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Cartwright IM, Colgan SP. The hypoxic tissue microenvironment as a driver of mucosal inflammatory resolution. Front Immunol 2023; 14:1124774. [PMID: 36742292 PMCID: PMC9890178 DOI: 10.3389/fimmu.2023.1124774] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/15/2022] [Accepted: 01/06/2023] [Indexed: 01/19/2023] Open
Abstract
On the backdrop of all acute inflammatory processes lies the activation of the resolution response. Recent years have witnessed an emerging interest in defining molecular factors that influence the resolution of inflammation. A keystone feature of the mucosal inflammatory microenvironment is hypoxia. The gastrointestinal tract, particularly the colon, exists in a state of physiological hypoxia and during active inflammation, this hypoxic state is enhanced as a result of infiltrating leukocyte oxygen consumption and the activation of oxygen consuming enzymes. Most evidence suggests that mucosal hypoxia promotes the active resolution of inflammation through a variety of mechanisms, including extracellular acidification, purine biosynthesis/salvage, the generation of specialized pro-resolving lipid mediators (ie. resolvins) and altered chemokine/cytokine expression. It is now appreciated that infiltrating innate immune cells (neutrophils, eosinophils, macrophages) have an important role in molding the tissue microenvironment to program an active resolution response. Structural or functional dysregulation of this inflammatory microenvironment can result in the loss of tissue homeostasis and ultimately progression toward chronicity. In this review, we will discuss how inflammatory hypoxia drives mucosal inflammatory resolution and its impact on other microenvironmental factors that influence resolution.
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Affiliation(s)
- Ian M. Cartwright
- Division of Gastroenterology and Hepatology, University of Colorado Anschutz Medical Campus, Aurora, CO, United States
- Department of Medicine, University of Colorado School of Medicine, Aurora, CO, United States
- Mucosal Inflammation Program, University of Colorado School of Medicine, Aurora, CO, United States
- Rocky Mountain Regional Veterans Affairs Medical Center, Aurora, CO, United States
| | - Sean P. Colgan
- Division of Gastroenterology and Hepatology, University of Colorado Anschutz Medical Campus, Aurora, CO, United States
- Department of Medicine, University of Colorado School of Medicine, Aurora, CO, United States
- Mucosal Inflammation Program, University of Colorado School of Medicine, Aurora, CO, United States
- Rocky Mountain Regional Veterans Affairs Medical Center, Aurora, CO, United States
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Gooneratne TD, Homer-Vanniasinkam S, Wijeyaratne SM. Beneficial Effects of Insulin on Ischemia Reperfusion Injury in Human Skeletal Muscle. Vasc Specialist Int 2022; 38:28. [PMID: 36216364 PMCID: PMC9550711 DOI: 10.5758/vsi.220018] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/19/2022] [Revised: 06/28/2022] [Accepted: 09/08/2022] [Indexed: 11/26/2022] Open
Abstract
Purpose Exaggerated leucocyte activity is a crucial step in the pathophysiology of skeletal muscle ischemia-reperfusion injury (IRI). We tested the hypothesis that insulin, via its' anti-leukocyte activity, attenuates skeletal muscle IRI in humans. Materials and Methods This randomized, blinded, placebo-controlled trial was conducted in patients with skeletal muscle ischemia who required revascularization. Treatment protocols were similar among them except for the insulin group, which received an infusion of insulin at 2.5 U/h. The degree of endothelial adhesiveness; leukocyte activity and pro-inflammatory status via P-selectin, tumor necrosis factor (TNF)-alpha, and myeloperoxidase (MPO) levels in the venous effluent; and clinical outcomes were measured. Results Twenty-four consenting patients were randomized to the insulin or control group. There were no significant differences between the two groups except for the median serum insulin level, which was higher in the insulin group (P<0.01). No serious intervention-related adverse events were observed. P-selectin (55.04-99.86 pg/mL; P<0.001), MPO (110.8-160.6 pg/mL; P<0.001), and TNF-alpha (12.16-36.01 pg/mL; P<0.001) levels demonstrated a significant increase post-reperfusion in the 'control' group, reaching a peak value at 2 hours post-reperfusion. The increase in all three markers from baseline was significantly diminished in the insulin group at the two-hour (P-selectin, P=0.001; MPO, P=0.001; TNF-alpha, P=0.005) and four-hour (P-selectin, P=0.003; MPO, P=0.002; TNF-alpha, P=0.01) intervals. The differences in clinical outcomes between the insulin and control groups were not statistically significant. Conclusion In clinical practice, insulin has the potential to attenuate the severity of skeletal muscle IRI inhibiting P-selectin, MPO, and TNF-alpha levels.
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Chen T, Niu L, Wang L, Zhou Q, Zhao X, Lai S, He X, He H, He M. Ferulic acid protects renal tubular epithelial cells against anoxia/reoxygenation injury mediated by AMPKα1. Free Radic Res 2022; 56:173-184. [PMID: 35382666 DOI: 10.1080/10715762.2022.2062339] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/04/2022]
Abstract
Anoxia/reoxygenation (A/R) injury causes dysfunction of rat renal tubular epithelial cells (NRK-52E), which is associated with excess reactive oxygen species (ROS) generation and eventually leads to apoptosis. Ferulic acid (FA), a phenolic acid, which is abundant in fruits and vegetables. FA possesses the properties of scavenging free radicals and cytoprotection against oxygen stress. In the study, the protective effects of FA against NRK-52E cells damage induced by A/R were explored and confirmed the role of AMP-activated protein kinaseα1 (AMPKα1). We found that after NRK-52E cells suffered A/R damage, FA pretreatment increased the cell viability and decreased LDH activity in culture medium in a concentration-dependent manner, the activities of endogenous antioxidant enzymes such as glutathione peroxidase, superoxide dismutase and catalase improved, intracellular ROS generation and malondialdehyde contents mitigated. In addition, pretreatment of 75 μM FA ameliorated mitochondrial dysfunction by A/R-injury and ultimately decreased apoptosis (25.3 ± 0.61 vs 12.1 ± 0.60), which was evidenced by preventing the release of cytochrome c from mitochondria to the cytoplasm. 75 μM FA pretreatment also significantly upregulated AMPKα1 expression (3.16 ± 0.18 folds) and phosphorylation (2.56 ± 0.13 folds). However, compound C, a specific AMPK inhibitor, significantly attenuated FA pretreatment's effects, as mentionedabove. These results firstly clarified that FA pretreatment attenuated NRK-52E cell damage induced by A/R via upregulating AMPKα1 expression and phosphorylation.
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Affiliation(s)
- Tianpeng Chen
- Institute of Cardiovascular Diseases, Jiangxi Academy of Clinical Medical Sciences, the First Affiliated Hospital of Nanchang University, Nanchang 330006, China
| | - Li Niu
- Jiangxi Provincial Key Laboratory of Basic Pharmacology, Nanchang University School of Pharmaceutical Science, Nanchang 330006, China
| | - Liang Wang
- Department of rehabilitation, the First Affiliated Hospital of Nanchang University, Nanchang 330006, China
| | - Qing Zhou
- Jiangxi Provincial Key Laboratory of Basic Pharmacology, Nanchang University School of Pharmaceutical Science, Nanchang 330006, China
| | - Xiaoyu Zhao
- Jiangxi Provincial Key Laboratory of Basic Pharmacology, Nanchang University School of Pharmaceutical Science, Nanchang 330006, China
| | - Songqing Lai
- Institute of Cardiovascular Diseases, Jiangxi Academy of Clinical Medical Sciences, the First Affiliated Hospital of Nanchang University, Nanchang 330006, China
| | - Xinlan He
- Jiangxi Provincial Key Laboratory of Basic Pharmacology, Nanchang University School of Pharmaceutical Science, Nanchang 330006, China
| | - Huan He
- Jiangxi Provincial Key Laboratory of Basic Pharmacology, Nanchang University School of Pharmaceutical Science, Nanchang 330006, China
| | - Ming He
- Institute of Cardiovascular Diseases, Jiangxi Academy of Clinical Medical Sciences, the First Affiliated Hospital of Nanchang University, Nanchang 330006, China
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De Galan C, De Vos M, Hindryckx P, Laukens D, Van Welden S. Long-Term Environmental Hypoxia Exposure and Haematopoietic Prolyl Hydroxylase-1 Deletion Do Not Impact Experimental Crohn's Like Ileitis. BIOLOGY 2021; 10:biology10090887. [PMID: 34571764 PMCID: PMC8464968 DOI: 10.3390/biology10090887] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 08/05/2021] [Revised: 08/23/2021] [Accepted: 09/03/2021] [Indexed: 11/16/2022]
Abstract
Simple Summary Hypoxia-induced signalling represents an important contributor to inflammatory bowel disease (IBD) pathophysiology. However, available data solely focus on colonic inflammation while the primary disease location in Crohn’s disease patients is the terminal ileum. Therefore, we explored the effects of environmental hypoxia and immune cell-specific deletion of oxygen sensor prolyl hydroxylase (PHD) 1 in a Crohn’s like ileitis mouse model. Five-week-old TNF∆ARE/+ mice and wildtype (WT) littermates were housed in normoxia (21% O2) or hypoxia (8% O2) for 10 weeks. Although environmental hypoxia increased both systemic as ileal markers of hypoxia, the body weight evolution in both WT and TNF∆ARE/+ mice was not affected. Interestingly, hypoxia did increase circulatory monocytes, ileal mononuclear phagocytes and proinflammatory cytokine expression in WT mice. However, no histological or inflammatory gene expression differences in the ileum could be identified between TNF∆ARE/+ mice housed in hypoxia versus normoxia nor between TNF∆ARE/+ and WT mice with additional loss of immune cell-specific Phd1 expression. This is the first study showing that long-term environmental hypoxia or haematopoietic Phd1-deletion does not impact experimental ileitis. Therefore, it strongly questions whether targeting hypoxia-induced signalling via currently available PHD inhibitors would exert an immune suppressive effect in IBD patients with ileal inflammation. Abstract Environmental hypoxia and hypoxia-induced signalling in the gut influence inflammatory bowel disease pathogenesis, however data is limited to colitis. Hence, we investigated the effect of environmental hypoxia and immune cell-specific deletion of oxygen sensor prolyl hydroxylase (PHD) 1 in a Crohn’s like ileitis mouse model. Therefore, 5-week-old C57/BL6 TNF∆ARE/+ mice and wildtype (WT) littermates were housed in normoxia (21% O2) or hypoxia (8% O2) for 10 weeks. Systemic inflammation was assessed by haematology. Distal ileal hypoxia was evaluated by pimonidazole staining. The ileitis degree was scored on histology, characterized via qPCR and validated in haematopoietic Phd1-deficient TNF∆ARE/+ mice. Our results demonstrated that hypoxia did not impact body weight evolution in WT and TNF∆ARE/+ mice. Hypoxia increased red blood cell count, haemoglobin, haematocrit and increased pimonidazole intensity in the ileum. Interestingly, hypoxia evoked an increase in circulatory monocytes, ileal mononuclear phagocytes and proinflammatory cytokine expression in WT mice. Despite these alterations, no histological or ileal gene expression differences could be identified between TNF∆ARE/+ mice housed in hypoxia versus normoxia nor between haematopoietic Phd1-deficient TNF∆ARE/+ and their WT counterparts. Therefore, we demonstrated for the first time that long-term environmental hypoxia or haematopoietic Phd1-deletion does not impact experimental ileitis development.
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Affiliation(s)
- Cara De Galan
- Department of Internal Medicine and Paediatrics, Ghent University, 9000 Ghent, Belgium; (C.D.G.); (M.D.V.); (P.H.); (D.L.)
- Ghent Gut Inflammation Group (GGIG), Ghent University, 9000 Ghent, Belgium
- VIB Centre for Inflammation Research, 9000 Ghent, Belgium
| | - Martine De Vos
- Department of Internal Medicine and Paediatrics, Ghent University, 9000 Ghent, Belgium; (C.D.G.); (M.D.V.); (P.H.); (D.L.)
- Ghent Gut Inflammation Group (GGIG), Ghent University, 9000 Ghent, Belgium
| | - Pieter Hindryckx
- Department of Internal Medicine and Paediatrics, Ghent University, 9000 Ghent, Belgium; (C.D.G.); (M.D.V.); (P.H.); (D.L.)
- Department of Gastroenterology, Ghent University Hospital, 9000 Ghent, Belgium
| | - Debby Laukens
- Department of Internal Medicine and Paediatrics, Ghent University, 9000 Ghent, Belgium; (C.D.G.); (M.D.V.); (P.H.); (D.L.)
- Ghent Gut Inflammation Group (GGIG), Ghent University, 9000 Ghent, Belgium
- VIB Centre for Inflammation Research, 9000 Ghent, Belgium
| | - Sophie Van Welden
- Department of Internal Medicine and Paediatrics, Ghent University, 9000 Ghent, Belgium; (C.D.G.); (M.D.V.); (P.H.); (D.L.)
