title: Endothelial progenitor cells improve directly and indirectly early vascularization of mesenchymal stem cell-driven bone regeneration in a critical bone defect in rats.
abstract: Early vascularization of a composite in a critical bone defect is a prerequisite for ingrowth of osteogenic reparative cells to regenerate bone, since lack of vessels does not ensure a sufficient nutritional support of the bone graft. The innovation of this study was to investigate  the direct and indirect effects of endothelial progenitor cells (EPCs) and cotransplanted mesenchymal stem cells (MSCs) on the in vivo neovascularization activity in a critical size defect at the early phase of endochondral ossification. Cultivated human EPCs and MSCs were loaded onto β-TCP  in vitro. A critical-sized bone defect (5 mm) was created surgically in the femoral diaphysis of adult athymic rat and stabilized with an external fixateur. The bone defects were filled with β-TCP, MSCs seeded on β-TCP, EPCs seeded on β-TCP, and coculture of MSCs and EPCs seeded  on β-TCP or autologous bone of rat. After 1 week, the rats were sacrificed. Using quantitative CD34 immunohistochemistry as well as qualitative analysis of vascularization (staining of MHC and VEGF) in decalcified serial sections were performed by means of an image analysis system. Fluorescence  microscopy analyzed the direct effects and indirect effects of human implanted EPCs for vessel formation at bone regeneration site. Formation of a primitive vascular plexus was also detectable in the β-TCP, MSC, or autologous bone group, but on a significantly higher level if EPCs alone  or combined with MSCs were transplanted. Moreover, highest amount of vascularization were detected when EPCs and MSCs together were implanted. Early vascularization is improved by transplanted EPCs, which formed new vessels directly. Indeed the indirect effect of EPCs to vascularization is  much higher. Transplanted EPC release chemotactic factors (VEGF) to recruit EPCs of the host and stimulate vascularization in the bone defect. Transplantation of human EPCs displays a promising approach to improve early vascularization of a scaffold in a critical bone defect. Moreover, coculture  of EPCs and MSCs demonstrate also a synergistic effect on new vessel formation and seems to be a potential osteogenic construct for in vivo application.
authors.affiliation: Department of Trauma Surgery, Johann-Wolfgang-Goethe University, Frankfurt/Main, Germany. caroline.seebach@kgu.de
authors.forename: C
authors.initials: C
authors.lastname: Seebach
authors.forename: D
authors.initials: D
authors.lastname: Henrich
authors.forename: K
authors.initials: K
authors.lastname: Wilhelm
authors.forename: J H
authors.initials: JH
authors.lastname: Barker
authors.forename: I
authors.initials: I
authors.lastname: Marzi
mesh_terms: D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism; D001847:Bone Diseases / Q000378:metabolism / Q000473:pathology*; D001861:Bone Regeneration / Q000502:physiology*; D016025:Bone Transplantation; D002130:Calcium Phosphates / Q000494:pharmacology; D002478:Cells, Cultured; D018920:Coculture Techniques; D004727:Endothelium / Q000166:cytology*; D006801:Humans; D008297:Male; D045164:Mesenchymal Stem Cell Transplantation; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D018919:Neovascularization, Physiologic / Q000187:drug effects; D010012:Osteogenesis / Q000187:drug effects; D051381:Rats; D011923:Rats, Nude; D033581:Stem Cell Transplantation; D013234:Stem Cells / Q000166:cytology*; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism
chemical_list: D018952:Antigens, CD34; D002130:Calcium Phosphates; D042461:Vascular Endothelial Growth Factor A; C485817:beta-tricalcium phosphate