Regulation of gene expression in pulmonary inflammation and differentiation : a role for C/EBP transcription factors
Author: Roos, Abraham B
Date: 2012-06-07
Location: Thoraxaulan N2:U1, Karolinska Universitetssjukhuset, Solna
Time: 09.00
Department: Inst för medicin, Solna / Dept of Medicine, Solna
Abstract
CCAAT/enhancer-binding protein (C/EBP) transcription factors play essential roles in gene regulation. The lung-enriched isoform C/EBPα is known to inhibit proliferation, promote differentiation and stimulate gene expression characteristic of the mature differentiated pulmonary epithelium. C/EBPβ, also enriched in the lung, plays a role in cell differentiation and the regulation of inflammatory and host defense genes in several organs. The activity of C/EBPβ is decreased in smokers with chronic obstructive pulmonary disease (COPD), indicating a role in COPD pathogenesis. The objective of this thesis was to investigate the unique or overlapping roles of C/EBPα and C/EBPβ in lung epithelial differentiation, and to assess the contribution of C/EBPβ in regulating pulmonary inflammation.
To investigate unique vs. overlapping roles of C/EBPα and C/EBPβ in the lung, the pulmonary phenotype of mice lacking C/EBPα (CebpaΔLE mice), C/EBPβ (CebpbΔLE mice) or both C/EBPα and C/EBPβ (CebpaΔLE; CebpbΔLE mice) specifically in the lung epithelium, all generated by SFTPC-Cre mediated excision, was investigated. Cell culture experiments suggested that C/EBPα and C/EBPβ bind the same elements within a lung-specific promoter, and that their functions are partially overlapping. Pre-natal CebpaΔLE mice and CebpaΔLE; CebpbΔLE mice displayed immature lungs similar to the lungs of premature infants, and CebpaΔLE; CebpbΔLE mice exhibited even more impaired airway epithelial cell differentiation than the CebpaΔLE mice. The proportion of CebpaΔLE mice that survived and reached adulthood spontaneously developed a majority of the histopathological hallmarks of COPD, possibly caused by infiltrating inflammatory cells – similar to what is observed in COPD and what is mechanistically proposed to drive COPD pathogenesis. These findings are indicative of a relationship between immature lungs at birth, C/EBPs and the development of inflammatory lung disease.
Considering the previous documentation of decreased airway epithelial C/EBPβ activity in smokers with COPD, C/EBPβ could have a role in COPD pathogenesis. The role of C/EBPβ in regulating inflammatory and innate immune responses in the lung was on this account investigated by employing a translational approach encompassing clinical samples as well as in vitro and in vivo experiments. CEBPB was significantly down-regulated in the airway epithelium of both current and former smokers compared to never-smokers, and in cigarette smoke extract-treated primary human airway epithelial cells in vitro, suggesting that C/EBPβ plays a role in smoking-induced disease. Supporting this, inhibition of CEBPB in human airway cells in vitro resulted in a compromised inflammatory response to smoke. Moreover, cigarette smoke-exposed CebpbΔLE mice displayed reduced respiratory neutrophilia and induction of inflammatory mediators, including the neutrophil chemoattractant Groa, compared to smoke- exposed controls. LPS-challenged CebpbΔLE mice also exhibited blunted respiratory neutrophilia and lower pulmonary expression of Groa, compared to LPS-challenged control littermates. In addition, suppression of LPS-induced neutrophilia and inflammatory gene expression by formoterol, a long acting β2-adrenoceptor agonist used in treatment of COPD, was impaired in CebpbΔLE mice. C/EBP transactivation was increased by treatment with formoterol in vitro, possibly through a β2-adrenoceptor and cAMP-dependent mechanism. This demonstrates that both inflammatory as well as anti-inflammatory stimuli involve regulation of gene transcription by C/EBPβ.
Taken together, these findings demonstrate that C/EBPα and C/EBPβ play pivotal and partly overlapping roles in airway epithelial differentiation, and that C/EBP and the lung epithelium orchestrates inflammatory responses as well as anti-inflammatory signaling by β2-adrenoceptor agonists in the lung. Thus, C/EBPs may influence tissue regeneration in lung homeostasis and disease as well as inflammatory and anti-inflammatory signaling, and are potential contributors to COPD pathogenesis.
To investigate unique vs. overlapping roles of C/EBPα and C/EBPβ in the lung, the pulmonary phenotype of mice lacking C/EBPα (CebpaΔLE mice), C/EBPβ (CebpbΔLE mice) or both C/EBPα and C/EBPβ (CebpaΔLE; CebpbΔLE mice) specifically in the lung epithelium, all generated by SFTPC-Cre mediated excision, was investigated. Cell culture experiments suggested that C/EBPα and C/EBPβ bind the same elements within a lung-specific promoter, and that their functions are partially overlapping. Pre-natal CebpaΔLE mice and CebpaΔLE; CebpbΔLE mice displayed immature lungs similar to the lungs of premature infants, and CebpaΔLE; CebpbΔLE mice exhibited even more impaired airway epithelial cell differentiation than the CebpaΔLE mice. The proportion of CebpaΔLE mice that survived and reached adulthood spontaneously developed a majority of the histopathological hallmarks of COPD, possibly caused by infiltrating inflammatory cells – similar to what is observed in COPD and what is mechanistically proposed to drive COPD pathogenesis. These findings are indicative of a relationship between immature lungs at birth, C/EBPs and the development of inflammatory lung disease.
