Experimental variation of the level and the ratio of angiogenic and osteogenic signaling affects the spatiotemporal expression of bone-specific markers and organization of bone formation in ectopic sites
- 262 Downloads
The aim of the present study was to test the hypothesis that the ratio of angiogenic and osteogenic signaling affects ectopic bone formation when delivered in different amounts.
Materials and methods
Porous composite PDLLA/CaCO3 scaffolds were loaded with rhBMP2 and rhVEGF in different dosage combinations and implanted into the gluteal muscles of 120 adult male Wistar rats. Bone formation and expression of alkaline phosphatase and Runx2 were quantified by histomorphometry. Spatial distribution across the scaffolds was assessed by using a grid that discriminated between the periphery and center of the scaffolds.
The evaluation showed that the combined delivery of bone morphogenetic protein BMP2 and VEGF in different dosage combinations did not enhance the overall quantity of ectopic bone formation compared to the delivery of BMP2 alone. The addition of VEGF generally upregulated Runx2 after 4 weeks, which may have retarded terminal osteogenic differentiation. However, slow combined delivery of 1.5–2.0 μg BMP2 combined with 50 ng VEGF165 over a period of 5 weeks supported a more even distribution of bone formation across the implanted scaffolds whereas higher amounts of VEGF did not elicit this effect.
The findings suggest that structural organization rather than the quantity of ectopic bone formation is affected by the dosage and the ratio of BMP2 and VEGF levels at the observed intervals.
The development of carriers for dual growth factor delivery has to take into account the necessity to carefully balance the ratio of growth release.
KeywordsBone morphogenetic protein Vascular endothelial growth factor Osteogenesis Runx2 Alkaline phosphatase
The authors greatly value the help of Mrs. Jutta Schulz and Dr. Behrens during the laboratory experiments and the support of Dr. Sven. Backhaus and Dr. Thomas Annen during the preparation of the scaffolds. They also wish to thank Mrs. Kant and Mrs. Schäfer for their valuable help in histologic processing.
N. Moser performed the animal experiments, scanned the histologic specimens, and co-authored the manuscript. J. Goldstein performed the histomorphometric evaluation of bone formation. P. Kaufmann scanned the histologic specimens, collected the data, and co-authored the manuscript. M. Epple fabricated the scaffolds. H. Schliephake performed the morphometry of immunohistochemical staining and co-authored the manuscript.
This work has been funded by a grant from the Federal Ministry of Education and Research (BMBF) (No. 13N10003).
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
All applicable international, national, and/or institutional guidelines for the care and use of animals were followed.
For this type of study, formal consent is not required.
- 5.Hernandez A, Reyes R, Sanchez E, Rodriguez-Evora M, Delgado A, Evora C (2012) In vivo osteogenic response to different ratios of BMP-2 and VEGF released from a biodegradable porous system. J Biomed Mater Res Part A 100A:2382–2391Google Scholar
- 8.Lohse N, Moser N, Backhaus S, Annen T, Epple M, Schliephake H (2015) Continuous delivery of rhBMP2 and rhVEGF165 at a certain ratio enhances bone formation in mandibular defects over the delivery of rhBMP2 alone—an experimental study in rats. J Control Release 220:201–209CrossRefPubMedGoogle Scholar
- 10.Geuze RE, Theyse LF, Kempen DH, Hazewinkel HA, Kraak HY, Oner FC, Dhert WJ, Alblas J (2012) A differential effect of bone morphogenetic protein-2 and vascular endothelial growth factor release timing on osteogenesis at ectopic and orthotopic sites in a large-animal model. Tissue Eng Part A 18:2052–2062CrossRefPubMedPubMedCentralGoogle Scholar
- 12.Young S, Patel ZS, Kretlow JD, Murphy MB, Mountziaris PM, Baggett LS, Ueda H, Tabata Y, Jansen JA, Wong M, Mikos AG (2009) Dose effect of dual delivery of vascular endothelial growth factor and bone morphogenetic protein-2 on bone regeneration in a rat critical-size defect model. Tissue Eng Part A. 15:2347–2362CrossRefPubMedPubMedCentralGoogle Scholar
- 23.Sacchetti B, Funari A, Remoli C, Giannicola G, Kogler G, Liedtke S, Cossu G, Serafini M, Sampaolesi M, Tagliafico E, Tenedin E, Saggio I, Robey PG, Riminucci M, Bianco P (2016) No identical “mesenchymal stem cells” at different times and sites: human committed progenitors of distinct origin and differentiation potential are incorporated as adventitial cells in microvessels. Stem Cells Rep 14:897–913CrossRefGoogle Scholar