Cigarette smoke (CS) is the primary cause of chronic obstructive pulmonary disease (COPD), an effect that is, in part, due to intense oxidant stress. Clearance of apoptotic cells (efferocytosis) is a ...critical regulator of lung homeostasis, which is defective in smokers and in patients with COPD, suggesting a role in disease pathogenesis.
We hypothesized that CS would impair efferocytosis through oxidant-dependent activation of RhoA, a known inhibitor of this process.
We investigated the effect of CS on efferocytosis in vivo and ex vivo, using acute, subacute, and long-term mouse exposure models.
Acute and subacute CS exposure suppressed efferocytosis by alveolar macrophages in a dose-dependent, reversible, and cell type-independent manner, whereas more intense CS exposure had an irreversible effect. In contrast, CS did not alter ingestion through the Fc gamma receptor. The inhibitory effect of CS on apoptotic cell clearance depended on oxidants, because the effect was blunted in oxidant-resistant ICR mice, and was prevented by either genetic or pharmacologic antioxidant strategies in vivo and ex vivo. CS inhibited efferocytosis through oxidant-dependent activation of the RhoA-Rho kinase pathway because (1) CS activated RhoA, (2) antioxidants prevented RhoA activation by CS, and (3) inhibitors of the RhoA-Rho kinase pathway reversed the suppressive effect of CS on apoptotic cell clearance in vivo and ex vivo.
These findings advance the hypothesis that impaired efferocytosis may contribute to the pathogenesis of COPD and suggest the therapeutic potential of drugs targeting the RhoA-Rho kinase pathway.
1 Division of Pulmonary Sciences and Critical Care Medicine, Department of Medicine, University of Colorado Denver, Aurora; ;
2 Department of Pediatrics, National Jewish Health, Denver, Colorado;
3 ...Department of Medicine, National Jewish Health, Denver, Colorado; and ;
4 Toronto General Hospital Research Institute of the University Health Network, Division of Respirology, Department of Medicine, University of Toronto, Toronto, Ontario, Canada
Submitted 28 January 2009
; accepted in final form 20 July 2009
Cystic fibrosis (CF) is caused by mutated CF transmembrane conductance regulator (CFTR) and is characterized by robust airway inflammation and accumulation of apoptotic cells. Phagocytosis of apoptotic cells (efferocytosis) is a pivotal regulator of inflammation, because it prevents postapoptotic necrosis and actively suppresses release of a variety of proinflammatory mediators, including IL-8. Because CF is associated with accumulation of apoptotic cells, inappropriate levels of IL-8, and robust inflammation, we sought to determine whether CFTR deficiency specifically impairs efferocytosis and its regulation of inflammatory mediator release. Here we show that CFTR deficiency directly interferes with efferocytosis by airway epithelium, an effect that is not due to altered binding of apoptotic cells to epithelial cells or altered expression of efferocytosis receptors. In contrast, expression of RhoA, a known negative regulator of efferocytosis, is substantially increased in CFTR-deficient cells, and inhibitors of RhoA or its downstream effector Rho kinase normalize efferocytosis in these cells. Impaired efferocytosis appears to be mediated through an amiloride-sensitive ion channel, because amiloride restores phagocytic competency in CFTR-deficient cells. Finally, ineffective efferocytosis in CFTR-deficient cells appears to have proinflammatory consequences, because apoptotic cells enhance IL-8 release by these cells, but not by wild-type controls. Therefore, in CF, dysregulated efferocytosis may lead to accumulation of apoptotic cells and impaired regulation of the inflammatory response and, ultimately, may suggest a new therapeutic target.
efferocytosis; Rho GTPase; inflammation; epithelial cells
Address for reprint requests and other correspondence: R. W. Vandivier, Division of Pulmonary Sciences and Critical Care Medicine, Univ. of Colorado Denver, Research Bldg. 2, 12700 E. 19th Ave. Box C272, Aurora, CO 80045 (e-mail: Bill.Vandivier{at}ucdenver.edu ).
Laboratory-based reporting of hospital-onset (HO) Methicillin-Resistant Staphylococcus aureus (MRSA) bacteremia is used by regulatory agencies as a method to decrease the incidence of MRSA bacteremia ...in acute care settings. Although decolonization, hand hygiene, and isolation strategies are used, many facilities continue to have high rates of HO-MRSA. To better understand the potential risk factors and underlying causes of patients developing a HO-MRSA bacteremia, a chart review of patients that developed a HO Staphylococcus aureus (SA) and meeting defined criteria was reviewed to identify potential strategies towards prevention efforts.
