title: Acute and chronic alcohol exposure impair the phagocytosis of apoptotic cells and enhance the pulmonary inflammatory response.
abstract: 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.
authors.affiliation: Division of Pulmonary Sciences and Critical Care Medicine, University of Colorado Denver, Anschutz Medical Campus, Research 2, Box C272, 9th Floor, 12700 East 19th Avenue, Aurora, CO 80045, USA. Darren.boe@ucdenver.edu
authors.forename: Darren M
authors.initials: DM
authors.lastname: Boé
authors.forename: Tiffany R
authors.initials: TR
authors.lastname: Richens
authors.forename: Sarah A
authors.initials: SA
authors.lastname: Horstmann
authors.forename: Ellen L
authors.initials: EL
authors.lastname: Burnham
authors.forename: William J
authors.initials: WJ
authors.lastname: Janssen
authors.forename: Peter M
authors.initials: PM
authors.lastname: Henson
authors.forename: Marc
authors.initials: M
authors.lastname: Moss
authors.forename: R William
authors.initials: RW
authors.lastname: Vandivier
mesh_terms: D000577:Amides / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D001992:Bronchoalveolar Lavage Fluid / Q000737:chemistry / Q000166:cytology; D002478:Cells, Cultured; D004195:Disease Models, Animal; D004305:Dose-Response Relationship, Drug; D004334:Drug Administration Schedule; D000431:Ethanol / Q000008:administration & dosage / Q000009:adverse effects*; D005260:Female; D008070:Lipopolysaccharides; D016676:Macrophages, Alveolar / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D010587:Phagocytosis / Q000187:drug effects* / Q000502:physiology; D011014:Pneumonia / Q000139:chemically induced / Q000503:physiopathology*; D011725:Pyridines / Q000494:pharmacology; D012128:Respiratory Distress Syndrome / Q000139:chemically induced / Q000378:metabolism / Q000503:physiopathology*; D020741:rho GTP-Binding Proteins / Q000032:analysis; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors; D020742:rhoA GTP-Binding Protein
chemical_list: D000577:Amides; D008070:Lipopolysaccharides; D011725:Pyridines; C108830:Y 27632; D000431:Ethanol; D054460:rho-Associated Kinases; C526380:RhoA protein, mouse; D020741:rho GTP-Binding Proteins; D020742:rhoA GTP-Binding Protein