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Macrophages play a crucial role in all stages of cutaneous wound healing responses and dysregulation of macrophage function can result in derailed wound repair. The phenotype of macrophages is influenced by the wound microenvironment and evolves during healing from a more pro-inflammatory (M1) profile in early stages, to a less inflammatory pro-healing (M2) phenotype in later stages of repair. The aim of the current study was to investigate the potential of exogenous administration of M2 macrophages to promote wound healing in an experimental mouse model of cutaneous injury. Bone marrow derived macrophages were stimulated in-vitro with IL-4 or IL-10 to obtain two different subsets of M2-polarized cells, M2a or M2c respectively. Polarized macrophages were injected into full-thickness excisional skin wounds of either C57BL/6 or diabetic db/db mice. Control groups were injected with non-polarized (M0) macrophages or saline. Our data indicate that despite M2 macrophages exhibit an anti-inflammatory phenotype in-vitro, they do not improve wound closure in wild type mice while they delay healing in diabetic mice. Examination of wounds on day 15 post-injury indicated delayed re-epithelialization and persistence of neutrophils in M2 macrophage treated diabetic wounds. Therefore, topical application of ex-vivo generated M2 macrophages is not beneficial and contraindicated for cell therapy of skin wounds. [ABSTRACT FROM AUTHOR]

Abstract: The aim of the current study was to investigate the effect of nicotine in an experimental mouse model of cutaneous injury and healing responses, during the inflammatory phase of repair. Nicotine injection in full-thickness excisional skin wounds minimally affected inflammatory mediators like TNF, IL-6 and IL-12 while it induced a down-regulation in the expression of growth factors like VEGF, PDGF, TGF-β1 and TGF-β2, and the anti-inflammatory cytokine IL-10. Analysis of wound closure rate indicated no significant differences between nicotine and saline injected controls. In-vitro studies using bone marrow derived macrophages, resident peritoneal macrophages and RAW 264.7 macrophages, indicated that nicotine down-regulates TNF production. Moreover, nicotine was shown to down-regulate VEGF, PDGF and TGF-β1 in both bone marrow derived macrophages and RAW 264.7 cells. Using an NF-κB luciferase reporter RAW 264.7 cell line, we show that nicotine effects are minimally dependent on NF-κB inhibition. Moreover, nicotinic acetylcholine receptor (nAChR) subunit expression analyses indicated that while β2 nAChR subunit is expressed in mouse macrophages, α7 nAChR is not. In conclusion, while skin inflammatory parameters were not significantly affected by nicotine, a down-regulation of growth factor expression in both mouse skin and macrophages was observed. Reduced growth factor expression by nicotine might contribute, at least in part, to the overall detrimental effects of tobacco use in wound healing and skin diseases. [Copyright &y& Elsevier]

Activation of the transcription factor NF-&kgr;B appears to be involved in different stages of atherogenesis. In this paper we investigate the role of NF-&kgr;B inhibitor I&kgr;Bα in atherosclerosis. Myeloid-specific deletion of I&kgr;Bα results in larger and more advanced lesions in LDL-R-deficient mice without affecting the compositional phenotype of the plaques or systemic inflammatory markers in the plasma. We show that IkBa-deleted macrophages display enhanced adhesion to an in vitro endothelial cell layer, coinciding with an increased expression of the chemokine CCL5. Also, in vivo we found that IkBαdel mice had more leukocytes adhering to the luminal side of the endothelial cell layers that cover the atherosclerotic plaques. Moreover, we introduce ER-MP58 in this paper as a new immunohistochemical tool for quantifying newly recruited myeloid cells in the atherosclerotic lesion. This staining confirms that in IkBαdel mice more leukocytes are attracted to the plaques. In conclusion, we show that I&kgr;Bα deletion in myeloid cells promotes atherogenesis, probably through an induced leukocyte recruitment to plaques. [ABSTRACT FROM AUTHOR]

Background: The p38&agr; Mitogen-Activated Protein Kinase (MAPK) regulates stress- and inflammation-induced cellular responses. Factors implicated in the development of atherosclerosis including modified low-density lipoprotein (LDL), cytokines and even shear stress induce p38 activation in endothelial cells and macrophages, which may be important for plaque formation. This study investigates the effects of endothelial- and macrophage-specific deficiency of p38&agr; in atherosclerosis development, in Apolipoprotein E deficient (ApoE-/-) mice. Methodology/Principal Findings: ApoE-/- mice with macrophage or endothelial cell-specific p38&agr; deficiency were fed a high cholesterol diet (HCD) for 10 weeks and atherosclerosis development was assessed by histological and molecular methods. Surprisingly, although p38&agr;-deficiency strongly attenuated oxidized LDL-induced expression of molecules responsible for monocyte recruitment in endothelial cell cultures in vitro, endothelial-specific p38&agr; ablation in vivo did not affect atherosclerosis development. Similarly, macrophage specific deletion of p38&agr; did not affect atherosclerotic plaque development in ApoE-/- mice. Conclusions: Although previous studies implicated p38&agr; signaling in atherosclerosis, our in vivo experiments suggest that p38&agr; function in endothelial cells and macrophages does not play an important role in atherosclerotic plaque formation in ApoE deficient mice. [ABSTRACT FROM AUTHOR]

