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CCL5–CCR5 axis mediates skin wound healing by promoting endothelial progenitor cell accumulation
Abstracts / Cytokine 48 (2009) 91–137 Secondary necrosis of apoptotic neutrophils induced by the human cathelicidin LL-37 is not proinflammatory to phagocytosing macrophages Hsin-Ni Li 1, Peter G. Barlow 1, Johan Bylund 2, Annie Mackellar 1, Åse Björstad 2, James Conlon 1, Pieter S. Hiemstra 3, Chris Haslett 1, Mohini Gray 1, A. John Simpson 1, Adriano G. Rossi 1, Donald J. Davidson 1, 1 MRC/ University of Edinburgh Centre for Inflammation Research, Queen’s Medical Research Institute, 47 Little France Crescent, Edinburgh EH16 4TJ, Scotland, 2 Department of Rheumatology, University of Gothenburg, Sweden, 3 Department of Pulmonology, Leiden University Medical Center, The Netherlands Cathelicidins are cationic host defence peptides (CHDP) with microbicidal, antiendotoxic, and multiple immunomodulatory properties. The importance of these peptides to innate host defence is indicated by the increased susceptibility to infection of individuals with morbus Kostmann and cathelicidin deficient mice, and the associations between cathelicidin levels and the pathogenesis of certain chronic diseases. Neutrophils (PMN) are the main reservoir of cathelicidins and play key roles in first line defence against infection. The appropriate regulation of PMN function, death, and clearance is critical to innate immunity and the efferocytosis of apoptotic PMN, in contrast to necrotic cells, is proposed to promote the resolution of inflammation. We demonstrate that the human cathelicidin LL-37 rapidly induced secondary necrosis of apoptotic human PMNs and identify the essential C-terminal region of LL-37 required this activity. LL-37-induced secondary necrosis did not affect PMN ingestion by human monocyte-derived macrophages and, in contrast to expectation, was not proinflammatory. The anti-inflammatory effects of apoptotic PMN on activated macrophages were retained and even potentiated in LL-37-induced secondarily necrotic PMN. In addition to the induction of secondary necrosis, LL-37 also promoted PMN granule contents release. Thus, we suggest that LL-37 induces swift secondary necrosis of PMN during inflammation without promoting macrophage inflammation, but may mediate host damage by releasing potentially harmful granules under chronic or dysregulated conditions.
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Molecular Preventive Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan, 4 Division of Molecular Bioregulation, Cancer Research Institute, Kanazawa University, Kanazawa, Japan Endothelial progenitor cells (EPCs) are a circulating, bone marrow-derived cell population that appears to participate in vasculogenesis and tissue repair. However, the mechanisms by which EPCs home to remodeling tissues still remain unclear. Here, we show that CC chemokine receptor 5 (CCR5) is crucial for the homing of EPCs to skin wound in mice. We found that EPCs, characterized by CD34+/Flk-1+ cells, expressed CCR5 protein and existed in peripheral blood and skin tissue after wounding in wild-type (WT) mice. Although EPCs were mobilized from bone marrow into circulation in both WT and CCR5 / mice to a similar extent after wounding, the EPC homing to the wounds was significantly decreased in CCR5 / mice compared with that in WT mice. Consistent with this, CCR5 / mice exhibited attenuated neovascularization at the wound sites, which appeared to result in an impaired wound healing as evidenced by delayed wound closure and diminished granulation tissue formation. Transfer of BM cells from WT, but not from CCR5 / donor mice, restored wound healing along with augmented neovascularization in CCR5 / recipient mice. Moreover, injected EPCs migrated to the wounds and were incorporated into capillaries after systemic administration with wounding. We also confirmed that CC chemokine ligand CCL5, but not CCL3 and CCL4, induced migration of ex vivo expanded EPCs in a dose-dependent manner in vitro. Together, our data suggests that CCL5–CCR5 interaction are essential for skin wound healing through the homing of EPCs to wound. doi:10.1016/j.cyto.2009.07.486
doi:10.1016/j.cyto.2009.07.484
PP2-109 Using luminex (xMAP) technology to assay for inflammatory cytokines in cell culture supernatants from daudi and ovcar-3 cells treated with interferon
