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Abstract Background Improved understanding of the interactions between bone cells and endothelial cells involved in osteogenesis should aid the development of new strategies for bone tissue engineering. The aim of the present study was to determine whether direct communication between bone marrow stromal cells (MSC) and human umbilical vein endothelial cells (EC) could influence the osteogenic potential of MSC in osteogenic factor-free medium. Methods After adding EC to MSC in a direct-contact system, cell viability and morphology were investigated with the WST assay and immnostaining. The effects on osteogenic differentiation of adding EC to MSC was systematically tested by the using Superarray assay and results were confirmed with real-time PCR. Results Five days after the addition of EC to MSC in a ratio of 1:5 (EC/MSC) significant increases in cell proliferation and cellular bridges between the two cell types were detected, as well as increased mRNA expression of alkaline phosphatase (ALP). This effect was greater than that seen with addition of osteogenic factors such as dexamethasone, ascorbic acid and β-glycerophosphate to the culture medium. The expression of transcription factor Runx2 was enhanced in MSC incubated with osteogenic stimulatory medium, but was not influenced by induction with EC. The expression of Collagen type I was not influenced by EC but the cells grown in the osteogenic factor-free medium exhibited higher expression than those cultured with osteogenic stimulatory medium. Conclusion These results show that co-culturing of EC and MSC for 5 days influences osteogenic differentiation of MSC, an effect that might be independent of Runx2, and enhances the production of ALP by MSC.

Our recent in vitro study demonstrated that endothelial cells (ECs) might influence the differentiation of bone marrow stromal cells (BMSCs). Therefore, the aim of this study was to describe this effect in vivo, using a rat calvarial bone defect model. BMSCs were isolated from femurs of two-donor Lewis rats and expanded in α-minimum essential medium containing 10% fetal bovine serum. One fifth of BMSCs were induced and differentiated into ECs in an Endothelial Cell Growth Medium-2 and then characterized by a flow cytometry. The remaining BMSCs were cultured in freshly prepared osteogenic stimulatory medium, containing dexamethasone, ascorbic acid and β-glycerophosphate. Either BMSCs alone (BMSC-group) or co-cultured ECs/BMSCs (CO-group) were seeded into poly( L-lactide- co-1,5-dioxepan-2-one) [poly(LLA- co-DXO)] scaffolds, cultured in spinner flasks, and then implanted into symmetrical calvarial defects prepared in recipient rats. The animals were sacrificed after 2 months. The formation of new bone was evaluated by radiography and histology and by the expression of osteogenic markers using reverse transcriptase-polymerized chain reaction (RT-PCR). To investigate vessel formation, histological staining was performed with EC's markers. The radiographical and histological results showed more rapid bone formation in the CO- than in the BMSC-group. However, the expression of EC's marker was similar on both groups by histological analysis after 2 months postoperatively. Furthermore, the CO-group exhibited greater expression of osteogenic markers as demonstrated by RT-PCR. The results are consistent with the previous in vitro findings that poly(LLA- co-DXO) scaffold might be suitable candidate for bone tissue engineering. In vivo, bone regeneration was enhanced by a construct of the polymer scaffold loaded with co-cultured cells. © 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part A:, 2011. [ABSTRACT FROM AUTHOR]

Our recent in vitro study demonstrated that endothelial cells (ECs) might influence the differentiation of bone marrow stromal cells (BMSCs). Therefore, the aim of this study was to describe this effect in vivo, using a rat calvarial bone defect model. BMSCs were isolated from femurs of two-donor Lewis rats and expanded in α-minimum essential medium containing 10% fetal bovine serum. One fifth of BMSCs were induced and differentiated into ECs in an Endothelial Cell Growth Medium-2 and then characterized by a flow cytometry. The remaining BMSCs were cultured in freshly prepared osteogenic stimulatory medium, containing dexamethasone, ascorbic acid and β-glycerophosphate. Either BMSCs alone (BMSC-group) or co-cultured ECs/BMSCs (CO-group) were seeded into poly(L-lactide-co-1,5-dioxepan-2-one) [poly(LLA-co-DXO)] scaffolds, cultured in spinner flasks, and then implanted into symmetrical calvarial defects prepared in recipient rats. The animals were sacrificed after 2 months. The formation of new bone was evaluated by radiography and histology and by the expression of osteogenic markers using reverse transcriptase-polymerized chain reaction (RT-PCR). To investigate vessel formation, histological staining was performed with EC's markers. The radiographical and histological results showed more rapid bone formation in the CO- than in the BMSC-group. However, the expression of EC's marker was similar on both groups by histological analysis after 2 months postoperatively. Furthermore, the CO-group exhibited greater expression of osteogenic markers as demonstrated by RT-PCR. The results are consistent with the previous in vitro findings that poly(LLA-co-DXO) scaffold might be suitable candidate for bone tissue engineering. In vivo, bone regeneration was enhanced by a construct of the polymer scaffold loaded with co-cultured cells., (2010 Wiley Periodicals, Inc.)