- Ghent Gut Inflammation Group (GGIG), Ghent University, 9000 Ghent, Belgium
- VIB Centre for Inflammation Research, 9000 Ghent, Belgium
- Correspondence: ; Tel.: +32-9-332-58-30
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Mucosal acidosis elicits a unique molecular signature in epithelia and intestinal tissue mediated by GPR31-induced CREB phosphorylation. Proc Natl Acad Sci U S A 2021; 118:2023871118. [PMID: 33972436 DOI: 10.1073/pnas.2023871118] [Citation(s) in RCA: 16] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022] Open
Abstract
Metabolic changes associated with tissue inflammation result in significant extracellular acidosis (EA). Within mucosal tissues, intestinal epithelial cells (IEC) have evolved adaptive strategies to cope with EA through the up-regulation of SLC26A3 to promote pH homeostasis. We hypothesized that EA significantly alters IEC gene expression as an adaptive mechanism to counteract inflammation. Using an unbiased RNA sequencing approach, we defined the impact of EA on IEC gene expression to define molecular mechanisms by which IEC respond to EA. This approach identified a unique gene signature enriched in cyclic AMP response element-binding protein (CREB)-regulated gene targets. Utilizing loss- and gain-of-function approaches in cultured epithelia and murine colonoids, we demonstrate that EA elicits prominent CREB phosphorylation through cyclic AMP-independent mechanisms that requires elements of the mitogen-activated protein kinase signaling pathway. Further analysis revealed that EA signals through the G protein-coupled receptor GPR31 to promote induction of FosB, NR4A1, and DUSP1. These studies were extended to an in vivo murine model in conjunction with colonization of a pH reporter Escherichia coli strain that demonstrated significant mucosal acidification in the TNFΔARE model of murine ileitis. Herein, we observed a strong correlation between the expression of acidosis-associated genes with bacterial reporter sfGFP intensity in the distal ileum. Finally, the expression of this unique EA-associated gene signature was increased during active inflammation in patients with Crohn's disease but not in the patient control samples. These findings establish a mechanism for EA-induced signals during inflammation-associated acidosis in both murine and human ileitis.
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Pantalone JM, Liu S, Olaloye OO, Prochaska EC, Yanowitz T, Riley MM, Buland JR, Brozanski BS, Good M, Konnikova L. Gestational Age-Specific Complete Blood Count Signatures in Necrotizing Enterocolitis. Front Pediatr 2021; 9:604899. [PMID: 33718296 PMCID: PMC7952609 DOI: 10.3389/fped.2021.604899] [Citation(s) in RCA: 12] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/10/2020] [Accepted: 01/29/2021] [Indexed: 12/18/2022] Open
Abstract
Objective: Necrotizing enterocolitis (NEC) is characterized by peripheral cell abnormalities, yet few studies have analyzed the complete blood count (CBC) specifically by gestational age (GA). Our objective was to describe GA-specific immune abnormalities in NEC through a comprehensive analysis of the CBC differential. Methods: Using a cohort of 246 infants (177 cases, 69 controls) admitted to neonatal intensive care units at a single institution, we retrospectively analyzed CBCs around illness onset in NEC cases compared with controls. Cases included surgical NEC (S-NEC, 34.5%) and medical NEC (M-NEC, 65.5%). Infants were divided into those born at GA <33 and ≥33 weeks. Differences in CBC values were described as absolute and percent changes at NEC onset from baseline and at antibiotic completion after NEC. We used machine learning algorithms based on the CBC at NEC to generate predictive models for diagnosis. Results: At NEC onset, there was an acute drop in monocytes and lymphocytes along with a rise in bands in S-NEC infants born <33 weeks compared with M-NEC. In comparison, both M-NEC and S-NEC ≥33 weeks had a percent drop in neutrophils at diagnosis compared with controls. At antibiotic completion, monocytes in S-NEC <33 weeks significantly rose compared with M-NEC, yet for S-NEC ≥33 weeks, bands significantly dropped compared with M-NEC. Predictive modeling was able to accurately predict S-NEC from M-NEC and controls. Conclusion: There are discrete leukocyte patterns in NEC based on GA. The CBC at diagnosis may be useful in identifying patients who will require surgery.
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Affiliation(s)
- Julia M Pantalone
- University of Pittsburgh School of Medicine, Pittsburgh, PA, United States
| | - Silvia Liu
- Department of Pathology, University of Pittsburgh, Pittsburgh, PA, United States
| | - Oluwabunmi O Olaloye
- Department of Pediatrics, University of Pittsburgh, Pittsburgh, PA, United States.,Department of Pediatrics, Yale School of Medicine, New Haven, CT, United States
| | - Erica C Prochaska
- Department of Pediatrics, University of Pittsburgh, Pittsburgh, PA, United States.,Division of Infectious Diseases, Department of Pediatrics, Johns Hopkins University School of Medicine, Baltimore, MD, United States
| | - Toby Yanowitz
- Department of Pediatrics, University of Pittsburgh, Pittsburgh, PA, United States
| | - Melissa M Riley
- Department of Pediatrics, University of Pittsburgh, Pittsburgh, PA, United States.,Department of Pediatrics, Washington University School of Medicine, St. Louis, MO, United States
| | - Justin R Buland
- Department of Pediatrics, University of Pittsburgh, Pittsburgh, PA, United States.,Department of Pediatrics, Washington University School of Medicine, St. Louis, MO, United States
| | - Beverly S Brozanski
- Department of Pediatrics, University of Pittsburgh, Pittsburgh, PA, United States.,Department of Pediatrics, Washington University School of Medicine, St. Louis, MO, United States
| | - Misty Good
- Department of Pediatrics, University of Pittsburgh, Pittsburgh, PA, United States.,Department of Pediatrics, Washington University School of Medicine, St. Louis, MO, United States
| | - Liza Konnikova
- University of Pittsburgh School of Medicine, Pittsburgh, PA, United States.,Department of Pediatrics, University of Pittsburgh, Pittsburgh, PA, United States.,Department of Pediatrics, Yale School of Medicine, New Haven, CT, United States.,Department of Immunology, University of Pittsburgh, Pittsburgh, PA, United States.,Department of Developmental Biology, University of Pittsburgh, Pittsburgh, PA, United States
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Khawaja AA, Chong DLW, Sahota J, Mikolasch TA, Pericleous C, Ripoll VM, Booth HL, Khan S, Rodriguez-Justo M, Giles IP, Porter JC. Identification of a Novel HIF-1α-α Mβ 2 Integrin-NET Axis in Fibrotic Interstitial Lung Disease. Front Immunol 2020; 11:2190. [PMID: 33178179 PMCID: PMC7594517 DOI: 10.3389/fimmu.2020.02190] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/05/2020] [Accepted: 08/11/2020] [Indexed: 12/18/2022] Open
Abstract
Neutrophilic inflammation correlates with mortality in fibrotic interstitial lung disease (ILD) particularly in the most severe form, idiopathic pulmonary fibrosis (IPF), although the underlying mechanisms remain unclear. Neutrophil function is modulated by numerous factors, including integrin activation, inflammatory cytokines and hypoxia. Hypoxia has an important role in inflammation and may also contribute to pulmonary disease. We aimed to determine how neutrophil activation occurs in ILD and the relative importance of hypoxia. Using lung biopsies and bronchoalveolar lavage (BAL) fluid from ILD patients we investigated the extent of hypoxia and neutrophil activation in ILD lungs. Then we used ex vivo neutrophils isolated from healthy volunteers and BAL from patients with ILD and non-ILD controls to further investigate aberrant neutrophil activation in hypoxia and ILD. We demonstrate for the first time using intracellular staining, HIF-1α stabilization in neutrophils and endothelial cells in ILD lung biopsies. Hypoxia enhanced both spontaneous (+1.31-fold, p < 0.05) and phorbol 12-myristate 13-acetate (PMA)-induced (+1.65-fold, p < 0.001) neutrophil extracellular trap (NET) release, neutrophil adhesion (+8.8-fold, <0.05), and trans-endothelial migration (+1.9-fold, p < 0.05). Hypoxia also increased neutrophil expression of the αM (+3.1-fold, p < 0.001) and αX (+1.6-fold, p < 0.01) integrin subunits. Interestingly, NET formation was induced by αMβ2 integrin activation and prevented by cation chelation. Finally, we observed NET-like structures in IPF lung sections and in the BAL from ILD patients, and quantification showed increased cell-free DNA content (+5.5-fold, p < 0.01) and MPO-citrullinated histone H3 complexes (+21.9-fold, p < 0.01) in BAL from ILD patients compared to non-ILD controls. In conclusion, HIF-1α upregulation may augment neutrophil recruitment and activation within the lung interstitium through activation of β2 integrins. Our results identify a novel HIF-1α- αMβ2 integrin axis in NET formation for future exploration in therapeutic approaches to fibrotic ILD.
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Affiliation(s)
- Akif A. Khawaja
- Centre for Inflammation and Tissue Repair, University College London, London, United Kingdom
- Centre for Rheumatology, University College London, London, United Kingdom
| | - Deborah L. W. Chong
- Centre for Inflammation and Tissue Repair, University College London, London, United Kingdom
| | - Jagdeep Sahota
- Centre for Inflammation and Tissue Repair, University College London, London, United Kingdom
| | - Theresia A. Mikolasch
- Centre for Inflammation and Tissue Repair, University College London, London, United Kingdom
- Interstitial Lung Disease Service, University College London Hospital NHS Foundation Trust, London, United Kingdom
| | - Charis Pericleous
- National Heart and Lung Institute, Imperial College London, London, United Kingdom
| | - Vera M. Ripoll
- Centre for Rheumatology, University College London, London, United Kingdom
| | - Helen L. Booth
- Interstitial Lung Disease Service, University College London Hospital NHS Foundation Trust, London, United Kingdom
| | - Saif Khan
- Institute of Nuclear Medicine, University College London, London, United Kingdom
| | - Manuel Rodriguez-Justo
- Department of Histopathology, University College London Hospital NHS Foundation Trust, London, United Kingdom
| | - Ian P. Giles
- Centre for Rheumatology, University College London, London, United Kingdom
| | - Joanna C. Porter
- Centre for Inflammation and Tissue Repair, University College London, London, United Kingdom
- Interstitial Lung Disease Service, University College London Hospital NHS Foundation Trust, London, United Kingdom
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Kim R, Attayek PJ, Wang Y, Furtado KL, Tamayo R, Sims CE, Allbritton NL. An in vitro intestinal platform with a self-sustaining oxygen gradient to study the human gut/microbiome interface. Biofabrication 2019; 12:015006. [PMID: 31519008 PMCID: PMC6933551 DOI: 10.1088/1758-5090/ab446e] [Citation(s) in RCA: 64] [Impact Index Per Article: 10.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
An oxygen gradient formed along the length of colonic crypts supports stem-cell proliferation at the normoxic crypt base while supporting obligate anaerobe growth in the anoxic colonic lumen. Primary human colonic epithelial cells derived from human gastrointestinal stem cells were cultured within a device possessing materials of tailored oxygen permeability to produce an oxygen-depleted luminal (0.8% ± 0.1% O2) and oxygen-rich basal (11.1% ± 0.5% O2) compartment. This oxygen difference created a stable oxygen gradient across the colonic epithelial cells which remained viable and properly polarized. Facultative and obligate anaerobes Lactobacillus rhamnosus, Bifidobacterium adolescentis, and Clostridium difficile grew readily within the luminal compartment. When formed along the length of an in vitro crypt, the oxygen gradient facilitated cell compartmentalization within the crypt by enhancing confinement of the proliferative cells to the crypt base. This platform provides a simple system to create a physiological oxygen gradient across an intestinal mimic while simultaneously supporting anaerobe co-culture.
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Affiliation(s)
- Raehyun Kim
- Joint Department of Biomedical Engineering, University of North Carolina, Chapel Hill, and North Carolina State University, Raleigh, North Carolina
| | - Peter J. Attayek
- Joint Department of Biomedical Engineering, University of North Carolina, Chapel Hill, and North Carolina State University, Raleigh, North Carolina
| | - Yuli Wang
- Department of Chemistry, University of North Carolina, Chapel Hill, North Carolina
| | - Kathleen L. Furtado
- Department of Microbiology and Immunology, University of North Carolina, Chapel Hill, North Carolina
| | - Rita Tamayo
- Department of Microbiology and Immunology, University of North Carolina, Chapel Hill, North Carolina
| | - Christopher E. Sims
- Department of Chemistry, University of North Carolina, Chapel Hill, North Carolina
| | - Nancy L. Allbritton
- Joint Department of Biomedical Engineering, University of North Carolina, Chapel Hill, and North Carolina State University, Raleigh, North Carolina
- Department of Chemistry, University of North Carolina, Chapel Hill, North Carolina
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9
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Triner D, Shah YM. Hypoxic Regulation of Neutrophils in Cancer. Int J Mol Sci 2019; 20:ijms20174189. [PMID: 31461847 PMCID: PMC6747474 DOI: 10.3390/ijms20174189] [Citation(s) in RCA: 13] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/24/2019] [Revised: 08/21/2019] [Accepted: 08/23/2019] [Indexed: 12/28/2022] Open
Abstract
Neutrophils have been well-characterized for their role in the host anti-microbial response. However, it is now appreciated that neutrophils have a critical role in tumorigenesis and tumor progression in the majority of solid tumors. Recent studies have indicated a critical role for hypoxia in regulating neutrophil function in tumors. Furthermore, neutrophil-specific expression of hypoxia-inducible transcription factors may represent a novel therapeutic target for human cancer. In this review, we highlight the function of neutrophils in cancer and the role of the neutrophil hypoxic response in regulating the neoplastic progression of cancer.
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Affiliation(s)
- Daniel Triner
- Department of Molecular & Integrative Physiology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Yatrik M Shah
- Department of Molecular & Integrative Physiology, University of Michigan Medical School, Ann Arbor, MI 48109, USA.
