Anabolic Effects of 1,25-Dihydroxyvitamin D3 on Osteoblasts Are Enhanced by Vascular Endothelial Growth Factor Produced by Osteoblasts and by Growth Factors Produced by Endothelial Cells*

Abstract

Human osteoblast-like cells (HOB) produce vascular endothelial growth factor (VEGF), the steady state level of which is stimulated by 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3]. As osteoblasts and endothelial cells are proximally located in skeletal tissue, we investigated the anabolic effects of 1,25-(OH)2D3 and VEGF on HOB cocultured with endothelial cells. When HOB with high alkaline phosphatase (Al-P) activity and human umbilical vein endothelial cells (HUVEC) with little activity were cultured together, Al-P activity increased, accompanied by an increase in cell number. When HOB and HUVEC were cultured separately, 1,25-(OH)2D3 did not directly stimulate [3H]thymidine incorporation into HUVEC, but stimulated it in the presence of HOB. VEGF did not directly stimulate the Al-P activity of HOB but stimulated it in the presence of HUVEC. The conditioned medium of HOB stimulated the proliferation of HUVEC, and this was partially blocked by anti-VEGF antibody. Conversely, the conditioned medium of HUVEC increased Al-P activity and [3H]thymidine incorporation into HOB, and this was partially blocked by antiinsulin-like growth factor I antibody and BQ-123, a specific antagonist of the endothelin-1 (ET-1) receptor. 1,25-(OH)2D3 stimulated the release of VEGF and ET-1 from HOB and HUVEC, respectively. Furthermore, the 1,25-(OH)2D3-induced release of VEGF was enhanced in HOB cocultured with HUVEC. A quantitative reverse transcription-PCR study revealed that genes for VEGF receptors (Flt-1 and KDR) were expressed in HUVEC, but not in HOB, and that 1,25-(OH)2D3 increased the levels of expression of VEGF receptor genes in endothelial cells only when cocultured with HOB. In summary, we demonstrated that 1,25-(OH)2D3 exerts an anabolic effect on osteoblasts by enhancing their production of VEGF, which stimulates its receptors on endothelial cells, followed by increased production of osteotropic growth factors, such as insulin-like growth factor I and ET-1. These in vitro findings suggest that the VEGF/VEGF receptor system may be involved in both bone formation and bone remodeling in vivo.

Electromagnetic Wave Irradiation Promotes Osteoblastic Cell Proliferation and Up-Regulates Growth Factors via Activation of the ERK1/2 and p38 MAPK Pathways

“…VEGF secreted from osteoblasts functions as a paracrine factor for neovascularization and causes endothelial cells to proliferate and stimulate blood vessel formation in bone tissue [41,42]. In accordance with our present results, it has been reported that a biphasic electrical current coupled with an electromagnetic field can stimulate cell proliferation and up-regulate VEGF expression in osteoblasts [30].…”

Gene Expression Profiles and Transcription Factors Involved in Parathyroid Hormone Signaling in Osteoblasts Revealed by Microarray and Bioinformatics

“…Endothelial cells lining the bone are in close proximity with osteoblasts and osteoprogenitor cells. Extensive cross-talk exists between endothelial cells and osteoblasts since endothelial cells stimulate the differentiation of osteoblasts by secreting endothelin-1 (ET-1), and osteoblasts stimulate proliferation of endothelial cells by secreting vascular endothelial growth factor (VEGF) (32,33). The microarray data provide a deeper understanding of this cross-talk.…”

Vascular Endothelial Growth Factor (VEGF)-induced Up-regulation of CCN1 in Osteoblasts Mediates Proangiogenic Activities in Endothelial Cells and Promotes Fracture Healing

“…VEGF is involved in conversion of the soft, cartilaginous callus to a hard, bony callus during fracture repair (12). Furthermore, VEGF has chemotactic effects on osteoblasts and may directly enhance their activity in vitro (13) or may indirectly induce their proliferation and differentiation by stimulating endothelial cells to produce osteoanabolic growth factors (14).…”