M-CSF signals through the MAPK/ERK pathway via Sp1 to induce VEGF production and induces angiogenesis in vivo.
BACKGROUND
M-CSF recruits mononuclear phagocytes which regulate processes such as angiogenesis and metastases in tumors. VEGF is a potent activator of angiogenesis as it promotes endothelial cell proliferation and new blood vessel formation. Previously, we reported that in vitro M-CSF induces the expression of biologically-active VEGF from human monocytes.


RESULTS
In this study, we demonstrate the molecular mechanism of M-CSF-induced VEGF production. Using a construct containing the VEGF promoter linked to a luciferase reporter, we found that a mutation reducing HIF binding to the VEGF promoter had no significant effect on luciferase production induced by M-CSF stimulation. Further analysis revealed that M-CSF induced VEGF through the MAPK/ERK signaling pathway via the transcription factor, Sp1. Thus, inhibition of either ERK or Sp1 suppressed M-CSF-induced VEGF at the mRNA and protein level. M-CSF also induced the nuclear localization of Sp1, which was blocked by ERK inhibition. Finally, mutating the Sp1 binding sites within the VEGF promoter or inhibiting ERK decreased VEGF promoter activity in M-CSF-treated human monocytes. To evaluate the biological significance of M-CSF induced VEGF production, we used an in vivo angiogenesis model to illustrate the ability of M-CSF to recruit mononuclear phagocytes, increase VEGF levels, and enhance angiogenesis. Importantly, the addition of a neutralizing VEGF antibody abolished M-CSF-induced blood vessel formation.


CONCLUSIONS
These data delineate an ERK- and Sp1-dependent mechanism of M-CSF induced VEGF production and demonstrate for the first time the ability of M-CSF to induce angiogenesis via VEGF in vivo.
The Dorothy M. Davis Heart and Lung Research Institute, The Ohio State University, Columbus, Ohio, United States of America.
Jennifer M
JM
Curry
Tim D
TD
Eubank
Ryan D
RD
Roberts
Yijie
Y
Wang
Nabendu
N
Pore
Amit
A
Maity
Clay B
CB
Marsh
D021581:Active Transport, Cell Nucleus; D000818:Animals; D002478:Cells, Cultured; D006801:Humans; D020935:MAP Kinase Signaling System; D016173:Macrophage Colony-Stimulating Factor / Q000502:physiology*; D051379:Mice; D009000:Monocytes; D018919:Neovascularization, Physiologic; D016329:Sp1 Transcription Factor / Q000378:metabolism / Q000502:physiology*; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis* / Q000235:genetics
D016329:Sp1 Transcription Factor; D042461:Vascular Endothelial Growth Factor A; D016173:Macrophage Colony-Stimulating Factor