- Division of Gastroenterology, Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI 48109, USA.
- Rogel Cancer Center, University of Michigan Medical School, Ann Arbor, MI 48109, USA.
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10
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Abstract
Redox signalling in the gastrointestinal mucosa is held in an intricate balance. Potent microbicidal mechanisms can be used by infiltrating immune cells, such as neutrophils, to protect compromised mucosae from microbial infection through the generation of reactive oxygen species. Unchecked, collateral damage to the surrounding tissue from neutrophil-derived reactive oxygen species can be detrimental; thus, maintenance and restitution of a breached intestinal mucosal barrier are paramount to host survival. Redox reactions and redox signalling have been studied for decades with a primary focus on contributions to disease processes. Within the past decade, an upsurge of exciting findings have implicated subtoxic levels of oxidative stress in processes such as maintenance of mucosal homeostasis, the control of protective inflammation and even regulation of tissue wound healing. Resident gut microbial communities have been shown to trigger redox signalling within the mucosa, which expresses similar but distinct enzymes to phagocytes. At the fulcrum of this delicate balance is the colonic mucosal epithelium, and emerging evidence suggests that precise control of redox signalling by these barrier-forming cells may dictate the outcome of an inflammatory event. This Review will address both the spectrum and intensity of redox activity pertaining to host-immune and host-microbiota crosstalk during homeostasis and disease processes in the gastrointestinal tract.
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11
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Simmen S, Cosin-Roger J, Melhem H, Maliachovas N, Maane M, Baebler K, Weder B, Maeyashiki C, Spanaus K, Scharl M, de Vallière C, Zeitz J, Vavricka SR, Hausmann M, Rogler G, Ruiz PA. Iron Prevents Hypoxia-Associated Inflammation Through the Regulation of Nuclear Factor-κB in the Intestinal Epithelium. Cell Mol Gastroenterol Hepatol 2018; 7:339-355. [PMID: 30704983 PMCID: PMC6357696 DOI: 10.1016/j.jcmgh.2018.10.006] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/04/2018] [Revised: 09/24/2018] [Accepted: 10/01/2018] [Indexed: 12/15/2022]
Abstract
BACKGROUND & AIMS Hypoxia-associated pathways influence the development of inflammatory bowel disease. Adaptive responses to hypoxia are mediated through hypoxia-inducible factors, which are regulated by iron-dependent hydroxylases. Signals reflecting oxygen tension and iron levels in enterocytes regulate iron metabolism. Conversely, iron availability modulates responses to hypoxia. In the present study we sought to elucidate how iron influences the responses to hypoxia in the intestinal epithelium. METHODS Human subjects were exposed to hypoxia, and colonic biopsy specimens and serum samples were collected. HT-29, Caco-2, and T84 cells were subjected to normoxia or hypoxia in the presence of iron or the iron chelator deferoxamine. Changes in inflammatory gene expression and signaling were assessed by quantitative polymerase chain reaction and Western blot. Chromatin immunoprecipitation was performed using antibodies against nuclear factor (NF)-κB and primers for the promoter of tumor necrosis factor (TNF) and interleukin (IL)1β. RESULTS Human subjects presented reduced levels of ferritin in the intestinal epithelium after hypoxia. Hypoxia reduced iron deprivation-associated TNF and IL1β expression in HT-29 cells through the induction of autophagy. Contrarily, hypoxia triggered TNF and IL1β expression, and NF-κB activation in Caco-2 and T84 cells. Iron blocked autophagy in Caco-2 cells, while reducing hypoxia-associated TNF and IL1β expression through the inhibition of NF-κB binding to the promoter of TNF and IL1β. CONCLUSIONS Hypoxia promotes iron mobilization from the intestinal epithelium. Hypoxia-associated autophagy reduces inflammatory processes in HT-29 cells. In Caco-2 cells, iron uptake is essential to counteract hypoxia-induced inflammation. Iron mobilization into enterocytes may be a vital protective mechanism in the hypoxic inflamed mucosa.
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Affiliation(s)
- Simona Simmen
- Department of Gastroenterology and Hepatology, University Hospital Zurich, University of Zurich, Zurich, Switzerland
| | - Jesus Cosin-Roger
- Department of Gastroenterology and Hepatology, University Hospital Zurich, University of Zurich, Zurich, Switzerland
| | - Hassan Melhem
- Department of Gastroenterology and Hepatology, University Hospital Zurich, University of Zurich, Zurich, Switzerland
| | - Nikolaos Maliachovas
- Department of Gastroenterology and Hepatology, University Hospital Zurich, University of Zurich, Zurich, Switzerland
| | - Max Maane
- Department of Gastroenterology and Hepatology, University Hospital Zurich, University of Zurich, Zurich, Switzerland
| | - Katharina Baebler
- Department of Gastroenterology and Hepatology, University Hospital Zurich, University of Zurich, Zurich, Switzerland
| | - Bruce Weder
- Department of Gastroenterology and Hepatology, University Hospital Zurich, University of Zurich, Zurich, Switzerland
| | - Chiaki Maeyashiki
- Department of Gastroenterology and Hepatology, University Hospital Zurich, University of Zurich, Zurich, Switzerland
| | - Katharina Spanaus
- Institute of Clinical Chemistry, University Hospital Zurich, University of Zurich, Zurich, Switzerland
| | - Michael Scharl
- Department of Gastroenterology and Hepatology, University Hospital Zurich, University of Zurich, Zurich, Switzerland
| | - Cheryl de Vallière
- Department of Gastroenterology and Hepatology, University Hospital Zurich, University of Zurich, Zurich, Switzerland
| | - Jonas Zeitz
- Department of Gastroenterology and Hepatology, University Hospital Zurich, University of Zurich, Zurich, Switzerland,Center of Gastroenterology, Clinic Hirslanden, Zurich, Switzerland
| | - Stephan R. Vavricka
- Department of Gastroenterology and Hepatology, University Hospital Zurich, University of Zurich, Zurich, Switzerland
| | - Martin Hausmann
- Department of Gastroenterology and Hepatology, University Hospital Zurich, University of Zurich, Zurich, Switzerland
| | - Gerhard Rogler
- Department of Gastroenterology and Hepatology, University Hospital Zurich, University of Zurich, Zurich, Switzerland
| | - Pedro A. Ruiz
- Department of Gastroenterology and Hepatology, University Hospital Zurich, University of Zurich, Zurich, Switzerland,Correspondence Address correspondence to: Pedro A. Ruiz-Castro, PhD, Department of Gastroenterology and Hepatology, University of Zurich, Raemistrasse 100, 8091 Zurich, Switzerland.
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12
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Cellular Stress Responses and Gut Microbiota in Inflammatory Bowel Disease. Gastroenterol Res Pract 2018; 2018:7192646. [PMID: 30026758 PMCID: PMC6031203 DOI: 10.1155/2018/7192646] [Citation(s) in RCA: 18] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/31/2018] [Accepted: 05/08/2018] [Indexed: 12/11/2022] Open
Abstract
Progresses in the past two decades have greatly expanded our understanding of inflammatory bowel disease (IBD), an incurable disease with multifaceted and challenging clinical manifestations. The pathogenesis of IBD involves multiple processes on the cellular level, which include the stress response signaling such as endoplasmic reticulum (ER) stress, oxidative stress, and hypoxia. Under physiological conditions, the stress responses play key roles in cell survival, mucosal barrier integrity, and immunomodulation. However, they can also cause energy depletion, trigger cell death and tissue injury, promote inflammatory response, and drive the progression of clinical disease. In recent years, gut microflora has emerged as an essential pathogenic factor and therapeutic target for IBD. Altered compositional and metabolic profiles of gut microbiota, termed dysbiosis, are associated with IBD. Recent studies, although limited, have shed light on how ER stress, oxidative stress, and hypoxic stress interact with gut microorganisms, a potential source of stress in the microenvironment of gastrointestinal tract. Our knowledge of cellular stress responses in intestinal homeostasis as well as their cross-talks with gut microbiome will further our understanding of the pathogenesis of inflammatory bowel disease and probably open avenues for new therapies.
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13
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Van Welden S, Selfridge AC, Hindryckx P. Intestinal hypoxia and hypoxia-induced signalling as therapeutic targets for IBD. Nat Rev Gastroenterol Hepatol 2017; 14:596-611. [PMID: 28853446 DOI: 10.1038/nrgastro.2017.101] [Citation(s) in RCA: 88] [Impact Index Per Article: 11.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/18/2022]
Abstract
Tissue hypoxia occurs when local oxygen demand exceeds oxygen supply. In chronic inflammatory conditions such as IBD, the increased oxygen demand by resident and gut-infiltrating immune cells coupled with vascular dysfunction brings about a marked reduction in mucosal oxygen concentrations. To counter the hypoxic challenge and ensure their survival, mucosal cells induce adaptive responses, including the activation of hypoxia-inducible factors (HIFs) and modulation of nuclear factor-κB (NF-κB). Both pathways are tightly regulated by oxygen-sensitive prolyl hydroxylases (PHDs), which therefore represent promising therapeutic targets for IBD. In this Review, we discuss the involvement of mucosal hypoxia and hypoxia-induced signalling in the pathogenesis of IBD and elaborate in detail on the role of HIFs, NF-κB and PHDs in different cell types during intestinal inflammation. We also provide an update on the development of PHD inhibitors and discuss their therapeutic potential in IBD.
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Affiliation(s)
- Sophie Van Welden
- Department of Gastroenterology, Ghent University, De Pintelaan 185, 1K12-E, 9000 Ghent, Belgium
| | - Andrew C Selfridge
- Robarts Clinical Trials West, 4350 Executive Drive 210, San Diego, California 92121, USA
| | - Pieter Hindryckx
- Department of Gastroenterology, Ghent University, De Pintelaan 185, 1K12-E, 9000 Ghent, Belgium
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14
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Colgan SP, Campbell EL, Kominsky DJ. Hypoxia and Mucosal Inflammation. ANNUAL REVIEW OF PATHOLOGY-MECHANISMS OF DISEASE 2017; 11:77-100. [PMID: 27193451 DOI: 10.1146/annurev-pathol-012615-044231] [Citation(s) in RCA: 87] [Impact Index Per Article: 10.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/18/2022]
Abstract
Sites of inflammation are defined by significant changes in metabolic activity. Recent studies have suggested that O2 metabolism and hypoxia play a prominent role in inflammation so-called "inflammatory hypoxia," which results from a combination of recruited inflammatory cells (e.g., neutrophils and monocytes), the local proliferation of multiple cell types, and the activation of multiple O2-consuming enzymes during inflammation. These shifts in energy supply and demand result in localized regions of hypoxia and have revealed the important function off the transcription factor HIF (hypoxia-inducible factor) in the regulation of key target genes that promote inflammatory resolution. Analysis of these pathways has provided multiple opportunities for understanding basic mechanisms of inflammation and has defined new targets for intervention. Here, we review recent work addressing tissue hypoxia and metabolic control of inflammation and immunity.
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Affiliation(s)
- Sean P Colgan
- Department of Medicine, University of Colorado School of Medicine, Aurora, Colorado 80045; .,Mucosal Inflammation Program, University of Colorado School of Medicine, Aurora, Colorado 80045
| | - Eric L Campbell
- Department of Medicine, University of Colorado School of Medicine, Aurora, Colorado 80045; .,Mucosal Inflammation Program, University of Colorado School of Medicine, Aurora, Colorado 80045
| | - Douglas J Kominsky
- Mucosal Inflammation Program, University of Colorado School of Medicine, Aurora, Colorado 80045.,Department of Microbiology and Immunology, Montana State University, Bozeman, Montana 59717
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15
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Montgomery ST, Mall MA, Kicic A, Stick SM. Hypoxia and sterile inflammation in cystic fibrosis airways: mechanisms and potential therapies. Eur Respir J 2016; 49:13993003.00903-2016. [DOI: 10.1183/13993003.00903-2016] [Citation(s) in RCA: 65] [Impact Index Per Article: 7.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/05/2016] [Accepted: 08/31/2016] [Indexed: 12/20/2022]
Abstract
Cystic fibrosis is one of the most common autosomal recessive genetic diseases in Caucasian populations. Diagnosisvianewborn screening and targeted nutritional and antibiotic therapy have improved outcomes, however respiratory failure remains the key cause of morbidity and mortality. Progressive respiratory disease in cystic fibrosis is characterised by chronic neutrophilic airway inflammation associated with structural airway damage leading to bronchiectasis and decreased lung function. Mucus obstruction is a characteristic early abnormality in the cystic fibrosis airway, associated with neutrophilic inflammation often in the absence of detectable infection. Recent studies have suggested a link between hypoxic cell death and sterile neutrophilic inflammation in cystic fibrosis and other diseasesviathe IL-1 signalling pathway. In this review, we consider recent evidence regarding the cellular responses to respiratory hypoxia as a potential driver of sterile neutrophilic inflammation in the lung, current knowledge on hypoxia as a pathogenic mechanism in cystic fibrosis and the potential for current and future therapies to alleviate hypoxia-driven sterile inflammation.
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16
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Lodge KM, Thompson AAR, Chilvers ER, Condliffe AM. Hypoxic regulation of neutrophil function and consequences for Staphylococcus aureus infection. Microbes Infect 2016; 19:166-176. [PMID: 27789256 DOI: 10.1016/j.micinf.2016.10.005] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/28/2016] [Revised: 10/07/2016] [Accepted: 10/12/2016] [Indexed: 01/22/2023]
Abstract
Staphylococcal infection and neutrophilic inflammation can act in concert to establish a profoundly hypoxic environment. In this review we summarise how neutrophils and Staphylococcus aureus are adapted to function under hypoxic conditions, with a particular focus on the impaired ability of hypoxic neutrophils to effect Staphylococcus aureus killing.
