductivity test and from a rat calvarial defect model demonstrated a dose-dependent increase in local bone formation. The E. coli rhBMP-2 group (5 μg) not only induced complete regeneration of an 8-mm critical-sized defect at 4 weeks, but also led to new bone with the same bone mineral density as normal bone at 8 weeks, with the same efficiency as that of mammalian cell rhBMP-2 (5 μg). These uniformly favorable results provide evidence that the osteogenic activity of E. coli rhBMP-2 is not inferior to that of mammalian cell rhBMP-2 despite its low solubility and lack of gylcosylation. These results suggest that the application of E. coli rhBMP-2 in absorbable collagen sponge may be a promising equivalent to mammalian cell rhBMP-2 in bone tissue engineering.

Analysis of hydrolyzable polyethylene glycol hydrogels and deproteinized bone mineral as delivery systems for glycosylated and non-glycosylated bone morphogenetic protein-2

“…In agreement with literature [9,[14][15][16][17], we found the biological activity of the glycosylated and nonglycosylated rhBMP-2 to be alike in vitro ( Figure 7). The biological activity of both variants drops rapidly in cell culture medium at 37 °C.…”

“…Despite the fact that E. coli-derived proteins are nonglycosylated, the biological potential of non-glycosylated rhBMP-2 has been reported to have bone-inducing activity similar to the glycosylated form [14][15][16][17]. In a human clinical trial, we demonstrated the efficacy of non-glycosylated rhBMP-2 in a bone augmentation procedure [9].…”

Modulation of Stromal Cell-Derived Factor-1/CXC Chemokine Receptor 4 Axis Enhances rhBMP-2-Induced Ectopic Bone Formation

“…In our study, the concentrations used of 5 mg/kg of AMD3100, 18 200 ng of SDF-1 per ACS implant, 45,46 and 10 mg of BMP-2 [47][48][49] were based on previous publications from other groups. Additionally, one publication described the release kinetics of SDF-1 from collagen scaffolds, reporting that there is a rapid release of 50% of the SDF-1 in the first 24 h, and, therefore, a high gradient, followed by a notably slower release of an additional 10% for at least a week thereafter.…”

Bone Regeneration by Transplantation of Human Mesenchymal Stromal Cells in a Rabbit Mandibular Distraction Osteogenesis Model

“…To measure the presence of newly formed bone, a rectangular area of the central position in the distracted callus was selected as the region of interest (ROI) in two-dimensional images, as described previously. 30 The pixel zone representing ossification in the defined ROI was then reconstructed in 3D by creating a volume of interest in the lower and upper ranges of the threshold using grayscale units. After using CTAn 1.8 on each reconstructed BMP file, microarchitecture parameters, including the bone volume to the total volume ratio (BV/TV), trabecular thickness (Tb.Th), and trabecular separation (Tb.Sp), were obtained using a CTanalyzer in a direct 3D-based analysis of a surface-rendered volume model according to the manufacturer's instructions.…”