Laboratory data was pulled from January 2016 – July 2017. Data included in the export were all positive blood cultures that grew MSSA and MRSA. The data was further filtered to only include first incident blood cultures growing MSSA or MRSA that occurred on or after the fourth day of hospitalization. Chart review was performed to identify risk factors and clinical syndromes that could potentially lead to a HO-SA bacteremia.
We identified 130 inpatients with HO-SA bacteremia, of which 63 (48%) was HO-MRSA. The most common sources identified for patients with these bacteremias were pneumonia (27%), vascular devices (20%), and skin/soft tissue infections (23%). There was no significant difference in the distribution of these sources comparing HO-MRSA with HO-MSSA. A total of 42 (32%) patients were identified to have signs and symptoms consistent with infection on admission but did not have a blood culture collected on or after day 4 of admission. Of these 42 patients, 33 (79%) were admitted from the ED and 20 (48%) are immunosuppressed patients.
Pneumonia was the most common cause of HO-SA bacteremia. Strategies to prevent HO-MRSA should include processes to prevent hospital acquired pneumonia with prompt blood culture collection on admission in patients presenting with compatable signs and symptoms.
Efficient clearance of apoptotic cells from the lung by alveolar macrophages is important for the maintenance of tissue structure and function. Lung tissue from humans with emphysema contains ...increased numbers of apoptotic cells and decreased levels of vascular endothelial growth factor (VEGF). Mice treated with VEGF receptor inhibitors have increased numbers of apoptotic cells and develop emphysema. We hypothesized that VEGF regulates apoptotic cell clearance by alveolar macrophages (AM) via its interaction with VEGF receptor 1 (VEGF R1). Our data show that the uptake of apoptotic cells by murine AMs and human monocyte-derived macrophages is inhibited by depletion of VEGF and that VEGF activates Rac1. Antibody blockade or pharmacological inhibition of VEGF R1 activity also decreased apoptotic cell uptake ex vivo. Conversely, overexpression of VEGF significantly enhanced apoptotic cell uptake by AMs in vivo. These results indicate that VEGF serves a positive regulatory role via its interaction with VEGF R1 to activate Rac1 and enhance AM apoptotic cell clearance.
Background: Alcohol abuse increases the risk for acute respiratory distress syndrome (ARDS). Efferocytosis, the clearance of apoptotic cells, is important in the resolution of inflammation and is ...regulated by RhoA and rho kinase (ROCK) activation. The effects of alcohol on pulmonary Rho pathway activation and efferocytosis have not been determined. We hypothesize that acute and chronic alcohol exposure impair pulmonary efferocytosis, leading to heightened inflammation during ARDS.
Methods: For in vivo experiments, C57BL/6 mice received either a single intraperitoneal injection of alcohol or chronic ethanol‐in‐water for 8 weeks prior to intratracheal instillation of apoptotic cells or lipopolysaccharide (LPS). Bronchoalveolar lavage (BAL) was performed for cells counts, calculation of the phagocytic index (PI), and Rho activity measurements. For in vitro studies, primary alveolar macrophages were cultured in alcohol (25–100 mM) and then co‐cultured with apoptotic cells. RhoA activity was determined following alcohol exposure, and the PI was determined before and after treatment with the ROCK inhibitor, Y27632.
Results: Acute alcohol exposure was associated with impaired efferocytosis. Following LPS exposure, acute alcohol exposure was also associated with increased BAL neutrophils. Chronic alcohol exposure alone did not alter efferocytosis. However, following exposure to LPS, chronic alcohol exposure was associated with both impaired efferocytosis and increased BAL neutrophils. In vitro alcohol exposure caused a dose‐dependent decrease in efferocytosis. Despite the fact that RhoA activity was decreased by alcohol exposure and RhoA inhibition did not alter the effects of alcohol on efferocytosis, treatment with the Rho kinase inhibitor, Y27632, reversed the effects of alcohol on efferocytosis.