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Affiliation(s)
- Katharine M Lodge
- Department of Medicine, University of Cambridge School of Clinical Medicine, Cambridge University Hospitals, Hills Road, Cambridge, CB2 0QQ, UK
| | - A A Roger Thompson
- Department of Infection, Immunity and Cardiovascular Disease, University of Sheffield Medical School, Beech Hill Road, Sheffield, S10 2RX, UK
| | - Edwin R Chilvers
- Department of Medicine, University of Cambridge School of Clinical Medicine, Cambridge University Hospitals, Hills Road, Cambridge, CB2 0QQ, UK.
| | - Alison M Condliffe
- Department of Infection, Immunity and Cardiovascular Disease, University of Sheffield Medical School, Beech Hill Road, Sheffield, S10 2RX, UK
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17
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Blaisdell A, Crequer A, Columbus D, Daikoku T, Mittal K, Dey SK, Erlebacher A. Neutrophils Oppose Uterine Epithelial Carcinogenesis via Debridement of Hypoxic Tumor Cells. Cancer Cell 2015; 28:785-799. [PMID: 26678340 PMCID: PMC4698345 DOI: 10.1016/j.ccell.2015.11.005] [Citation(s) in RCA: 122] [Impact Index Per Article: 12.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/11/2015] [Revised: 08/20/2015] [Accepted: 11/17/2015] [Indexed: 12/19/2022]
Abstract
Polymorphonuclear neutrophils (PMNs) are largely considered to foster cancer development despite wielding an arsenal of cytotoxic agents. Using a mouse model of PTEN-deficient uterine cancer, we describe a surprising inhibitory role for PMNs in epithelial carcinogenesis. By inducing tumor cell detachment from the basement membrane, PMNs impeded early-stage tumor growth and retarded malignant progression. Unexpectedly, PMN recruitment and tumor growth control occurred independently of lymphocytes and cellular senescence and instead ensued as part of the tumor's intrinsic inflammatory response to hypoxia. In humans, a PMN gene signature correlated with improved survival in several cancer subtypes, including PTEN-deficient uterine cancer. These findings provide insight into tumor-associated PMNs and reveal a context-specific capacity for PMNs to directly combat tumorigenesis.
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MESH Headings
- Animals
- Bone Marrow Transplantation
- Carcinoma, Endometrioid/enzymology
- Carcinoma, Endometrioid/genetics
- Carcinoma, Endometrioid/immunology
- Carcinoma, Endometrioid/mortality
- Carcinoma, Endometrioid/pathology
- Carcinoma, Endometrioid/prevention & control
- Cell Adhesion
- Cell Hypoxia
- Cell Line, Tumor
- Cell Proliferation
- Chemotaxis
- Computational Biology
- Databases, Genetic
- Female
- Gene Expression Profiling
- Gene Transfer Techniques
- Humans
- Inflammation Mediators/metabolism
- Mice, Inbred C57BL
- Mice, Knockout
- Myeloid Differentiation Factor 88/deficiency
- Myeloid Differentiation Factor 88/genetics
- Neoplasm Staging
- Neutrophil Activation
- Neutrophil Infiltration
- Neutrophils/immunology
- Neutrophils/metabolism
- Ovarian Neoplasms/enzymology
- Ovarian Neoplasms/genetics
- Ovarian Neoplasms/immunology
- Ovarian Neoplasms/mortality
- Ovarian Neoplasms/pathology
- Ovarian Neoplasms/prevention & control
- Ovariectomy
- PTEN Phosphohydrolase/deficiency
- PTEN Phosphohydrolase/genetics
- Phagocytosis
- Receptors, Colony-Stimulating Factor/deficiency
- Receptors, Colony-Stimulating Factor/genetics
- Receptors, Interleukin-8B/genetics
- Receptors, Interleukin-8B/metabolism
- Survival Analysis
- Time Factors
- Tumor Burden
- Tumor Microenvironment
- Uterus/enzymology
- Uterus/immunology
- Uterus/pathology
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Affiliation(s)
- Adam Blaisdell
- Department of Pathology, NYU School of Medicine, New York, NY 10016, USA
| | - Amandine Crequer
- Department of Pathology, NYU School of Medicine, New York, NY 10016, USA
| | - Devin Columbus
- Department of Pathology, NYU School of Medicine, New York, NY 10016, USA
| | - Takiko Daikoku
- Division of Reproductive Sciences, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA
| | - Khush Mittal
- Department of Pathology, NYU School of Medicine, New York, NY 10016, USA
| | - Sudhansu K Dey
- Division of Reproductive Sciences, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA
| | - Adrian Erlebacher
- Department of Pathology, NYU School of Medicine, New York, NY 10016, USA; NYU Cancer Institute, NYU Langone Medical Center, New York, NY 10016, USA.
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18
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Eosinophils and IL-33 Perpetuate Chronic Inflammation and Fibrosis in a Pediatric Population with Stricturing Crohn's Ileitis. Inflamm Bowel Dis 2015; 21. [PMID: 26218140 PMCID: PMC4567482 DOI: 10.1097/mib.0000000000000512] [Citation(s) in RCA: 31] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
BACKGROUND Fibrostenosis and stricture are well-recognized endpoints in Crohn's disease (CD). We hypothesized that stricturing CD is characterized by eosinophilia and epithelial IL-33. We proposed that eosinophil exposure to IL-33 would perpetuate inflammatory chronicity and subsequent fibrostenosis. METHODS We performed a retrospective study of 74 children with inflammatory and stricturing ileal CD comparing clinicopathological features to immunohistochemical measures of eosinophilia and IL-33. To scrutinize eosinophil patterns, we developed a novel eosinophil peroxidase score encompassing number, distribution, and degranulation. Human eosinophils and intestinal fibroblasts were cultured with IL-33 and IL-13, and inflammatory and remodeling parameters were assessed. Antieosinophil therapy was also administered to the Crohn's-like ileitis model (SAMP1/SkuSlc). RESULTS Our novel eosinophil peroxidase score was more sensitive than H&E staining, revealing significant differences in eosinophil patterns, comparing inflammatory and stricturing pediatric CD. A significant relationship between ileal eosinophilia and complicated clinical/histopathological phenotype including fibrosis was determined. IL-33 induced significant eosinophil peroxidase secretion and IL-13 production. Exposure to eosinophils in the presence of IL-33, "primed" fibroblasts to increase proinflammatory cytokines (TNF-α, IL-1β, and IL-6), eosinophil-associated chemokines (CCL24 and CCL26), and IL-13Rα2 production. Production of fibrogenic molecules (collagen 1A2, fibronectin, and periostin) increased after exposure of "primed" fibroblasts to IL-13. Epithelial-IL-33 was increased in pediatric Crohn's ileitis and strongly associated with clinical and histopathological activity, ileal eosinophilia, and complicated fibrostenotic disease. SAMP1/SkuSlc eosinophil-targeted treatment resulted in significant improvements in inflammation and remodeling. CONCLUSIONS Our study of specimens from pediatric patients with ileal CD linked eosinophil patterns and IL-33 to fibrosis and suggested that these may contribute to the perpetuation of inflammation and subsequent stricture in pediatric CD.
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19
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Masterson JC, McNamee EN, Fillon SA, Hosford L, Harris R, Fernando SD, Jedlicka P, Iwamoto R, Jacobsen E, Protheroe C, Eltzschig HK, Colgan SP, Arita M, Lee JJ, Furuta GT. Eosinophil-mediated signalling attenuates inflammatory responses in experimental colitis. Gut 2015; 64:1236-47. [PMID: 25209655 PMCID: PMC4515997 DOI: 10.1136/gutjnl-2014-306998] [Citation(s) in RCA: 95] [Impact Index Per Article: 9.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/11/2014] [Accepted: 08/19/2014] [Indexed: 02/06/2023]
Abstract
OBJECTIVE Eosinophils reside in the colonic mucosa and increase significantly during disease. Although a number of studies have suggested that eosinophils contribute to the pathogenesis of GI inflammation, the expanding scope of eosinophil-mediated activities indicate that they also regulate local immune responses and modulate tissue inflammation. We sought to define the impact of eosinophils that respond to acute phases of colitis in mice. DESIGN Acute colitis was induced in mice by administration of dextran sulfate sodium, 2,4,6-trinitrobenzenesulfonic acid or oxazolone to C57BL/6J (control) or eosinophil deficient (PHIL) mice. Eosinophils were also depleted from mice using antibodies against interleukin (IL)-5 or by grafting bone marrow from PHIL mice into control mice. Colon tissues were collected and analysed by immunohistochemistry, flow cytometry and reverse transcription PCR; lipids were analysed by mass spectroscopy. RESULTS Eosinophil-deficient mice developed significantly more severe colitis, and their colon tissues contained a greater number of neutrophils, than controls. This compensatory increase in neutrophils was accompanied by increased levels of the chemokines CXCL1 and CXCL2, which attract neutrophils. Lipidomic analyses of colonic tissue from eosinophil-deficient mice identified a deficiency in the docosahexaenoic acid-derived anti-inflammatory mediator 10, 17- dihydroxydocosahexaenoic acid (diHDoHE), namely protectin D1 (PD1). Administration of an exogenous PD1-isomer (10S, 17S-DiHDoHE) reduced the severity of colitis in eosinophil-deficient mice. The PD1-isomer also attenuated neutrophil infiltration and reduced levels of tumour necrosis factor-α, IL-1β, IL-6 and inducible NO-synthase in colons of mice. Finally, in vitro assays identified a direct inhibitory effect of PD1-isomer on neutrophil transepithelial migration. CONCLUSIONS Eosinophils exert a protective effect in acute mouse colitis, via production of anti-inflammatory lipid mediators.
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Affiliation(s)
- Joanne C Masterson
- Section of Pediatric Gastroenterology, Hepatology and Nutrition, Gastrointestinal Eosinophilic Diseases Program, Department of Pediatrics; Digestive Health Institute, Children's Hospital Colorado, University of Colorado School of Medicine, Aurora, Colorado, USA,Mucosal Inflammation Program, University of Colorado School of Medicine, Aurora, Colorado, USA,University of Colorado School of Medicine, Aurora, Colorado, USA
| | - Eóin N McNamee
- Mucosal Inflammation Program, University of Colorado School of Medicine, Aurora, Colorado, USA,University of Colorado School of Medicine, Aurora, Colorado, USA,Department of Anesthesiology, University of Colorado School of Medicine, Aurora, Colorado, USA
| | - Sophie A Fillon
- Section of Pediatric Gastroenterology, Hepatology and Nutrition, Gastrointestinal Eosinophilic Diseases Program, Department of Pediatrics; Digestive Health Institute, Children's Hospital Colorado, University of Colorado School of Medicine, Aurora, Colorado, USA,Mucosal Inflammation Program, University of Colorado School of Medicine, Aurora, Colorado, USA,University of Colorado School of Medicine, Aurora, Colorado, USA
| | - Lindsay Hosford
- Section of Pediatric Gastroenterology, Hepatology and Nutrition, Gastrointestinal Eosinophilic Diseases Program, Department of Pediatrics; Digestive Health Institute, Children's Hospital Colorado, University of Colorado School of Medicine, Aurora, Colorado, USA,Mucosal Inflammation Program, University of Colorado School of Medicine, Aurora, Colorado, USA,University of Colorado School of Medicine, Aurora, Colorado, USA
| | - Rachel Harris
- Section of Pediatric Gastroenterology, Hepatology and Nutrition, Gastrointestinal Eosinophilic Diseases Program, Department of Pediatrics; Digestive Health Institute, Children's Hospital Colorado, University of Colorado School of Medicine, Aurora, Colorado, USA,Mucosal Inflammation Program, University of Colorado School of Medicine, Aurora, Colorado, USA,University of Colorado School of Medicine, Aurora, Colorado, USA
| | - Shahan D Fernando
- Section of Pediatric Gastroenterology, Hepatology and Nutrition, Gastrointestinal Eosinophilic Diseases Program, Department of Pediatrics; Digestive Health Institute, Children's Hospital Colorado, University of Colorado School of Medicine, Aurora, Colorado, USA,Mucosal Inflammation Program, University of Colorado School of Medicine, Aurora, Colorado, USA,University of Colorado School of Medicine, Aurora, Colorado, USA
| | - Paul Jedlicka
- University of Colorado School of Medicine, Aurora, Colorado, USA,Department of Pathology, University of Colorado School of Medicine, Aurora, Colorado, USA
| | - Ryo Iwamoto
- Department of Health Chemistry, Graduate School of Pharmaceutical Sciences, University of Tokyo, Tokyo, Japan
| | - Elizabeth Jacobsen
- Laboratory for Metabolomics, RIKEN Center for Integrative Medical Sciences (IMS), Yokohama, Japan,Department of Biochemistry and Molecular Biology, Mayo Clinic Scottsdale, Scottsdale, Arizona, USA
| | - Cheryl Protheroe
- Laboratory for Metabolomics, RIKEN Center for Integrative Medical Sciences (IMS), Yokohama, Japan,Department of Biochemistry and Molecular Biology, Mayo Clinic Scottsdale, Scottsdale, Arizona, USA
| | - Holger K Eltzschig
- Mucosal Inflammation Program, University of Colorado School of Medicine, Aurora, Colorado, USA,University of Colorado School of Medicine, Aurora, Colorado, USA,Department of Anesthesiology, University of Colorado School of Medicine, Aurora, Colorado, USA
| | - Sean P Colgan
- Mucosal Inflammation Program, University of Colorado School of Medicine, Aurora, Colorado, USA,University of Colorado School of Medicine, Aurora, Colorado, USA,Department of Medicine, University of Colorado School of Medicine, Aurora, Colorado, USA
| | - Makoto Arita
- Department of Health Chemistry, Graduate School of Pharmaceutical Sciences, University of Tokyo, Tokyo, Japan,Laboratory for Metabolomics, RIKEN Center for Integrative Medical Sciences (IMS), Yokohama, Japan
| | - James J Lee
- Department of Biochemistry and Molecular Biology, Mayo Clinic Scottsdale, Scottsdale, Arizona, USA
| | - Glenn T Furuta
- Section of Pediatric Gastroenterology, Hepatology and Nutrition, Gastrointestinal Eosinophilic Diseases Program, Department of Pediatrics; Digestive Health Institute, Children's Hospital Colorado, University of Colorado School of Medicine, Aurora, Colorado, USA,Mucosal Inflammation Program, University of Colorado School of Medicine, Aurora, Colorado, USA,University of Colorado School of Medicine, Aurora, Colorado, USA
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20
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Salvadori M, Rosso G, Bertoni E. Update on ischemia-reperfusion injury in kidney transplantation: Pathogenesis and treatment. World J Transplant 2015; 5:52-67. [PMID: 26131407 PMCID: PMC4478600 DOI: 10.5500/wjt.v5.i2.52] [Citation(s) in RCA: 259] [Impact Index Per Article: 25.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/27/2014] [Revised: 01/12/2015] [Accepted: 04/29/2015] [Indexed: 02/05/2023] Open
Abstract
Ischemia/reperfusion injury is an unavoidable relevant consequence after kidney transplantation and influences short term as well as long-term graft outcome. Clinically ischemia/reperfusion injury is associated with delayed graft function, graft rejection, chronic rejection and chronic graft dysfunction. Ischemia/reperfusion affects many regulatory systems at the cellular level as well as in the renal tissue that result in a distinct inflammatory reaction of the kidney graft. Underlying factors of ischemia reperfusion include energy metabolism, cellular changes of the mitochondria and cellular membranes, initiation of different forms of cell death-like apoptosis and necrosis together with a recently discovered mixed form termed necroptosis. Chemokines and cytokines together with other factors promote the inflammatory response leading to activation of the innate immune system as well as the adaptive immune system. If the inflammatory reaction continues within the graft tissue, a progressive interstitial fibrosis develops that impacts long-term graft outcome. It is of particular importance in kidney transplantation to understand the underlying mechanisms and effects of ischemia/reperfusion on the graft as this knowledge also opens strategies to prevent or treat ischemia/reperfusion injury after transplantation in order to improve graft outcome.
