title: Nicotine stimulates nerve growth factor in lung fibroblasts through an NFκB-dependent mechanism.
abstract: BACKGROUND
  Airway hyperresponsiveness (AHR) is classically found in asthma, and persistent AHR is associated with poor asthma control. Although airway smooth muscle (ASM) cells play a critical pathophysiologic role in AHR, the paracrine contributions of surrounding cells such as fibroblasts to the contractile phenotype of ASM cells have not been examined fully. This study addresses the hypothesis that nicotine promotes a contractile ASM cell phenotype by stimulating fibroblasts to increase nerve growth factor (NGF) secretion into the environment.
  
  
  METHODS
  Primary lung fibroblasts isolated from wild type and α7 nicotinic acetylcholine receptor (α7 nAChR) deficient mice were treated with nicotine (50 µg/ml) in vitro for 72 hours. NGF levels were measured in culture media and in bronchoalveolar lavage (BAL) fluid from asthmatic, smoking and non-smoking subjects by ELISA. The role of the NFκB pathway in nicotine-induced NGF expression was investigated by measuring NFκB nuclear translocation, transcriptional activity, chromatin immunoprecipitation assays, and si-p65 NFκB knockdown. The ability of nicotine to stimulate a fibroblast-mediated, contractile ASM cell phenotype was confirmed by examining expression of contractile proteins in ASM cells cultured with fibroblast-conditioned media or BAL fluid.
  
  
  RESULTS
  NGF levels were elevated in the bronchoalveolar lavage fluid of nicotine-exposed mice, current smokers, and asthmatic children. Nicotine increased NGF secretion in lung fibroblasts in vitro in a dose-dependent manner and stimulated NFκB nuclear translocation, p65 binding to the NGF promoter, and NFκB transcriptional activity. These responses were attenuated in α7 nAChR deficient fibroblasts and in wild type fibroblasts following NFκB inhibition. Nicotine-treated, fibroblast-conditioned media increased expression of contractile proteins in ASM cells.
  
  
  CONCLUSIONS
  Nicotine stimulates NGF release by lung fibroblasts through α7 nAChR and NFκB dependent pathways. These novel findings suggest that the nicotine-α7 nAChR-NFκB- NGF axis may provide novel therapeutic targets to attenuate tobacco smoke-induced AHR.
authors.affiliation: Atlanta VA Medical Center, Decatur, Georgia, United States of America; Division of Pulmonary, Allergy and Critical Care Medicine, Emory University School of Medicine, Atlanta, Georgia, United States of America.
authors.forename: Cherry
authors.initials: C
authors.lastname: Wongtrakool
authors.affiliation: Division of Pulmonary, Allergy and Critical Care Medicine, Emory University School of Medicine, Atlanta, Georgia, United States of America.
authors.forename: Kora
authors.initials: K
authors.lastname: Grooms
authors.affiliation: Atlanta VA Medical Center, Decatur, Georgia, United States of America; Division of Pulmonary, Allergy and Critical Care Medicine, Emory University School of Medicine, Atlanta, Georgia, United States of America.
authors.forename: Kaiser M
authors.initials: KM
authors.lastname: Bijli
authors.affiliation: University of Washington, Seattle, Washington, United States of America.
authors.forename: Kristina
authors.initials: K
authors.lastname: Crothers
authors.affiliation: Department of Pediatrics, Emory University School of Medicine, Atlanta, Georgia, United States of America.
authors.forename: Anne M
authors.initials: AM
authors.lastname: Fitzpatrick
authors.affiliation: Atlanta VA Medical Center, Decatur, Georgia, United States of America; Division of Pulmonary, Allergy and Critical Care Medicine, Emory University School of Medicine, Atlanta, Georgia, United States of America.
authors.forename: C Michael
authors.initials: CM
authors.lastname: Hart
mesh_terms: D000818:Animals; D005347:Fibroblasts / Q000187:drug effects* / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D008168:Lung / Q000166:cytology*; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism*; D020932:Nerve Growth Factor / Q000378:metabolism*; D009538:Nicotine / Q000494:pharmacology*; D064569:alpha7 Nicotinic Acetylcholine Receptor / Q000378:metabolism
chemical_list: D016328:NF-kappa B; D064569:alpha7 Nicotinic Acetylcholine Receptor; D009538:Nicotine; D020932:Nerve Growth Factor