Considering the previous documentation of decreased airway epithelial C/EBPβ activity in smokers with COPD, C/EBPβ could have a role in COPD pathogenesis. The role of C/EBPβ in regulating inflammatory and innate immune responses in the lung was on this account investigated by employing a translational approach encompassing clinical samples as well as in vitro and in vivo experiments. CEBPB was significantly down-regulated in the airway epithelium of both current and former smokers compared to never-smokers, and in cigarette smoke extract-treated primary human airway epithelial cells in vitro, suggesting that C/EBPβ plays a role in smoking-induced disease. Supporting this, inhibition of CEBPB in human airway cells in vitro resulted in a compromised inflammatory response to smoke. Moreover, cigarette smoke-exposed CebpbΔLE mice displayed reduced respiratory neutrophilia and induction of inflammatory mediators, including the neutrophil chemoattractant Groa, compared to smoke- exposed controls. LPS-challenged CebpbΔLE mice also exhibited blunted respiratory neutrophilia and lower pulmonary expression of Groa, compared to LPS-challenged control littermates. In addition, suppression of LPS-induced neutrophilia and inflammatory gene expression by formoterol, a long acting β2-adrenoceptor agonist used in treatment of COPD, was impaired in CebpbΔLE mice. C/EBP transactivation was increased by treatment with formoterol in vitro, possibly through a β2-adrenoceptor and cAMP-dependent mechanism. This demonstrates that both inflammatory as well as anti-inflammatory stimuli involve regulation of gene transcription by C/EBPβ.
Taken together, these findings demonstrate that C/EBPα and C/EBPβ play pivotal and partly overlapping roles in airway epithelial differentiation, and that C/EBP and the lung epithelium orchestrates inflammatory responses as well as anti-inflammatory signaling by β2-adrenoceptor agonists in the lung. Thus, C/EBPs may influence tissue regeneration in lung homeostasis and disease as well as inflammatory and anti-inflammatory signaling, and are potential contributors to COPD pathogenesis.
List of papers:
I. Didon L, Roos AB, Elmberger GP, Gonzalez FJ, Nord M. Lung-specific Inactivation of C/EBPα Causes a Pathological Pattern Characteristic of COPD. European Respiratory Journal. 2010; 35(1): 186–197.
Fulltext (DOI)
Pubmed
View record in Web of Science®
II. Roos AB, Berg T, Barton JL, Didon L and Nord M. Airway Epithelial Cell Differentiation During Lung Organogenesis Requires C/EBPα and C/EBPβ. Developmental Dynamics. 2012; 241(5): 911-23.
Fulltext (DOI)
Pubmed
View record in Web of Science®
III. Didon L, Barton JL, Roos AB, Gaschler GJ, Bauer CMT, Berg T, Stämpfli M R, Nord M. Lung Epithelial C/EBPβ is Necessary for the Integrity of Inflammatory Responses to Cigarette Smoke. American Journal of Respiratory and Critical Care Medicine. 2011; 184(2): 233-42.
Fulltext (DOI)
Pubmed
IV. Roos AB, Barton JL, Miller-Larsson A, Dahlberg B, Berg T, Didon L, Nord, M. Lung Epithelial C/EBPβ Contributes to LPS-induced Neutrophilia and its Suppression by Formoterol. [Submitted]
I. Didon L, Roos AB, Elmberger GP, Gonzalez FJ, Nord M. Lung-specific Inactivation of C/EBPα Causes a Pathological Pattern Characteristic of COPD. European Respiratory Journal. 2010; 35(1): 186–197.
Fulltext (DOI)
Pubmed
View record in Web of Science®
II. Roos AB, Berg T, Barton JL, Didon L and Nord M. Airway Epithelial Cell Differentiation During Lung Organogenesis Requires C/EBPα and C/EBPβ. Developmental Dynamics. 2012; 241(5): 911-23.
Fulltext (DOI)
Pubmed
View record in Web of Science®
III. Didon L, Barton JL, Roos AB, Gaschler GJ, Bauer CMT, Berg T, Stämpfli M R, Nord M. Lung Epithelial C/EBPβ is Necessary for the Integrity of Inflammatory Responses to Cigarette Smoke. American Journal of Respiratory and Critical Care Medicine. 2011; 184(2): 233-42.
Fulltext (DOI)
Pubmed
IV. Roos AB, Barton JL, Miller-Larsson A, Dahlberg B, Berg T, Didon L, Nord, M. Lung Epithelial C/EBPβ Contributes to LPS-induced Neutrophilia and its Suppression by Formoterol. [Submitted]
Institution: Karolinska Institutet
Supervisor: Nord, Magnus
Issue date: 2012-05-09
Rights:
Publication year: 2012
ISBN: 978-91-7457-775-4
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