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21
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Wang G, Zhao T, Song X, Zhong W, Yu L, Hua W, Xing MMQ, Qiu X. A 3-D multicellular tumor spheroid on ultrathin matrix coated single cancer cells provides a tumor microenvironment model to study epithelial-to-mesenchymal transitions. Polym Chem 2015; 6:283-293. [DOI: 10.1039/c4py01161a] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/30/2024]
Abstract
We report a 3D tumor spheroid model to study epithelial-to-mesenchymal transitions (EMT) using ultra-thin matrix coated single cancer cells.
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Affiliation(s)
- Guobao Wang
- Department of Anatomy
- Guangdong Provincial Key Laboratory of Construction and Detection in Tissue Engineering
- Southern Medical University
- Guangzhou 510515
- P.R. China
| | - Tingting Zhao
- Department of Anatomy
- Guangdong Provincial Key Laboratory of Construction and Detection in Tissue Engineering
- Southern Medical University
- Guangzhou 510515
- P.R. China
| | - Xiaoping Song
- Department of Anatomy
- Guangdong Provincial Key Laboratory of Construction and Detection in Tissue Engineering
- Southern Medical University
- Guangzhou 510515
- P.R. China
| | - Wen Zhong
- Department of Textile Sciences
- University of Manitoba
- Winnipeg
- Canada
| | - Lei Yu
- Department of Anatomy
- Guangdong Provincial Key Laboratory of Construction and Detection in Tissue Engineering
- Southern Medical University
- Guangzhou 510515
- P.R. China
| | - Wenxi Hua
- Department of Anatomy
- Guangdong Provincial Key Laboratory of Construction and Detection in Tissue Engineering
- Southern Medical University
- Guangzhou 510515
- P.R. China
| | - Malcolm M. Q. Xing
- Department of Mechanical and Manitoba Institute of Child Health
- University of Manitoba
- Winnipeg, MB R3T 2N2
- Canada
| | - Xiaozhong Qiu
- Department of Anatomy
- Guangdong Provincial Key Laboratory of Construction and Detection in Tissue Engineering
- Southern Medical University
- Guangzhou 510515
- P.R. China
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22
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Harris AJ, Thompson AR, Whyte MK, Walmsley SR. HIF-mediated innate immune responses: cell signaling and therapeutic implications. HYPOXIA 2014; 2:47-58. [PMID: 27774466 PMCID: PMC5045056 DOI: 10.2147/hp.s50269] [Citation(s) in RCA: 43] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Indexed: 12/28/2022]
Abstract
Leukocytes recruited to infected, damaged, or inflamed tissues during an immune response must adapt to oxygen levels much lower than those in the circulation. Hypoxia inducible factors (HIFs) are key mediators of cellular responses to hypoxia and, as in other cell types, HIFs are critical for the upregulation of glycolysis, which enables innate immune cells to produce adenosine triphosphate anaerobically. An increasing body of evidence demonstrates that hypoxia also regulates many other innate immunological functions, including cell migration, apoptosis, phagocytosis of pathogens, antigen presentation and production of cytokines, chemokines, and angiogenic and antimicrobial factors. Many of these functions are mediated by HIFs, which are not only stabilized posttranslationally by hypoxia, but also transcriptionally upregulated by inflammatory signals. Here, we review the role of HIFs in the responses of innate immune cells to hypoxia, both in vitro and in vivo, with a particular focus on myeloid cells, on which the majority of studies have so far been carried out.
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Affiliation(s)
- Alison J Harris
- Academic Unit of Respiratory Medicine, Department of Infection and Immunity, University of Sheffield, Sheffield, UK
| | - Aa Roger Thompson
- Academic Unit of Respiratory Medicine, Department of Infection and Immunity, University of Sheffield, Sheffield, UK
| | - Moira Kb Whyte
- Academic Unit of Respiratory Medicine, Department of Infection and Immunity, University of Sheffield, Sheffield, UK
| | - Sarah R Walmsley
- Academic Unit of Respiratory Medicine, Department of Infection and Immunity, University of Sheffield, Sheffield, UK
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Pathogenesis of human enterovirulent bacteria: lessons from cultured, fully differentiated human colon cancer cell lines. Microbiol Mol Biol Rev 2014; 77:380-439. [PMID: 24006470 DOI: 10.1128/mmbr.00064-12] [Citation(s) in RCA: 52] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023] Open
Abstract
Hosts are protected from attack by potentially harmful enteric microorganisms, viruses, and parasites by the polarized fully differentiated epithelial cells that make up the epithelium, providing a physical and functional barrier. Enterovirulent bacteria interact with the epithelial polarized cells lining the intestinal barrier, and some invade the cells. A better understanding of the cross talk between enterovirulent bacteria and the polarized intestinal cells has resulted in the identification of essential enterovirulent bacterial structures and virulence gene products playing pivotal roles in pathogenesis. Cultured animal cell lines and cultured human nonintestinal, undifferentiated epithelial cells have been extensively used for understanding the mechanisms by which some human enterovirulent bacteria induce intestinal disorders. Human colon carcinoma cell lines which are able to express in culture the functional and structural characteristics of mature enterocytes and goblet cells have been established, mimicking structurally and functionally an intestinal epithelial barrier. Moreover, Caco-2-derived M-like cells have been established, mimicking the bacterial capture property of M cells of Peyer's patches. This review intends to analyze the cellular and molecular mechanisms of pathogenesis of human enterovirulent bacteria observed in infected cultured human colon carcinoma enterocyte-like HT-29 subpopulations, enterocyte-like Caco-2 and clone cells, the colonic T84 cell line, HT-29 mucus-secreting cell subpopulations, and Caco-2-derived M-like cells, including cell association, cell entry, intracellular lifestyle, structural lesions at the brush border, functional lesions in enterocytes and goblet cells, functional and structural lesions at the junctional domain, and host cellular defense responses.
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Eguchi T, Hamanaka K, Kondo R, Saito G, Shiina T, Koizumi T, Yoshida K. Lung re-expansion following one-lung ventilation induces neutrophil cytoskeletal rearrangements in rats. Ann Thorac Cardiovasc Surg 2013; 20:276-83. [PMID: 23801182 DOI: 10.5761/atcs.oa.13.02247] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022] Open
Abstract
PURPOSE To investigate the morphological and functional behavior of neutrophils during and after one-lung ventilation (OLV). METHODS We utilized an OLV rat model system and performed 3 hours of OLV followed by either re-expansion (RE) and 30 minutes of two-lung ventilation (TLV) (RE group), only two-lung ventilation (TLV group), or only OLV (OLV group). Cytoskeletal rearrangements of circulating neutrophils were assessed by determining the localization of filamentous actin (F-actin). In addition, the number of sequestered neutrophils in the lung capillary and the cytokine-induced neutrophil chemoattractant 1 (CINC-1) levels in the plasma were determined. RESULTS The F-actin rimmed neutrophils in the RE group increased after RE, but did not increase in the other groups. In the RE group, the sequestered neutrophils in the ventilated lung were significantly more numerous, and the plasma CINC-1 levels were significantly higher than in the other groups. CONCLUSIONS Lung RE following OLV induces cytoskeletal rearrangements in circulating neutrophils and would thereby promote their sequestration in the lung capillaries. The plasma CINC-1 elevation after RE can be involved in neutrophil recruitment.
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Affiliation(s)
- Takashi Eguchi
- Department of Thoracic Surgery, Shinshu University School of Medicine, Matsumoto, Nagano, Japan
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25
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Li WA, Moore-Langston S, Chakraborty T, Rafols JA, Conti AC, Ding Y. Hyperglycemia in stroke and possible treatments. Neurol Res 2013; 35:479-91. [PMID: 23622737 DOI: 10.1179/1743132813y.0000000209] [Citation(s) in RCA: 44] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/04/2023]
Abstract
Hyperglycemia affects approximately one-third of acute ischemic stroke patients and is associated with poor clinical outcomes. In experimental and clinical stroke studies, hyperglycemia has been shown to be detrimental to the penumbral tissue for several reasons. First, hyperglycemia exacerbates both calcium imbalance and the accumulation of reactive oxygen species (ROS) in neurons, leading to increased apoptosis. Second, hyperglycemia fuels anaerobic energy production, causing lactic acidosis, which further stresses neurons in the penumbral regions. Third, hyperglycemia decreases blood perfusion after ischemic stroke by lowering the availability of nitric oxide (NO), which is a crucial mediator of vasodilation. Lastly, hyperglycemia intensifies the inflammatory response after stroke, causing edema, and hemorrhage through disruption of the blood brain barrier and degradation of white matter, which leads to a worsening of functional outcomes. Many neuroprotective treatments addressing hyperglycemia in stroke have been implemented in the past decade. Early clinical use of insulin provided mixed results due to insufficiently controlled glucose levels and heterogeneity of patient population. Recently, however, the latest Stroke Hyperglycemia Insulin Network Effort trial has addressed the shortcomings of insulin therapy. While glucagon-like protein-1 administration, hyperbaric oxygen preconditioning, and ethanol therapy appear promising, these treatments remain in their infancy and more research is needed to better understand the mechanisms underlying hyperglycemia-induced injuries. Elucidation of these mechanistic pathways could lead to the development of rational treatments that reduce hyperglycemia-associated injuries and improve functional outcomes for ischemic stroke patients.
