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Epigenomic alterations and gene expression profiles in respiratory epithelia exposed to cigarette smoke condensate.

ABSTRACT: Limited information is available regarding epigenomic events mediating initiation and progression of tobacco-induced lung cancers. In this study, we established an in vitro system to examine epigenomic effects of cigarette smoke in respiratory epithelia. Normal human small airway epithelial cells and cdk-4/hTERT-immortalized human bronchial epithelial cells (HBEC) were cultured in normal media with or without cigarette smoke condensate (CSC) for up to 9 months under potentially relevant exposure conditions. Western blot analysis showed that CSC mediated dose- and time-dependent diminution of H4K16Ac and H4K20Me3, while increasing relative levels of H3K27Me3; these histone alterations coincided with decreased DNA methyltransferase 1 (DNMT1) and increased DNMT3b expression. Pyrosequencing and quantitative RT-PCR experiments revealed time-dependent hypomethylation of D4Z4, NBL2, and LINE-1 repetitive DNA sequences; up-regulation of H19, IGF2, MAGE-A1, and MAGE-A3; activation of Wnt signaling; and hypermethylation of tumor suppressor genes such as RASSF1A and RAR-beta, which are frequently silenced in human lung cancers. Array-based DNA methylation profiling identified additional novel DNA methylation targets in soft-agar clones derived from CSC-exposed HBEC; a CSC gene expression signature was also identified in these cells. Progressive genomic hypomethylation and locoregional DNA hypermethylation induced by CSC coincided with a dramatic increase in soft-agar clonogenicity. Collectively, these data indicate that cigarette smoke induces 'cancer-associated' epigenomic alterations in cultured respiratory epithelia. This in vitro model may prove useful for delineating early epigenetic mechanisms regulating gene expression during pulmonary carcinogenesis.

SUBMITTER: Liu F

PROVIDER: S-EPMC6329296 | biostudies-literature | 2010 Jun

REPOSITORIES: biostudies-literature

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Epigenomic alterations and gene expression profiles in respiratory epithelia exposed to cigarette smoke condensate.

Liu F F Killian J K JK Yang M M Walker R L RL Hong J A JA Zhang M M Davis S S Zhang Y Y Hussain M M Xi S S Rao M M Meltzer P A PA Schrump D S DS

Oncogene 20100503 25

Limited information is available regarding epigenomic events mediating initiation and progression of tobacco-induced lung cancers. In this study, we established an in vitro system to examine epigenomic effects of cigarette smoke in respiratory epithelia. Normal human small airway epithelial cells and cdk-4/hTERT-immortalized human bronchial epithelial cells (HBEC) were cultured in normal media with or without cigarette smoke condensate (CSC) for up to 9 months under potentially relevant exposure ...[more]