PP2-107 Absence of IFN-c accelerates thrombus resolution through enhanced MMP-9 and VEGF expression
Joseph Bekisz, Hana Schmeisser, David Stephany, Kathryn Zoon, Poster Presentation II Using luminex (xMAP) technology to assay for inflammatory cytokines in cell culture supernatants from daudi and ovcar-3 cells treated with interferon Joseph Bekisz, Hana Schmeisser, David Stephany, Kathryn Zoon, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, USA
Toshikazu Kondo, Mizuho Nosaka, Yuko Ishida, Akihiko Kimura, Yumi Kuninaka, Masanori Inui, Naofumi Mukaida, Poster Presentation II Absence of IFN-gamma accelerates thrombus resolution through enhanced MMP9 and VEGF expression Toshikazu Kondo 1, Mizuho Nosaka 1, Yuko Ishida 1, Akihiko Kimura 1, Yumi Kuninaka 1, Masanori Inui 2, Naofumi Mukaida 1,2, 1 Department of Forensic Medicine, Wakayama Medical University, Wakayama, Japan, 2 Department of Immunology, Institute of Advanced Medicine We explored the pathophysiological roles of IFN-c in the resolution of venous thrombi. Upon the ligation of the inferior vena cava (IVC), venous thrombi developed progressively until 5 days, and remained similar sizes at 10 days. Concomitantly, intrathrombotic IFN-c contents were elevated progressively with an increase of post-ligation intervals. When IFN-c -deficient (IFN-c = ) mice were treated in the same manner, thrombus size was similar to that in wild-type (WT) mice until 5 days after the IVC ligation, but it was apparently smaller at 10 and 14 days, compared with WT mice. Intrathrombotic collagen-positive areas and intrathrombotic hydroxyproline contents were remarkably reduced later than 10 days after IVC ligation in IFNc = mice, compared with WT mice. However, there were no significant differences in intrathrombotic chemokine expression and leukocyte recruitment between both strains. Matrix metalloproteinase (MMP)-9 but not MMP-2 mRNA expression was higher at the late phase in IFN-c = mice, than WT mice. Concomitantly, MMP-9 enzyme activities were higher in IFN-c = mice, than WT mice. Moreover, intrathrombotic vascular areas were increased in IFN-c = mice, together with enhanced vascular endothelial growth factor (VEGF) gene expression, compared with WT mice. Furthermore, the administration of anti-IFN-c mAb accelerated the thrombus resolution in WT mice. Taken together, IFN-c can have a detrimental role in the thrombus resolution by suppressing MMP-9 and VEGF expression and can be a good molecular target for the treatment of deep vein thrombosis.
One goal of our research is to develop new interferon alpha constructs which will exhibit equal to or greater antiviral and antiproliferative activities but reduced side effects many of which may be related to inflammatory cytokines induced by the IFN treatment. Hybrid interferons composed of IFN-alpha 2c and IFN-alpha 21b were examined on Daudi (Burkitt’s Lymphoma) and OVCAR-3 (Ovarian carcinoma) cells for their antiviral and antiproliferative activities as well as the expression profiles of inflammatory cytokines and chemokines. Luminex (xMAP) technology was employed with this analysis which allows one to probe for a variety of analytes in a single sample and to quantitate the amount of that analyte in pg/ml. Three timepoints (24, 48 and 72 h) were examined at which cells were either left untreated or treated with IFN-a 2c (5 ng/ml). Cells culture supernatants were used for analysis. Preliminary results showed expected levels of IFN-alpha 2 given the amount of IFN-alpha used for treatment. Enhanced levels of IL-1 alpha were detected in both OVCAR-3 and Daudi samples, whereas IL-6 and IL-8 were upregulated in OVCAR-3 cells only. All three of these cytokines are associated with inflammation. Interestingly, the two macrophage inflammatory proteins (MIP-1a and MIP-1b) were upregulated in Daudi cells but not detected at all in IFN-treated OVCAR cells. TNF-a and TNF-b levels in both cell lines were essentially non-detectable. By far, the analyte expressed to the greatest extent in both cell lines was IP-10, a chemokine known to be induced by IFN-c and recently shown to be induced differentially by some IFN-a subtypes. Future work will include treating various cell lines with the IFN-a hybrids generated in our laboratory so that both the cell- specific and IFN-specific expression profiles can be determined and correlated with antiviral and antiproliferative activities and their ability to induce inflammatory cytokines and chemokines.