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Affiliation(s)
- William A Li
- Department of Neurological Surgery, Wayne State University School of Medicine, Detroit, MI, USA
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26
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Hypoxia: a double-edged sword of immunity. J Mol Med (Berl) 2011; 89:657-65. [DOI: 10.1007/s00109-011-0724-8] [Citation(s) in RCA: 50] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/10/2010] [Revised: 01/05/2011] [Accepted: 01/06/2011] [Indexed: 12/12/2022]
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Clinical significance of a myeloperoxidase gene polymorphism and inducible nitric oxide synthase expression in cirrhotic patients with hepatopulmonary syndrome. ACTA ACUST UNITED AC 2010; 30:437-42. [DOI: 10.1007/s11596-010-0445-1] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/29/2009] [Indexed: 10/19/2022]
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Mirpuri J, Brazil JC, Berardinelli AJ, Nasr TR, Cooper K, Schnoor M, Lin PW, Parkos CA, Louis NA. Commensal Escherichia coli reduces epithelial apoptosis through IFN-alphaA-mediated induction of guanylate binding protein-1 in human and murine models of developing intestine. THE JOURNAL OF IMMUNOLOGY 2010; 184:7186-95. [PMID: 20483731 DOI: 10.4049/jimmunol.0903116] [Citation(s) in RCA: 28] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/23/2022]
Abstract
Appropriate microbial colonization protects the developing intestine by promoting epithelial barrier function and fostering mucosal tolerance to luminal bacteria. Commensal flora mediate their protective effects through TLR9-dependent activation of cytokines, such as type I IFNs (alpha, beta) and IL-10. Although IFN-beta promotes apoptosis, IFN-alpha activates specific antiapoptotic target genes whose actions preserve epithelial barrier integrity. We have recently identified guanylate binding protein-1 (GBP-1) as an antiapoptotic protein, regulated by both type I and type II IFNs, that promotes intestinal epithelial barrier integrity in mature intestine. However, the mechanisms by which commensal bacteria regulate epithelial apoptosis during colonization of immature intestine and the contributions of GBP-1 are unknown. The healthy newborn intestine is initially colonized with bacterial species present in the maternal gastrointestinal tract, including nonpathogenic Escherichia coli. Therefore, we examined the influence of commensal E. coli on cytokine expression and candidate mediators of apoptosis in preweaned mice. Specifically, enteral exposure of 2 wk-old mice to commensal E. coli for 24 h selectively increased both IFN-alphaA and GBP-1 mRNA expression and prevented staurosporine-induced epithelial apoptosis. Exogenous IFN-alphaA treatment also induced GBP-1 expression and protected against staurosporine-induced apoptosis in a GBP-1 dependent manner, both in vitro and ex vivo. These findings identify a role for IFN-alphaA-mediated GBP-1 expression in the prevention of intestinal epithelial apoptosis by commensal bacteria. Thus IFN-alphaA mediates the beneficial effects of commensal bacteria and may be a promising therapeutic target to promote barrier integrity and prevent the inappropriate inflammatory responses seen in developing intestine as in necrotizing enterocolitis.
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Affiliation(s)
- Julie Mirpuri
- Division of Neonatal-Perinatal Medicine, Department of Pediatrics, Emory University School of Medicine, Spelman College, Atlanta, GA 30322, USA
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Abstract
The abnormal decrease or the lack of oxygen supply to cells and tissues is called hypoxia. This condition is commonly seen in various diseases such as rheumatoid arthritis and atherosclerosis, also in solid cancers. Pre-clinical and clinical studies have shown that hypoxic cancers are extremely aggressive, resistant to standard therapies (chemotherapy and radiotherapy), and thus very difficult to eradicate. Hypoxia affects both the tumor and the immune cells via various pathways. This review summarizes the most common effects of hypoxia on immune cells that play a key role in the anti-tumor response, the limitation of current therapies, and the potential solutions that were developed for hypoxic malignancies.
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Affiliation(s)
- Patricia Yotnda
- Center for Cell and Gene Therapy, Baylor College of Medicine, One Baylor Plaza, 77030, Houston, TX, USA.
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30
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Frick JS, MacManus CF, Scully M, Glover LE, Eltzschig HK, Colgan SP. Contribution of adenosine A2B receptors to inflammatory parameters of experimental colitis. THE JOURNAL OF IMMUNOLOGY 2009; 182:4957-64. [PMID: 19342675 DOI: 10.4049/jimmunol.0801324] [Citation(s) in RCA: 126] [Impact Index Per Article: 7.9] [Reference Citation Analysis] [Abstract] [Subscribe] [Scholar Register] [Indexed: 01/08/2023]
Abstract
Inflammatory diseases influence tissue metabolism, significantly altering the profile of extracellular adenine nucleotides. A number of studies have suggested that adenosine (Ado) may function as an endogenously generated anti-inflammatory molecule. Given the central role of intestinal epithelial cells to the development of colitis, we hypothesized that specific Ado receptors would contribute to disease resolution in mucosal inflammation as modeled by dextran sodium sulfate (DSS) colitis. Initial profiling studies revealed that murine intestinal epithelial cells express predominantly the Ado A2B receptor (AA2BR) and to a lesser extent AA2AR. Guided by these results, we examined the contribution of AA2BR to colitis. Initial studies indicated that the severity of colitis was increased in Aa2br(-/-) mice relative to Aa2br(+/+) controls, as reflected by increased weight loss, colonic shortening, and disease activity indices. Likewise, enteral administration of the selective AA2BR inhibitor PSB1115 to Aa2br(+/+) mice resulted in a similar increase in severity of DSS colitis. Cytokine profiling of colonic tissue revealed specific deficiencies in IL-10 in Aa2br(-/-) mice relative to controls. Extensions of these findings in cultured human intestinal epithelial cells revealed that stable Ado analogs induce IL-10 mRNA and protein and that such increases can be blocked with PSB1115. Taken together, these studies indicate a central regulatory role for AA2BR-modulated IL-10 in the acute inflammatory phase of DSS colitis, thereby implicating AA2BR as an endogenously protective molecule expressed on intestinal epithelial cells.
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Affiliation(s)
- Julia-Stefanie Frick
- Institute for Medical Microbiology and Hygiene, University Hospital, Tübingen, Germany
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Keely S, Glover LE, MacManus CF, Campbell EL, Scully MM, Furuta GT, Colgan SP. Selective induction of integrin beta1 by hypoxia-inducible factor: implications for wound healing. FASEB J 2008; 23:1338-46. [PMID: 19103643 DOI: 10.1096/fj.08-125344] [Citation(s) in RCA: 77] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
Because of localized vascular damage and increased tissue oxygen demand, wound healing occurs in a relatively hypoxic microenvironment. These features are particularly relevant to wound healing and fibrosis in chronic inflammatory conditions, such as Crohn's disease and ulcerative colitis. In these studies, we sought to identify the contribution of hypoxia to mechanisms of wound repair in a model of the intestinal submucosa. Initial studies revealed that hypoxia promotes wound healing, as modeled by an increase in intestinal fibroblast-mediated collagen gel contraction. Guided by results from transcriptional profiling, we identified the selective induction of fibroblast integrin beta1 (ITGB1) by hypoxia. Further analysis revealed that hypoxia, as well as pharmacological activators of hypoxia-inducible factor (HIF), induce fibroblast beta1 integrin mRNA, protein, and function by as much as 4-fold. Cloning and analysis of the beta1 integrin gene promoter revealed a 10 +/- 0.8-fold increase in promoter activity in response to hypoxia, and subsequent studies identified a functional DNA binding region for HIF in the ITGB1 gene promoter. Mutational analysis of the HIF binding site within the ITGB1 promoter resulted in a significant loss of ITGB1 hypoxia-inducibility. As proof of principle, studies in a murine model of colitis revealed a correlation between colitic disease severity and tissue ITGB1 expression (R(2)=0.80). Taken together, these results demonstrate that hypoxia induces fibroblast ITGB1 expression and function by transcriptional mechanisms dependent on HIF.
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Affiliation(s)
- Simon Keely
- Mucosal Inflammation Program, Department of Medicine, University of Colorado Denver, Aurora, CO 80045, USA
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Robinson A, Keely S, Karhausen J, Gerich ME, Furuta GT, P.Colgan S. Mucosal protection by hypoxia-inducible factor prolyl hydroxylase inhibition. Gastroenterology 2008; 134:145-55. [PMID: 18166352 PMCID: PMC2194638 DOI: 10.1053/j.gastro.2007.09.033] [Citation(s) in RCA: 306] [Impact Index Per Article: 18.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/07/2007] [Accepted: 09/13/2007] [Indexed: 12/02/2022]
Abstract
BACKGROUND & AIMS A number of recent studies have implicated tissue hypoxia in both acute and chronic inflammatory diseases, particularly as they relate to mucosal surfaces lined by epithelial cells. In this context, a protective role for the transcriptional regulator hypoxia-inducible factor (HIF) was shown through conditional deletion of epithelial HIF-1alpha in a murine model of colitis. Here, we hypothesized that pharmacologic activation of HIF would similarly provide a protective adaptation to murine colitic disease. METHODS For these purposes, we used a novel prolyl hydroxylase (PHD) inhibitor (FG-4497) that readily stabilizes HIF-1alpha and subsequently drives the expression downstream of HIF target genes (eg, erythropoietin). RESULTS Our results show that the FG-4497-mediated induction of HIF-1alpha provides an overall beneficial influence on clinical symptoms [weight loss, colon length, tissue tumor necrosis factor-alpha (TNFalpha)] in murine trinitrobenzene sulfonic acid (TNBS) colitis, most likely because of their barrier protective function and wound healing during severe tissue hypoxia at the site of inflammation. CONCLUSIONS Taken together these findings emphasize the role of epithelial HIF-1alpha during inflammatory diseases in the colon and may provide the basis for a therapeutic use of PHD inhibitors in inflammatory mucosal disease.
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Affiliation(s)
- Andreas Robinson
- Mucosal Inflammation Program, Division of Gastroenterology, University of Colorado Health Sciences Center, Denver, CO
| | - Simon Keely
- Mucosal Inflammation Program, Division of Gastroenterology, University of Colorado Health Sciences Center, Denver, CO
| | - Jörn Karhausen
- Department of Anesthesiology and Intensive Care Medicine, Tübingen University Hospital, Tubingen, Germany
| | - Mark E. Gerich
- Mucosal Inflammation Program, Division of Gastroenterology, University of Colorado Health Sciences Center, Denver, CO
| | - Glenn T. Furuta
- Mucosal Inflammation Program, Division of Gastroenterology, University of Colorado Health Sciences Center, Denver, CO, Department of Pediatrics, Section of Pediatric Gastroenterology, Hepatology and Nutrition, The Children's Hospital, Denver, CO
| | - Sean P.Colgan
- Mucosal Inflammation Program, Division of Gastroenterology, University of Colorado Health Sciences Center, Denver, CO,Correspondence to: Sean P. Colgan, Ph.D., Mucosal Inflammation Program, BRB Room 702, 4200 E. 9th Ave Denver, CO 80220, USA. Office phone: 303−315−1064 Lab phone: 303−315−1068 E-mail:
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Carbon dioxide directly suppresses spontaneous migration, chemotaxis, and free radical production of human neutrophils. Surg Endosc 2007; 22:1813-7. [PMID: 18074179 DOI: 10.1007/s00464-007-9703-0] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/08/2007] [Revised: 09/11/2007] [Accepted: 10/04/2007] [Indexed: 12/20/2022]
Abstract
BACKGROUND Carbon dioxide (CO(2)) insufflation during laparoscopy has been shown to dampen the systemic stress response to surgery. This is related to a suppression of peritoneal macrophage functions. In vivo data suggest that CO(2) can also affect neutrophils (polymorphonuclear cells, PMNs), the most abundant cell type in the inflamed peritoneal cavity. Nonetheless, the direct effects of CO(2) on PMNs have not yet been investigated. METHOD PMNs were isolated from peripheral blood of healthy volunteers and incubated with (1) CO(2) (100% CO(2), pH 6.2), (2) hypoxic control (95% helium/5% CO(2), pH 7.4), and (3) control (95% air/5% CO(2), pH 7.4). Spontaneous and IL-8-induced migrations (chemokinesis and chemotaxis) during 2 h of exposure to different gases were measured with a transwell chamber system. The release of reactive oxygen species (ROS, luminometry) was determined after 15-min and 2-h exposures. In other sets of experiments, PMNs were exposed for 2 h or 4 h and kept under normal conditions for 18 h with lipopolysaccharide (LPS) stimulation thereafter. Final viability and apoptosis were assessed with fluorometry. RESULTS Exposure to 100% CO(2) completely blocked spontaneous and IL-8 induced migration of PMNs (p < 0.001 vs. controls). Neutrophil migration was slightly diminished in the hypoxic control group. PMA-stimulated ROS production was reduced even after short exposure to 100% CO(2)(p < 0.05). We observed a slight increase of caspase-3/7 activity after exposure to 100% CO(2) and/or hypoxia; however, total viability was not affected. CONCLUSIONS CO(2) incubation directly and temporarily suppresses the proinflammatory functions of PMNs; this is caused only partially by the concomitant hypoxia. This effect will contribute to the dampened inflammatory response to laparoscopic surgery. Further studies are needed to investigate whether the temporary suppression of neutrophil functions could affect the clearance of bacterial contaminations.
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Van Linden A, Eltzschig HK. Role of pulmonary adenosine during hypoxia: extracellular generation, signaling and metabolism by surface adenosine deaminase/CD26. Expert Opin Biol Ther 2007; 7:1437-47. [PMID: 17727332 DOI: 10.1517/14712598.7.9.1437] [Citation(s) in RCA: 37] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/05/2022]
Abstract
Numerous parallels exist between limited oxygen availability (hypoxia) and acute inflammation. The lungs in particular are prone to acute inflammation during hypoxia, resulting in pulmonary edema, vascular leakage and neutrophil infiltration. The innate response elicited by hypoxia is associated with increased extracellular adenosine effects. Although studies on acute pulmonary hypoxia show a protective role of extracellular adenosine by attenuating pulmonary edema and excessive inflammation, chronic elevation of pulmonary adenosine may be detrimental. Adenosine deaminase (ADA)-deficient mice, for example, develop signs of chronic pulmonary injury in association with highly elevated levels of adenosine. Thus, the authors hypothesized the existence of hypoxia-elicited clearance mechanisms to offset deleterious influences of chronically elevated adenosine. Such studies indicated a second response to hypoxia characterized by pulmonary induction of ADA and CD26. In fact, hypoxia-inducible ADA is enzymatically active and tethered on the outside of the membrane via CD26 to form a complex capable of degrading extracellular adenosine to inosine. This paper reviews metabolic and transcriptional changes of extracellular adenosine generation, signaling and degradation during acute and prolonged pulmonary hypoxia.