Background: The aim of the current study was to investigate the role of p55 TNF Receptor (p55 TNFR), the main signaling receptor for the pro-inflammatory cytokine tumor necrosis factor (TNF), in the development of two vascular disorders: atherosclerosis and angiotensin (Ang) II-induced abdominal aortic aneurysms (AAA). Methodology/Principal Findings: p55 TNFR deficient mice were crossed to an LDL receptor deficient background and were induced for the development of either atherosclerosis or AngII-induced AAA, and compared to littermate controls, wild-type for p55 TNFR expression. p55 TNFR deficient mice developed 43% smaller atherosclerotic lesions in the aortic sinuses compared to controls. Moreover, expression of CD68, a macrophage specific marker, exhibited a 50% reduction in the aortic arches. Decreased atherosclerosis correlated with a strong down-regulation in the expression of adhesion molecules, such as VCAM-1 and ICAM-1, by p55 TNFR deficient endothelium. In addition, expression levels of the pro-inflammatory cytokines and chemokines TNF, IL-6, MCP-1 and RANTES were significantly reduced in aortas of p55 TNFR deficient mice. In contrast, in the AngII-induced model of AAA, p55 TNFR deficiency correlated with a slight trend towards increased aneurismal lethality, but the incidence of aortic rupture due to a dissecting aneurysm, and the expansion of the suprarenal aorta were not significantly different compared to controls. Conclusion/Significance: We found that p55 TNFR expression promotes atherosclerosis, among other mechanisms, by enhancing expression of endothelial adhesion molecules, while it seems to have no major role in the development of AngIIinduced AAA. [ABSTRACT FROM AUTHOR]

Summary: Atherosclerosis is a progressive disorder of the arterial wall and the underlying cause of cardiovascular diseases such as heart attack and stroke. Today, atherosclerosis is recognized as a complex disease with a strong inflammatory component. The nuclear factor-κB (NF-κB) signaling pathway regulates inflammatory responses and has been implicated in atherosclerosis. Here, we addressed the function of NF-κB signaling in vascular endothelial cells in the pathogenesis of atherosclerosis in vivo. Endothelium-restricted inhibition of NF-κB activation, achieved by ablation of NEMO/IKKγ or expression of dominant-negative IκBα specifically in endothelial cells, resulted in strongly reduced atherosclerotic plaque formation in ApoE −/− mice fed with a cholesterol-rich diet. Inhibition of NF-κB abrogated adhesion molecule induction in endothelial cells, impaired macrophage recruitment to atherosclerotic plaques, and reduced expression of cytokines and chemokines in the aorta. Thus, endothelial NF-κB signaling orchestrates proinflammatory gene expression at the arterial wall and promotes the pathogenesis of atherosclerosis. [Copyright &y& Elsevier]

Background—Previous studies implicated Toll-like receptor signaling as a critical pathogenic pathway in atherosclerosis, but the cell-specific mechanisms by which Toll-like receptors act to control atherosclerotic plaque development remain poorly understood. Methods and Results—To study the cell-specific role of tumor necrosis factor receptor-associated factor 6 (TRAF6) in atherosclerosis, we generated ApoE-/- mice with endothelial cell- or myeloid cell-specific TRAF6 deficiency using Cre/LoxP-mediated gene targeting. Endothelial TRAF6 deficiency reduced atherosclerosis in female ApoE-/- mice by inhibiting nuclear factor-κB-dependent proinflammatory gene expression and monocyte adhesion to endothelial cells. In contrast, myeloid cell-specific TRAF6 deficiency caused exacerbated atherosclerosis, with larger plaques containing more necrotic areas in both male and female ApoE-/- mice. TRAF6-deficient macrophages showed impaired expression of the antiinflammatory and atheroprotective cytokine interleukin-10, elevated endoplasmic reticulum stress, increased sensitivity to oxidized low-density lipoprotein-induced apoptosis, and reduced capacity to clear apoptotic cells. Thus, the reduced antiinflammatory properties, coupled with increased sensitivity to apoptosis and impaired efferocytosis capacity of TRAF6-deficient macrophages, result in exacerbated atherosclerosis development in TR AF6MYKO/ApoE-/- mice. Conclusion—Toll-like receptor-mediated TRAF6 signaling acts in endothelial cells to promote atherosclerosis but displays atheroprotective, antiinflammatory and prosurvival functions in myeloid cells. [ABSTRACT FROM AUTHOR]