doi:10.1016/j.cyto.2009.07.485
doi:10.1016/j.cyto.2009.07.487
PP2-108 CCL5–CCR5 axis mediates skin wound healing by promoting endothelial progenitor cell accumulation
PP2-110 Prostaglandin E2 (PGE2) induces IL-6 in human orbital fibroblasts through a CREB-dependent mechanism
Yuko Ishida, Akihiko Kimura, Masanori Inui, Yumi Kuninaka, Kouji Matsushima, Naofumi Mukaida, Toshikazu Kondo, Poster Presentation II CCL5–CCR5 axis mediates skin wound healing by promoting endothelial progenitor cell accumulation Yuko Ishida 1, Akihiko Kimura 1, Masanori Inui 2, Yumi Kuninaka 1, Kouji Matsushima 3, Naofumi Mukaida 4, Toshikazu Kondo 1, 1 Department of Forensic Medicine, Wakayama Medical University, Wakayama, Japan, 2 Department of Molecular Immunology, Institute of Advanced Medicine, Wakayama Medical University, Wakayama, Japan, 3 Department of
Nupur Raychaudhuri, Terry J. Smith, Poster Presentation II Prostaglandin E2 (PGE2) induces IL-6 in human orbital fibroblasts through a CREBdependent mechanism Nupur Raychaudhuri, Terry J. Smith, Kellogg Eye Center, University of Michigan, Ann Arbor, MI, USA Background: IL-6 has been implicated in the pathogenesis of Graves’ disease and its inflammatory orbital manifestation, thyroid-associated ophthalmopathy (TAO).
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Molecular Preventive Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan, 4 Division of Molecular Bioregulation, Cancer Research Institute, Kanazawa University, Kanazawa, Japan Endothelial progenitor cells (EPCs) are a circulating, bone marrow-derived cell population that appears to participate in vasculogenesis and tissue repair. However, the mechanisms by which EPCs home to remodeling tissues still remain unclear. Here, we show that CC chemokine receptor 5 (CCR5) is crucial for the homing of EPCs to skin wound in mice. We found that EPCs, characterized by CD34+/Flk-1+ cells, expressed CCR5 protein and existed in peripheral blood and skin tissue after wounding in wild-type (WT) mice. Although EPCs were mobilized from bone marrow into circulation in both WT and CCR5 / mice to a similar extent after wounding, the EPC homing to the wounds was significantly decreased in CCR5 / mice compared with that in WT mice. Consistent with this, CCR5 / mice exhibited attenuated neovascularization at the wound sites, which appeared to result in an impaired wound healing as evidenced by delayed wound closure and diminished granulation tissue formation. Transfer of BM cells from WT, but not from CCR5 / donor mice, restored wound healing along with augmented neovascularization in CCR5 / recipient mice. Moreover, injected EPCs migrated to the wounds and were incorporated into capillaries after systemic administration with wounding. We also confirmed that CC chemokine ligand CCL5, but not CCL3 and CCL4, induced migration of ex vivo expanded EPCs in a dose-dependent manner in vitro. Together, our data suggests that CCL5–CCR5 interaction are essential for skin wound healing through the homing of EPCs to wound. doi:10.1016/j.cyto.2009.07.486
doi:10.1016/j.cyto.2009.07.484
PP2-109 Using luminex (xMAP) technology to assay for inflammatory cytokines in cell culture supernatants from daudi and ovcar-3 cells treated with interferon
PP2-107 Absence of IFN-c accelerates thrombus resolution through enhanced MMP-9 and VEGF expression
Joseph Bekisz, Hana Schmeisser, David Stephany, Kathryn Zoon, Poster Presentation II Using luminex (xMAP) technology to assay for inflammatory cytokines in cell culture supernatants from daudi and ovcar-3 cells treated with interferon Joseph Bekisz, Hana Schmeisser, David Stephany, Kathryn Zoon, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, USA
Toshikazu Kondo, Mizuho Nosaka, Yuko Ishida, Akihiko Kimura, Yumi Kuninaka, Masanori Inui, Naofumi Mukaida, Poster Presentation II Absence of IFN-gamma accelerates thrombus resolution through enhanced MMP9 and VEGF expression Toshikazu Kondo 1, Mizuho Nosaka 1, Yuko Ishida 1, Akihiko Kimura 1, Yumi Kuninaka 1, Masanori Inui 2, Naofumi Mukaida 1,2, 1 Department of Forensic Medicine, Wakayama Medical University, Wakayama, Japan, 2 Department of Immunology, Institute of Advanced Medicine We explored the pathophysiological roles of IFN-c in the resolution of venous thrombi. Upon the ligation of the inferior vena cava (IVC), venous thrombi developed progressively until 5 days, and remained similar sizes at 10 days. Concomitantly, intrathrombotic IFN-c contents were elevated progressively with an increase of post-ligation intervals. When IFN-c -deficient (IFN-c = ) mice were treated in the same manner, thrombus size was similar to that in wild-type (WT) mice until 5 days after the IVC ligation, but it was apparently smaller at 10 and 14 days, compared with WT mice. Intrathrombotic collagen-positive areas and intrathrombotic hydroxyproline contents were remarkably reduced later than 10 days after IVC ligation in IFNc = mice, compared with WT mice. However, there were no significant differences in intrathrombotic chemokine expression and leukocyte recruitment between both strains. Matrix metalloproteinase (MMP)-9 but not MMP-2 mRNA expression was higher at the late phase in IFN-c = mice, than WT mice. Concomitantly, MMP-9 enzyme activities were higher in IFN-c = mice, than WT mice. Moreover, intrathrombotic vascular areas were increased in IFN-c = mice, together with enhanced vascular endothelial growth factor (VEGF) gene expression, compared with WT mice. Furthermore, the administration of anti-IFN-c mAb accelerated the thrombus resolution in WT mice. Taken together, IFN-c can have a detrimental role in the thrombus resolution by suppressing MMP-9 and VEGF expression and can be a good molecular target for the treatment of deep vein thrombosis.