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Affiliation(s)
- Annemie Van Linden
- University of Colorado Health Science Center, Mucosal Inflammation Program, Department of Anesthesiology, Perioperative Care and Pain Medicine, Biochemistry Research Building (BRB), Room 852, 4200 E. 9th Avenue, Campus Box B113, Denver, CO 80262, USA
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Zhang HY, Han DW, Zhao ZF, Liu MS, Wu YJ, Chen XM, Ji C. Multiple pathogenic factor-induced complications of cirrhosis in rats: a new model of hepatopulmonary syndrome with intestinal endotoxemia. World J Gastroenterol 2007. [PMID: 17659698 DOI: 10.3748/wjg.13.6385] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM To develop and characterize a practical model of Hepatopulmonary syndrome (HPS) in rats. METHODS The experimental animals were randomized into five feeding groups: (1) control (fed standard diet), (2) control plus intraperitoneal injection with lipopolysaccharide (LPS), (3) cirrhosis (fed a diet of maize flour, lard, cholesterol, and alcohol plus subcutaneously injection with carbon tetrachloride (CCl(4)) oil solution), (4) cirrhosis plus LPS, and (5) cirrhosis plus glycine and LPS. The blood, liver and lung tissues of rats were sampled for analysis and characterization. Technetium 99m-labeled macroaggregated albumin (Tc99m-MAA) was used to test the dilatation of pulmonary microvasculature. RESULTS Typical cirrhosis and subsequent hepato-pulmonary syndrome was observed in the cirrhosis groups after an 8 wk feeding period. In rats with cirrhosis, there were a decreased PaO(2) and PaCO(2) in arterial blood, markedly decreased arterial O(2) content, a significantly increased alveolar to arterial oxygen gradient, an increased number of bacterial translocated within mesenteric lymph node, a significant higher level of LPS and tumor necrosis factor-alpha (TNF-alpha) in plasma, and a significant greater ratio of Tc99m-MAA brain-over-lung radioactivity. After LPS administration in rats with cirrhosis, various pathological parameters got worse and pulmonary edema formed. The predisposition of glycine antagonized the effects of LPS and significantly alleviated various pathological alterations. CONCLUSION The results suggest that: (1) a characteristic rat model of HPS can be non-invasively induced by multiple pathogenic factors including high fat diet, alcohol, cholesterol and CCl(4); (2) this model can be used for study of hepatopulmonary syndrome and is clinically relevant; and (3) intestinal endotoxemia (IETM) and its accompanying cytokines, such as TNF-alpha, exert a crucial role in the pathogenesis of HPS in this model.
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Affiliation(s)
- Hui-Ying Zhang
- Director of Institute of Hepatology, Shanxi Medical University, 86 Xin-jian Nan Road, Taiyuan 030001, Shanxi Province, China
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Zhang HY, Han DW, Zhao ZF, Liu MS, Wu YJ, Chen XM, Ji C. Multiple pathogenic factor-induced complications of cirrhosis in rats: A new model of hepatopulmonary syndrome with intestinal endotoxemia. World J Gastroenterol 2007; 13:3500-7. [PMID: 17659698 PMCID: PMC4146787 DOI: 10.3748/wjg.v13.i25.3500] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To develop and characterize a practical model of Hepatopulmonary syndrome (HPS) in rats.
METHODS: The experimental animals were randomized into five feeding groups: (1) control (fed standard diet), (2) control plus intraperitoneal injection with lipopolysaccharide (LPS), (3) cirrhosis (fed a diet of maize flour, lard, cholesterol, and alcohol plus subcutaneously injection with carbon tetrachloride (CCl4) oil solution), (4) cirrhosis plus LPS, and (5) cirrhosis plus glycine and LPS. The blood, liver and lung tissues of rats were sampled for analysis and characterization. Technetium 99m-labeled macroaggregated albumin (Tc99m-MAA) was used to test the dilatation of pulmonary microvasculature.
RESULTS: Typical cirrhosis and subsequent hepato-pulmonary syndrome was observed in the cirrhosis groups after an 8 wk feeding period. In rats with cirrhosis, there were a decreased PaO2 and PaCO2 in arterial blood, markedly decreased arterial O2 content, a significantly increased alveolar to arterial oxygen gradient, an increased number of bacterial translocated within mesenteric lymph node, a significant higher level of LPS and tumor necrosis factor-α (TNF-α) in plasma, and a significant greater ratio of Tc99m-MAA brain-over-lung radioactivity. After LPS administration in rats with cirrhosis, various pathological parameters got worse and pulmonary edema formed. The predisposition of glycine antagonized the effects of LPS and significantly alleviated various pathological alterations.
CONCLUSION: The results suggest that: (1) a characte-ristic rat model of HPS can be non-invasively induced by multiple pathogenic factors including high fat diet, alcohol, cholesterol and CCl4; (2) this model can be used for study of hepatopulmonary syndrome and is clinically relevant; and (3) intestinal endotoxemia (IETM) and its accompanying cytokines, such as TNF-α, exert a crucial role in the pathogenesis of HPS in this model.
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Affiliation(s)
- Hui-Ying Zhang
- Director of Institute of Hepatology, Shanxi Medical University, 86 Xin-jian Nan Road, Taiyuan 030001, Shanxi Province, China
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Louis NA, Hamilton KE, Canny G, Shekels LL, Ho SB, Colgan SP. Selective induction of mucin-3 by hypoxia in intestinal epithelia. J Cell Biochem 2007; 99:1616-27. [PMID: 16823775 DOI: 10.1002/jcb.20947] [Citation(s) in RCA: 108] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/19/2023]
Abstract
Epithelial cells line mucosal surfaces (e.g., lung, intestine) and critically function as a semipermeable barrier to the outside world. Mucosal organs are highly vascular with extensive metabolic demands, and for this reason, are particularly susceptible to diminished blood flow and resultant tissue hypoxia. Here, we pursue the hypothesis that intestinal barrier function is regulated in a protective manner by hypoxia responsive genes. We demonstrate by PCR confirmation of microarray data and by avidin blotting of immunoprecipitated human Mucin 3 (MUC3), that surface MUC3 expression is induced in T84 intestinal epithelial cells following exposure to hypoxia. MUC3 RNA is minimally detectable while surface protein expression is absent under baseline normoxic conditions. There is a robust induction in both the mRNA (first evident by 8 h) and protein expression, first observed and maximally expressed following 24 h hypoxia. This is followed by a subsequent decline in protein expression, which remains well above baseline at 48 h of hypoxia. Further, we demonstrate that this induction of MUC3 protein is associated with a transient increase in the barrier restorative peptide, intestinal trefoil factor (ITF). ITF not only colocalizes with MUC3, by confocal microscopy, to the apical surface of T84 cells following exposure to hypoxia, but is also found, by co-immunoprecipitation, to be physically associated with MUC3, following 24 h of hypoxia. In exploration of the mechanism of hypoxic regulation of mucin 3 expression, we demonstrated by luciferase assay that the full-length promoter for mouse Mucin 3 (Muc3) is hypoxia-responsive with a 5.08 +/- 1.76-fold induction following 24 h of hypoxia. Furthermore, analysis of both the human (MUC3A) and mouse (Muc3) promoters revealed potential HIF-1 binding sites which were shown by chromatin immunoprecipitation to bind the pivotal hypoxia-regulating transcription factor HIF-1alpha. Taken together, these studies implicate the HIF-1alpha mediated hypoxic induced expression of mucin 3 and associated ITF in the maintenance of intestinal barrier function under hypoxic conditions.
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Affiliation(s)
- Nancy A Louis
- Center for Experimental Therapeutics and Reperfusion Injury, Brigham and Women's Hospital, 75 Francis Street, Boston, MA 02115, USA.
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Colgan SP, Eltzschig HK, Eckle T, Thompson LF. Physiological roles for ecto-5'-nucleotidase (CD73). Purinergic Signal 2006; 2:351-60. [PMID: 18404475 PMCID: PMC2254482 DOI: 10.1007/s11302-005-5302-5] [Citation(s) in RCA: 398] [Impact Index Per Article: 20.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/13/2005] [Revised: 11/01/2005] [Accepted: 11/02/2005] [Indexed: 02/07/2023] Open
Abstract
Nucleotides and nucleosides influence nearly every aspect of physiology and pathophysiology. Extracellular nucleotides are metabolized through regulated phosphohydrolysis by a series of ecto-nucleotidases. The formation of extracellular adenosine from adenosine 5’-monophosphate is accomplished primarily through ecto-5’-nucleotidase (CD73), a glycosyl phosphatidylinositol-linked membrane protein found on the surface of a variety of cell types. Recent in vivo studies implicating CD73 in a number of tissue protective mechanisms have provided new insight into its regulation and function and have generated considerable interest. Here, we review contributions of CD73 to cell and tissue stress responses, with a particular emphasis on physiologic responses to regulated CD73 expression and function, as well as new findings utilizing Cd73-deficient animals.
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Affiliation(s)
- Sean P Colgan
- Center for Experimental Therapeutics, Brigham and Women’s Hospital, Harvard Medical School, Thorn Building 704, 75 Francis Street, Boston, Massachusetts, 02115, USA,
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Ibla JC, Khoury J, Kong T, Robinson A, Colgan SP. Transcriptional repression of Na-K-2Cl cotransporter NKCC1 by hypoxia-inducible factor-1. Am J Physiol Cell Physiol 2006; 291:C282-9. [PMID: 16571862 DOI: 10.1152/ajpcell.00564.2005] [Citation(s) in RCA: 29] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/30/2023]
Abstract
Tissue edema is commonly associated with hypoxia. Generally, such episodes of fluid accumulation are self-limiting. At present, little is known about mechanisms to compensate excessive fluid transport. Here we describe an adaptive mechanism to dampen fluid loss during hypoxia. Initial studies confirmed previous observations of attenuated electrogenic Cl- secretion after epithelial hypoxia. A screen of known ion transporters in Cl- -secreting epithelia revealed selective downregulation of Na-K-2Cl cotransporter NKCC1 mRNA, protein, and function. Subsequent studies identified transcriptional repression of NKCC1 mediated by hypoxia-inducible factor (HIF). Chromatin immunoprecipitation analysis identified a functional HIF binding site oriented on the antisense strand of genomic DNA downstream of the transcription start site corresponding to the NKCC1 5'-untranslated region. Additional in vivo studies using conditional Hif1a-null mice revealed that the loss of HIF-1alpha in Cl- -secreting epithelia results in a loss of NKCC1 repression. These studies describe a novel regulatory pathway for NKCC1 transcriptional repression by hypoxia. These results suggest that HIF-dependent repression of epithelial NKCC1 may provide a compensatory mechanism to prevent excessive fluid loss during hypoxia.
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Affiliation(s)
- Juan C Ibla
- Department of Anesthesia Perioperative and Pain Medicine, Children's Hospital, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA
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Louis NA, Hamilton KE, Kong T, Colgan SP. HIF-dependent induction of apical CD55 coordinates epithelial clearance of neutrophils. FASEB J 2006; 19:950-9. [PMID: 15923405 DOI: 10.1096/fj.04-3251com] [Citation(s) in RCA: 63] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/11/2022]
Abstract
Sites of inflammation are associated with dramatic shifts in tissue metabolism. Inflammation can result in significant tissue hypoxia, with resultant induction of hypoxia-responsive genes. Given this association, we hypothesized that neutrophil (PMN) ligands expressed on epithelial cells may be regulated by hypoxia. Initial studies confirmed earlier results that epithelial hypoxia enhances PMN transepithelial migration and promotes apical clearance of PMN from the epithelial surface. A screen of known PMN ligands revealed a surprisingly stable expression pattern in hypoxia. However, this screen identified one gene, CD55, as a highly hypoxia-inducible molecule expressed on the apical membrane of mucosal epithelia. Subsequent studies verified the induction of CD55 mRNA and protein expression by hypoxia. Overexpression of CD55 by transfection in nonhypoxic epithelia resulted in a similar pattern of apical PMN clearance, and peptide mimetics corresponding to the PMN binding site on DAF blocked such apical clearance of PMN. Studies directed at understanding molecular pathways of hypoxia inducibility revealed that a approximately 200 bp region of the CD55 gene conferred hypoxia inducibility for CD55. These studies identified a functional binding site for the transcriptional regulator hypoxia-inducible factor (HIF). Taken together, these results identify HIF-dependent induction of epithelial CD55 in the resolution of ongoing inflammation through clearance of apical PMN.