PMID: 20440268

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Project description:Rationale: The effects of hookah smoke on respiratory epithelia have not been well characterized. Objectives: To characterize and compare the effects of hookah tobacco smoke and conventional cigarette smoke on the epigenome and transcriptome of human respiratory epithelia. Methods: Normal human small airway epithelial cells and cyclin-dependent kinase 4/human telomerase reverse transcriptase–immortalized human bronchial epithelial cells were cultured for 5 days in normal media in the presence or absence of water pipe condensates or cigarette smoke condensates under relevant exposure conditions. CyQUANT assay (Thermo Fisher Scientific), RNA sequencing, quantitative reverse transcriptase–polymerase chain reaction, and Western blotting techniques were used to examine/compare the effects of hookah and cigarette smoke on cell proliferation, messenger RNA/microRNA expression, and global histone marks. Results: Water pipe condensates– and cigarette smoke condensates–mediated, dose-dependent growth-inhibitory effects in cultured respiratory epithelial cells were measured. Under conditions mediating equipotent mild to moderate growth-inhibitory effects, water pipe condensates as well as cigarette smoke condensates decreased histone H4 lysine 15 acetylation (H4K16ac) and histone H4 lysine 20 trimethylation (H4K20me3) levels in small airway epithelial cells and human bronchial epithelial cells; these histone alterations are putative “hallmarks of cancer.” RNA sequencing analysis demonstrated that cigarette smoke condensates and water pipe condensates mediated dose-dependent alterations in gene expression in small airway epithelial cells and human bronchial epithelial cells. A total of 873 genes were commonly regulated (fold change >2) by water pipe condensates (1.0 mg/ml) and cigarette smoke condensates (0.05 mg/ml) in small airway epithelial cells, whereas a total of 1,577 genes were commonly regulated by water pipe condensates (1.0 mg/ml) and cigarette smoke condensates (0.05 mg/ml) in human bronchial epithelial cells. Ingenuity Pathway Analysis (QIAGEN Bioinformatics) of the 100 genes commonly regulated by water pipe condensates and cigarette smoke condensates in small airway epithelial cells and human bronchial epithelial cells demonstrated that the top three activated upstream regulators are cigarette smoke (z-score?=?2.577), benzo[a]pyrene (z-score?=?2.905), and nuclear factor (erythroid-derived 2)-like 2 (NFE2L2, Nrf2) (z-score?=?2.531). Analysis of downstream effects indicated that growth of epithelial tissue was predicted to be increased (z-score?=?2.813; P?<?0.001). The top three canonical pathways included xenobiotic metabolism signaling, aryl hydrocarbon receptor signaling, and nicotine degradation III. Conclusions: These preliminary findings strongly suggest that hookah tobacco is not a safe alternative to cigarettes, and warrant further evaluation of the epigenomic effects of hookah smoke in human respiratory epithelia using our established laboratory models and correlative analyses in hookah smokers.

Project description:BackgroundThe coronavirus disease 2019 (COVID-19), which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has become a worldwide pandemic with over 627 million cases and over 6.5 million deaths. It was reported that smoking-related chronic obstructive pulmonary disease (COPD) might be a crucial risk for COVID-19 patients to develop severe condition. As cigarette smoke (CS) is the major risk factor for COPD, we hypothesize that barrier dysfunction and an altered cytokine response in CS-exposed airway epithelial cells may contribute to increased SARS-CoV-2-induced immune response that may result in increased susceptibility to severe disease. The aim of this study was to evaluate the role of CS on SARS-CoV-2-induced immune and inflammatory responses, and epithelial barrier integrity leading to airway epithelial damage.MethodsPrimary human airway epithelial cells were differentiated under air-liquid interface culture. Cells were then exposed to cigarette smoke medium (CSM) before infection with SARS-CoV-2 isolated from a local patient. The infection susceptibility, morphology, and the expression of genes related to host immune response, airway inflammation and damages were evaluated.ResultsCells pre-treated with CSM significantly caused higher replication of SARS-CoV-2 and more severe SARS-CoV-2-induced cellular morphological alteration. CSM exposure caused significant upregulation of long form angiotensin converting enzyme (ACE)2, a functional receptor for SARS-CoV-2 viral entry, transmembrane serine protease (TMPRSS)2 and TMPRSS4, which cleave the spike protein of SARS-CoV-2 to allow viral entry, leading to an aggravated immune response via inhibition of type I interferon pathway. In addition, CSM worsened SARS-CoV-2-induced airway epithelial cell damage, resulting in severe motile ciliary disorder, junctional disruption and mucus hypersecretion.ConclusionSmoking led to dysregulation of host immune response and cell damage as seen in SARS-CoV-2-infected primary human airway epithelia. These findings may contribute to increased disease susceptibility with severe condition and provide a better understanding of the pathogenesis of SARS-CoV-2 infection in smokers.

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Epigenomic alterations and gene expression profiles in respiratory epithelia exposed to cigarette smoke condensate.

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Epigenomic alterations and gene expression profiles in respiratory epithelia exposed to cigarette smoke condensate.

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