One goal of our research is to develop new interferon alpha constructs which will exhibit equal to or greater antiviral and antiproliferative activities but reduced side effects many of which may be related to inflammatory cytokines induced by the IFN treatment. Hybrid interferons composed of IFN-alpha 2c and IFN-alpha 21b were examined on Daudi (Burkitt’s Lymphoma) and OVCAR-3 (Ovarian carcinoma) cells for their antiviral and antiproliferative activities as well as the expression profiles of inflammatory cytokines and chemokines. Luminex (xMAP) technology was employed with this analysis which allows one to probe for a variety of analytes in a single sample and to quantitate the amount of that analyte in pg/ml. Three timepoints (24, 48 and 72 h) were examined at which cells were either left untreated or treated with IFN-a 2c (5 ng/ml). Cells culture supernatants were used for analysis. Preliminary results showed expected levels of IFN-alpha 2 given the amount of IFN-alpha used for treatment. Enhanced levels of IL-1 alpha were detected in both OVCAR-3 and Daudi samples, whereas IL-6 and IL-8 were upregulated in OVCAR-3 cells only. All three of these cytokines are associated with inflammation. Interestingly, the two macrophage inflammatory proteins (MIP-1a and MIP-1b) were upregulated in Daudi cells but not detected at all in IFN-treated OVCAR cells. TNF-a and TNF-b levels in both cell lines were essentially non-detectable. By far, the analyte expressed to the greatest extent in both cell lines was IP-10, a chemokine known to be induced by IFN-c and recently shown to be induced differentially by some IFN-a subtypes. Future work will include treating various cell lines with the IFN-a hybrids generated in our laboratory so that both the cell- specific and IFN-specific expression profiles can be determined and correlated with antiviral and antiproliferative activities and their ability to induce inflammatory cytokines and chemokines.
doi:10.1016/j.cyto.2009.07.485
doi:10.1016/j.cyto.2009.07.487
PP2-108 CCL5–CCR5 axis mediates skin wound healing by promoting endothelial progenitor cell accumulation
PP2-110 Prostaglandin E2 (PGE2) induces IL-6 in human orbital fibroblasts through a CREB-dependent mechanism
Yuko Ishida, Akihiko Kimura, Masanori Inui, Yumi Kuninaka, Kouji Matsushima, Naofumi Mukaida, Toshikazu Kondo, Poster Presentation II CCL5–CCR5 axis mediates skin wound healing by promoting endothelial progenitor cell accumulation Yuko Ishida 1, Akihiko Kimura 1, Masanori Inui 2, Yumi Kuninaka 1, Kouji Matsushima 3, Naofumi Mukaida 4, Toshikazu Kondo 1, 1 Department of Forensic Medicine, Wakayama Medical University, Wakayama, Japan, 2 Department of Molecular Immunology, Institute of Advanced Medicine, Wakayama Medical University, Wakayama, Japan, 3 Department of
Nupur Raychaudhuri, Terry J. Smith, Poster Presentation II Prostaglandin E2 (PGE2) induces IL-6 in human orbital fibroblasts through a CREBdependent mechanism Nupur Raychaudhuri, Terry J. Smith, Kellogg Eye Center, University of Michigan, Ann Arbor, MI, USA Background: IL-6 has been implicated in the pathogenesis of Graves’ disease and its inflammatory orbital manifestation, thyroid-associated ophthalmopathy (TAO).