Session Title: Cell-cell Adhesion, Cell Trafficking and Angiogenesis
Session Type: Abstract Submissions (ACR)
Background/Purpose: Angiopoietin (Ang) -1 and -2 signalling to the Tie2 tyrosine kinase receptor has an essential role in blood vessel remodeling and angiogenesis. Ang-1, Ang-2 and Tie2 are all expressed in rheumatoid arthritis (RA) synovial tissue. Activated Tie2 is prominently observed in RA synovial macrophages, both Ang-1 and Ang-2 can cooperate with TNF to induce macrophage IL-6 production, and Ang-1 signaling to Tie2 promotes disease persistence and progression in early RA. Here, we examined how macrophage differentiation regulates expression of Tie2 and macrophage responses to Ang-1 and Ang-2, as well as the impact of Tie2 signalling on macrophage production of secreted products elevated in the synovial fluid (SF) of early RA patients.
Methods: Human healthy donor peripheral blood mononuclear cells were isolated from buffy coats and differentiated into macrophages in the presence of Ang-1 and Ang-2, the pro-inflammatory/classically activating cyokines GM-CSF and IFN-γ, or in the presence of the anti-inflammatory/alternatively activating cytokines M-CSF or IL-10. The expression of macrophage polarization markers and Tie2 was analyzed by flow cytometry and quantitative (q)-PCR. Macrophages were stimulated with TNF in the presence or absence of Ang-1 or Ang-2 and effects on gene expression assessed using low density q-PCR arrays, ELISA and luminex. Monocyte chemotactic responses were assessed using 96-well transwell systems.
Results: Macrophage Tie2 protein and mRNA expression was observed under all conditions, but failed to correspond to pro- or anti-inflammatory phenotypes, as it was highest in macrophages differentiated in IL-10 and IFN-g. Ang-1 and Ang-2 failed to induce macrophage polarization. Each polarization condition displayed a distinct expression profile of angiogenic factors. Ang-1 and Ang-2 alone failed to influence gene expression. Ang-1, and to a lesser extent Ang-2, synergized with TNF to stimulate expression of similar gene profiles regardless of macrophage polarization conditions. Significantly enhanced TNF-induced CXCL2, CXCL-3, CXCL-9, IL-6 mRNA expression was observed. Neither Ang-1 nor Ang-2 stimulated monocyte chemotaxis. However, conditioned medium from macrophages stimulated with TNF in combination with Ang-1 or Ang-2 demonstrated significantly enhanced chemotactic activity for monocytes, compared to conditioned medium from macrophages stimulated with TNF alone. Ang-1, but not Ang-2, significantly cooperated with TNF to induce macrophage production of cytokines elevated in the SF of early RA patients, including TGFalpha, FGF2, and IL-12B.
Conclusion: Our results demonstrate that Tie2 is functionally expressed in macrophages, regardless of macrophage polarization conditions. Stimulation of macrophage Tie2 with Ang-1, and to a lesser extent Ang-2, enhances TNF induced pro-inflammatory cytokine and chemokine expression. These results suggest that Tie2 signaling, in combination with TNF, induce a pro-inflammatory and pro-angiogenic profile in differentiated macrophages, and provide a molecular basis for the role of Tie2 in promoting disease progression in both early and established RA.
C. A. Ambarus,
B. Malvar Fernandez,
D. L. Baeten,
P. P. Tak,
Employee of GlaxoSmithKline,
K. A. Reedquist,
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/tie2-signalling-induces-a-pro-inflammatory-and-pro-angiogenic-phenotype-in-differentiated-macrophages-independently-of-macrophage-polarization-conditions-and-contributes-to-production-of-cytokines/