Conclusions: Acute alcohol exposure impairs pulmonary efferocytosis, whereas exposure to chronic alcohol is only associated with impaired efferocytosis following LPS‐induced lung injury. Both forms of alcohol exposure are associated with increased alveolar neutrophil numbers in response to LPS. The acute effects of alcohol on efferocytosis appear to be mediated, at least in part, by RhoA‐independent activation of ROCK. Further studies are needed to dissect the differences between the effects of acute and chronic alcohol exposure on efferocytosis and to determine the effects of alcohol on alternative activators of ROCK.
We developed a 1-hour field enzyme immunoassay (EIA) for detecting antibody to Sin Nombre virus in deer mice (Peromyscus maniculatus). The assay specificity and sensitivity were comparable to those ...of a standard EIA. This test will permit identification of rodents with antibody to this and perhaps other hantaviruses.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, ODKLJ, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
We developed a 1-hour field enzyme immunoassay (EIA) for detecting antibody to Sin Nombre virus in deer mice (
Peromyscus maniculatus
). The assay specificity and sensitivity were comparable to those ...of a standard EIA. This test will permit identification of rodents with antibody to this and perhaps other hantaviruses.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, ODKLJ, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Efficient removal of apoptotic cells is essential for resolution of inflammation. Failure to clear dying cells can exacerbate lung injury and lead to persistent inflammation and autoimmunity. Here we ...show that TNFα blocks apoptotic cell clearance by alveolar macrophages and leads to proinflammatory responses in the lung. Compared with mice treated with intratracheal TNFα or exogenous apoptotic cells, mice treated with the combination of TNFα plus apoptotic cells demonstrated reduced apoptotic cell clearance from the lungs and increased recruitment of inflammatory leukocytes to the air spaces. Treatment with intratracheal TNFα had no effect on the removal of exogenous apoptotic cells from the lungs of TNFα receptor-1 (p55) and -2 (p75) double mutant mice and no effect on leukocyte recruitment. Bronchoalveolar lavage from mice treated with TNFα plus apoptotic cells contained increased levels of proinflammatory cytokines IL-6, KC, and MCP-1, but exhibited no change in levels of anti-inflammatory cytokines IL-10 and TGF-β. Administration of TNFα plus apoptotic cells during LPS-induced lung injury augmented neutrophil accumulation and proinflammatory cytokine production. These findings suggest that the presence of TNFα in the lung can alter the response of phagocytes to apoptotic cells leading to inflammatory cell recruitment and proinflammatory mediator production.
1 Program in Cell Biology, Department of Pediatrics, and ;
2 Department of Medicine, National Jewish Health, and ;
3 Division of Pulmonary Sciences and Critical Care Medicine, Department of Medicine, ...University of Colorado Health Sciences Center, Denver, Colorado; ;
4 Centro de Pesquisa Gonçalo Moniz Fiocruz, Salvador, Brazil; and ;
5 Department of Clinical Medicine, Institute of Tropical Medicine, Nagasaki University, Nagasaki, Japan
Submitted 19 November 2008
; accepted in final form 28 July 2009
Efficient removal of apoptotic cells is essential for resolution of inflammation. Failure to clear dying cells can exacerbate lung injury and lead to persistent inflammation and autoimmunity. Here we show that TNF blocks apoptotic cell clearance by alveolar macrophages and leads to proinflammatory responses in the lung. Compared with mice treated with intratracheal TNF or exogenous apoptotic cells, mice treated with the combination of TNF plus apoptotic cells demonstrated reduced apoptotic cell clearance from the lungs and increased recruitment of inflammatory leukocytes to the air spaces. Treatment with intratracheal TNF had no effect on the removal of exogenous apoptotic cells from the lungs of TNF receptor-1 (p55) and -2 (p75) double mutant mice and no effect on leukocyte recruitment. Bronchoalveolar lavage from mice treated with TNF plus apoptotic cells contained increased levels of proinflammatory cytokines IL-6, KC, and MCP-1, but exhibited no change in levels of anti-inflammatory cytokines IL-10 and TGF-β. Administration of TNF plus apoptotic cells during LPS-induced lung injury augmented neutrophil accumulation and proinflammatory cytokine production. These findings suggest that the presence of TNF in the lung can alter the response of phagocytes to apoptotic cells leading to inflammatory cell recruitment and proinflammatory mediator production.
apoptosis; phagocytosis
Address for reprint requests and other correspondence: W. J. Janssen, Dept. of Medicine, K729B, National Jewish Health, Denver, CO 80206 (e-mail: janssenw{at}njc.org ).
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