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Affiliation(s)
- Nancy A Louis
- Neonatology Division, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA
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Karhausen J, Kong T, Narravula S, Colgan SP. Induction of the von Hippel-Lindau tumor suppressor gene by late hypoxia limits HIF-1 expression. J Cell Biochem 2005; 95:1264-75. [PMID: 15962286 DOI: 10.1002/jcb.20489] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/11/2022]
Abstract
Hypoxia-inducible factor (HIF) remains the central focus of oxygen sensing during hypoxia. HIF is a heterodimeric transcription factor consisting of an oxygen-regulated alpha- and a constitutively expressed beta subunit. The von Hippel-Lindau tumor suppressor (pVHL) is a component of the E3 ubiquitin ligase complex and targets HIF-alpha to proteasomal degradation, but also is known to exert a significant control on HIF transactivation activity. However, the understanding of the full interaction between HIF and pVHL has been hindered by a lack in the understanding of pVHL regulation. Here, we report that pVHL itself is induced in prolonged hypoxia in a kinetic that parallels the observed downregulation of HIF-1alpha protein under such conditions. In addition, we document direct HIF-1alpha binding to the VHL promoter and identify a functional hypoxia response element (HRE) within the VHL promoter. Such induction of pVHL in hypoxia furthermore has functional implications for the HIF dependent hypoxic response, implicating a physiologically relevant feedback mechanism. These results provide an intriguing model, whereby HIF self-regulates expression through VHL and highlight the role of pVHL as a unifying mechanism of HIF regulation.
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Affiliation(s)
- Jörn Karhausen
- Center for Experimental Therapeutics and Reperfusion Injury, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA
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Dynamic purine signaling and metabolism during neutrophil-endothelial interactions. Purinergic Signal 2005; 1:229-39. [PMID: 18404508 PMCID: PMC2096542 DOI: 10.1007/s11302-005-6323-9] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/14/2004] [Revised: 03/07/2005] [Accepted: 03/23/2005] [Indexed: 01/09/2023] Open
Abstract
During episodes of hypoxia and inflammation, polymorphonuclear leukocytes (PMN) move into underlying tissues by initially passing between endothelial cells that line the inner surface of blood vessels (transendothelial migration, TEM). TEM creates the potential for disturbances in vascular barrier and concomitant loss of extravascular fluid and resultant edema. Recent studies have demonstrated a crucial role for nucleotide metabolism and nucleoside signaling during inflammation. These studies have implicated multiple adenine nucleotides as endogenous tissue protective mechanisms invivo. Here, we review the functional components of vascular barrier, identify strategies for increasing nucleotide generation and nucleoside signaling, and discuss potential therapeutic targets to regulate the vascular barrier during inflammation.
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Eltzschig HK, Thompson LF, Karhausen J, Cotta RJ, Ibla JC, Robson SC, Colgan SP. Endogenous adenosine produced during hypoxia attenuates neutrophil accumulation: coordination by extracellular nucleotide metabolism. Blood 2004; 104:3986-92. [PMID: 15319286 DOI: 10.1182/blood-2004-06-2066] [Citation(s) in RCA: 282] [Impact Index Per Article: 13.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Hypoxia is a well-documented inflammatory stimulus and results in tissue polymorphonuclear leukocyte (PMN) accumulation. Likewise, increased tissue adenosine levels are commonly associated with hypoxia, and given the anti-inflammatory properties of adenosine, we hypothesized that adenosine production via adenine nucleotide metabolism at the vascular surface triggers an endogenous anti-inflammatory response during hypoxia. Initial in vitro studies indicated that endogenously generated adenosine, through activation of PMN adenosine A(2A) and A(2B) receptors, functions as an antiadhesive signal for PMN binding to microvascular endothelia. Intravascular nucleotides released by inflammatory cells undergo phosphohydrolysis via hypoxia-induced CD39 ectoapyrase (CD39 converts adenosine triphosphate/adenosine diphosphate [ATP/ADP] to adenosine monophosphate [AMP]) and CD73 ecto-5'-nucleotidase (CD73 converts AMP to adenosine). Extensions of our in vitro findings using cd39- and cd73-null animals revealed that extracellular adenosine produced through adenine nucleotide metabolism during hypoxia is a potent anti-inflammatory signal for PMNs in vivo. These findings identify CD39 and CD73 as critical control points for endogenous adenosine generation and implicate this pathway as an innate mechanism to attenuate excessive tissue PMN accumulation.
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Affiliation(s)
- Holger K Eltzschig
- Brigham and Women's Hospital, Harvard Medical School, Thorn Building 704, 75 Francis Street, Boston, MA 02115, USA
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van de Wetering MD, Caron HN, Biezeveld M, Taminiau JAJM, ten Kate FJ, Spanjaard L, Kuijpers TW. Severity of enterocolitis is predicted by IL-8 in paediatric oncology patients. Eur J Cancer 2004; 40:571-8. [PMID: 14962725 DOI: 10.1016/j.ejca.2003.10.021] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/08/2003] [Revised: 09/15/2003] [Accepted: 10/31/2003] [Indexed: 11/20/2022]
Abstract
Enterocolitis in oncology patients remains an important complication, but there is a lack of insight into its likely severity from microbial, pathological and inflammatory aspects. Paediatric oncology patients admitted with neutropenic fever, who developed abdominal pain and diarrhoea, were monitored by the takers of rectal biopsies, cultures, and inflammatory marker measurements. Twenty-five patients were included (mean age 7.1 years). 8 patients (32%) needed intensive care treatment, 3 (12%) patients died. Gram-positive bacteraemia was diagnosed in 4 patients (16%). Most patients had negative blood and stool cultures. Predictors of a severe clinical course of the enterocolitis were an increased serum interleukin-8 (IL-8) (>1000 pg/ml) level and an increased serum C-reactive protein level (CRP) (>150 mg/l) level, both measured on the first day of clinical illness. Relative risks (RR) for admission to an Intensive Care Unit (ICU) were 11.3 (95% Confidence Interval (CI) 1.6-77.9) for elevated IL-8 levels and 6.4 (95% (CI) 0.92-45.1) for increased CRP levels. Rectal biopsies and pathology could not predict outcome (P=0.22). IL-8 analysis at the onset of enterocolitis symptoms can identify high-risk patients, which might be used clinically to design future intervention trials.
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Affiliation(s)
- M D van de Wetering
- Emma Children's Hospital, Academic Medical Center, F8-245, Meibergdreef 9, 1105 AZ Amsterdam, The Netherlands.
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Abstract
Although restoration of blood flow to an ischaemic organ is essential to prevent irreversible tissue injury, reperfusion per se may result in a local and systemic inflammatory response that may augment tissue injury in excess of that produced by ischaemia alone. Cellular damage after reperfusion of previously viable ischaemic tissues is defined as ischaemia-reperfusion (I-R) injury. I-R injury is characterized by oxidant production, complement activation, leucocyte-endothelial cell adhesion, platelet-leucocyte aggregation, increased microvascular permeability and decreased endothelium-dependent relaxation. In its severest form, I-R injury can lead to multiorgan dysfunction or death. Although our understanding of the pathophysiology of I-R injury has advanced significantly in the last decade, such experimentally derived concepts have yet to be fully integrated into clinical practice. Treatment of I-R injury is also confounded by the fact that inhibition of I-R-associated inflammation might disrupt protective physiological responses or result in immunosuppression. Thus, while timely reperfusion of the ischaemic area at risk remains the cornerstone of clinical practice, therapeutic strategies such as ischaemic preconditioning, controlled reperfusion, and anti-oxidant, complement or neutrophil therapy may significantly prevent or limit I-R-induced injury in humans.
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Affiliation(s)
- Holger K Eltzschig
- Clinical Associate Department of Cardiovascular Anesthesia, Texas Heart Institute, St. Luke's Episcopal Hospital, 6720 Bertner Avenue, Room 0520, MC1-226, Houston, TX 77030, USA
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Lawrence DW, Bruyninckx WJ, Louis NA, Lublin DM, Stahl GL, Parkos CA, Colgan SP. Antiadhesive role of apical decay-accelerating factor (CD55) in human neutrophil transmigration across mucosal epithelia. ACTA ACUST UNITED AC 2003; 198:999-1010. [PMID: 14530374 PMCID: PMC2194216 DOI: 10.1084/jem.20030380] [Citation(s) in RCA: 68] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
Neutrophil migration across mucosal epithelium during inflammatory episodes involves the precise orchestration of a number a cell surface molecules and signaling pathways. After successful migration to the apical epithelial surface, apically localized epithelial proteins may serve to retain PMN at the lumenal surface. At present, identification of apical epithelial ligands and their PMN counter-receptors remain elusive. Therefore, to define the existence of apical epithelial cell surface proteins involved in PMN–epithelial interactions, we screened a panel of antibodies directed against epithelial plasma membranes. This strategy identified one antibody (OE-1) that both localized to the apical cell membrane and significantly inhibited PMN transmigration across epithelial monolayers. Microsequence analysis revealed that OE-1 recognized human decay-accelerating factor (DAF, CD55). DAF is a highly glycosylated, 70–80-kD, glycosyl-phosphatidyinositol–linked protein that functions predominantly as an inhibitor of autologous complement lysis. DAF suppression experiments using antisense oligonucleotides or RNA interference revealed that DAF may function as an antiadhesive molecule promoting the release of PMN from the lumenal surface after transmigration. Similarly, peptides corresponding to the antigen recognition domain of OE-1 resulted in accumulation of PMN on the apical epithelial surface. The elucidation of DAF as an apical epithelial ligand for PMN provides a target for novel anti-inflammatory therapies directed at quelling unwanted inflammatory episodes.
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Affiliation(s)
- Donald W Lawrence
- Center for Experimental Therapeutics and Reperfusion Injury, Brigham and Women's Hospital, Harvard Medical School, 20 Shattuck St., Boston, MA 02115, USA
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Abstract
As a 'double-edged sword', neutrophil (polymorphonuclear leukocyte) migration across epithelial-lined organs is an important component of host defense, but it also results in epithelial pathophysiology and disease symptoms. There have been significant advances in better understanding the mechanisms of how leukocytes cross the vascular endothelium to exit the bloodstream; however, many of the mechanisms that govern polymorphonuclear leukocyte transepithelial migration are different and we are only just beginning to understand them. Recent findings include new junctional adhesion molecules and carbohydrate moieties as receptors for migrating neutrophils. In addition, new insights into leukocyte-epithelial signaling events have emerged that are beginning to shed light on the role of SIRP-CD47 interactions in regulating the rate of neutrophil transepithelial migration and how neutrophils modulate epithelial barrier function.
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Affiliation(s)
- Ke Zen
- Division of Gastrointestinal Pathology, Department of Pathology and Laboratory Medicine, Emory University, Whitehead Research Boulevard, Room 1053, 615 Michael Street, Atlanta, GA 30322, USA
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Crossley LJ. Neutrophil activation by fMLP regulates FOXO (forkhead) transcription factors by multiple pathways, one of which includes the binding of FOXO to the survival factor Mcl-1. J Leukoc Biol 2003; 74:583-92. [PMID: 12960271 DOI: 10.1189/jlb.0103020] [Citation(s) in RCA: 27] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/22/2023] Open
Abstract
Activation signals from bacterial stimuli set into motion a series of events that alter the abbreviated lifespan of neutrophils. These studies show that the bacterial chemoattractant, formyl-Met-Leu-Phe (fMLP), promotes the phosphorylation/inactivation of the FOXO subfamily of forkhead transcription factors (FKHR, FKHR-L1, and AFX) through the phosphatidylinositol-3-kinase/Akt (protein kinase B) and the RAS mitogen-activated protein kinase pathways. Furthermore, fMLP stimulation causes the inducible expression of the prosurvival Bcl-2 family member Mcl-1, which then binds to a complex containing FKHR. These studies show that fMLP-stimulated neutrophils coordinate the regulation of FOXO transcription factors and the survival factor Mcl-1, a mechanism that may allow neutrophils to alter their survival.
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Affiliation(s)
- Lisa J Crossley
- Department of Anesthesiology, Perioperative and Pain Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA.
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Eltzschig HK, Ibla JC, Furuta GT, Leonard MO, Jacobson KA, Enjyoji K, Robson SC, Colgan SP. Coordinated adenine nucleotide phosphohydrolysis and nucleoside signaling in posthypoxic endothelium: role of ectonucleotidases and adenosine A2B receptors. J Exp Med 2003; 198:783-96. [PMID: 12939345 PMCID: PMC2194189 DOI: 10.1084/jem.20030891] [Citation(s) in RCA: 411] [Impact Index Per Article: 18.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/08/2022] Open
Abstract
Limited oxygen delivery to tissues (hypoxia) is common in a variety of disease states. A number of parallels exist between hypoxia and acute inflammation, including the observation that both influence vascular permeability. As such, we compared the functional influence of activated polymorphonuclear leukocytes (PMN) on normoxic and posthypoxic endothelial cells. Initial studies indicated that activated PMN preferentially promote endothelial barrier function in posthypoxic endothelial cells (>60% increase over normoxia). Extension of these findings identified at least one soluble mediator as extracellular adenosine triphosphate (ATP). Subsequent studies revealed that ATP is coordinately hydrolyzed to adenosine at the endothelial cell surface by hypoxia-induced CD39 and CD73 (>20-and >12-fold increase in mRNA, respectively). Studies in vitro and in cd39-null mice identified these surface ecto-enzymes as critical control points for posthypoxia-associated protection of vascular permeability. Furthermore, insight gained through microarray analysis revealed that the adenosine A2B receptor (AdoRA2B) is selectively up-regulated by hypoxia (>5-fold increase in mRNA), and that AdoRA2B antagonists effectively neutralize ATP-mediated changes in posthypoxic endothelial permeability. Taken together, these results demonstrate transcription coordination of adenine nucleotide and nucleoside signaling at the vascular interface during hypoxia.
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Affiliation(s)
- Holger K Eltzschig
- Center for Experimental Therapeutics and Reperfusion Injury, Brigham and Women's Hospital, Harvard Medical School, 75 Francis Street, Boston, MA 02115, USA
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