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W.-K. Wu, A. D. Dick, D. O. Bates, L. B. Nicholson; VEGF Production in Macrophages Is Enhanced by Anti-Inflammatory Stimuli. Invest. Ophthalmol. Vis. Sci. 2009;50(13):3758.
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Following stimulation, macrophages can produce vascular endothelial growth factor (VEGF) which may play a role in pathological neovascularization. After they exit from the bone marrow, macrophages undergo a period of maturation that is directed by the local microenvironment. For example interferon gamma (IFN-γ) induces a M1 phenotype and IL-4 and IL-13 induce a M2 subtype. These populations are characterized by the productions of inducible nitric oxide synthase (iNOS) and arginase I respectively. In this study we have examined the relation between VEGF production and macrophage stimulation and investigated how macrophage phenotype influences to VEGF production.
Macrophages were derived from the bone marrow (BMDMs) or peritoneal cavity of C57BL/6 mice. BMDMs were generated by culture in the presence of M-CSF for 8 days in Teflon bags. These cells are naïve until they are transferred to tissue culture plastic. Cells were stimulated by prostaglandin E2 (PGE2), IFN- γ, lipopolysaccharide (LPS), and CGS21680 (CGS), a selective A2A receptor agonist. After 48 hrs supernatant was collected and we measured nitrite production by using the Griess reaction and IL-6, IL-10 and VEGF levels by ELISA. Levels of VEGF mRNA in stimulated macrophages were analyzed by Q-PCR and arginase I expression was determined by FACS.
Nitrite and IL-6 levels were up-regulated by LPS but decreased by addition of PGE2, which therefore exhibited anti-inflammatory properties in this situation. IL-10 production was stimulated by LPS and markedly up-regulated in the present of PGE2. Moreover, arginase was induced by PGE2 and CGS. This suggests that PGE2 and CGS are able to deliver anti-inflammatory signals to macrophages. Furthermore, PGE2 and CGS in combination with LPS triggered increased levels of VEGF production. In contrast, PGE2 also increased VEGF mRNA levels compared with activation with LPS/IFN-γ stimulation Taken together, this suggests that VEGF producing macrophages may be of an anti-inflammatory M2-like phenotype.
We conclude that macrophages that mature in an anti-inflammatory microenvironment may be a more effective source of VEGF than those that develop in a proinflammatory environment. This may partly explain the low level of neovascularization in uveitis. Further work will determine whether VEGF can itself modulate macrophage activation and phenotype.
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