Project description:In the process of seeking novel lung host defense regulators by analyzing genome-wide RNA sequence data from normal human airway epithelium, we detected expression of POU2AF1, a known transcription co-factor previously thought to be expressed only in lymphocytes. Lymphocyte contamination of human airway epithelial samples obtained by bronchoscopy and brushing was excluded by immunohistochemistry staining, the observation of up-regulation of POU2AF1 in purified airway basal stem/progenitor cells undergoing differentiation and analysis of differentiating single basal cell clones. Lentivirus-mediated up-regulation of POU2AF1 in airway basal cells induced up-regulation of host defense genes, including MX1, IFIT3, IFITM and known POU2AF1 downstream genes HLA-DRA, ID2, ID3, IL6, BCL6. Interestingly, expression of these genes paralleled changes of POU2AF1 expression during airway epithelium differentiation in vitro, suggesting POU2AF1 helps to maintain a "host defense tone" even in pathogen-free condition. Cigarette smoke, a known risk factor for airway infection, suppressed POU2AF1 expression both in vivo in humans and in vitro in human airway epithelial cultures, accompanied by deregulation of POU2AF1 downstream genes. Finally, enhancing POU2AF1 expression in human airway epithelium attenuated the suppression of host defense genes by smoking. Together, these findings suggest a novel function of POU2AF1 as a potential regulator of host defense genes in the human airway epithelium. Methods: Massive parallel RNA sequencing was used to compare the transcriptome of lentivirus mediated POU2AF1 or RFP (control) gene expression in human primary airway epithelial cells (3 samples per group). Uninfected basal cell was used as a further control. Conclusions: The genes up-regulated by POU2AF1 in human airway epithelial cells are mainly related to the intracellular or extracellular anti-pathogen response, suggesting POU2AF1 plays a role in airway epithelial host defense. By genome-wide-based screening, POU2AF1, a known lymphocyte transcription co-factor, was found to be expressed in human airway epithelium and regulate host defense genes. It might be a drug target as smoking-compromised host defense is associated with down-regulation of POU2AF1. In this Series, human airway epithelial cell transcriptomes (3 uninfected without treatment, 3 infected with lenti-RFP virus and 3 infected with lenti-POU2AF1 virus) were compared using massive parallel RNA sequencing (Illumina HiSeq 2000).

Project description:In the process of seeking novel lung host defense regulators by analyzing genome-wide RNA sequence data from normal human airway epithelium, we detected expression of POU2AF1, a known transcription co-factor previously thought to be expressed only in lymphocytes. Lymphocyte contamination of human airway epithelial samples obtained by bronchoscopy and brushing was excluded by immunohistochemistry staining, the observation of up-regulation of POU2AF1 in purified airway basal stem/progenitor cells undergoing differentiation and analysis of differentiating single basal cell clones. Lentivirus-mediated up-regulation of POU2AF1 in airway basal cells induced up-regulation of host defense genes, including MX1, IFIT3, IFITM and known POU2AF1 downstream genes HLA-DRA, ID2, ID3, IL6, BCL6. Interestingly, expression of these genes paralleled changes of POU2AF1 expression during airway epithelium differentiation in vitro, suggesting POU2AF1 helps to maintain a "host defense tone" even in pathogen-free condition. Cigarette smoke, a known risk factor for airway infection, suppressed POU2AF1 expression both in vivo in humans and in vitro in human airway epithelial cultures, accompanied by deregulation of POU2AF1 downstream genes. Finally, enhancing POU2AF1 expression in human airway epithelium attenuated the suppression of host defense genes by smoking. Together, these findings suggest a novel function of POU2AF1 as a potential regulator of host defense genes in the human airway epithelium. Methods: Massive parallel RNA sequencing was used to compare the transcriptome of lentivirus mediated POU2AF1 or RFP (control) gene expression in human primary airway epithelial cells (3 samples per group). Uninfected basal cell was used as a further control. Conclusions: The genes up-regulated by POU2AF1 in human airway epithelial cells are mainly related to the intracellular or extracellular anti-pathogen response, suggesting POU2AF1 plays a role in airway epithelial host defense. This Series represents samples complementary to those in GSE60989.

Project description:In the process of seeking novel lung host defense regulators by analyzing genome-wide RNA sequence data from normal human airway epithelium, we detected expression of POU2AF1, a known transcription co-factor previously thought to be expressed only in lymphocytes. Lymphocyte contamination of human airway epithelial samples obtained by bronchoscopy and brushing was excluded by immunohistochemistry staining, the observation of up-regulation of POU2AF1 in purified airway basal stem/progenitor cells undergoing differentiation and analysis of differentiating single basal cell clones. Lentivirus-mediated up-regulation of POU2AF1 in airway basal cells induced up-regulation of host defense genes, including MX1, IFIT3, IFITM and known POU2AF1 downstream genes HLA-DRA, ID2, ID3, IL6, BCL6. Interestingly, expression of these genes paralleled changes of POU2AF1 expression during airway epithelium differentiation in vitro, suggesting POU2AF1 helps to maintain a "host defense tone" even in pathogen-free condition. Cigarette smoke, a known risk factor for airway infection, suppressed POU2AF1 expression both in vivo in humans and in vitro in human airway epithelial cultures, accompanied by deregulation of POU2AF1 downstream genes. Finally, enhancing POU2AF1 expression in human airway epithelium attenuated the suppression of host defense genes by smoking. Together, these findings suggest a novel function of POU2AF1 as a potential regulator of host defense genes in the human airway epithelium. Methods: Massive parallel RNA sequencing was used to compare the transcriptome of lentivirus mediated POU2AF1 or RFP (control) gene expression in human primary airway epithelial cells (3 samples per group). Uninfected basal cell was used as a further control. Conclusions: The genes up-regulated by POU2AF1 in human airway epithelial cells are mainly related to the intracellular or extracellular anti-pathogen response, suggesting POU2AF1 plays a role in airway epithelial host defense.

Project description:The toll-like receptors (TLRs) are important components of the respiratory epithelium host innate defense, enabling the airway surface to recognize and respond to a variety of insults in inhaled air. Based on the knowledge that smokers are more susceptible to pulmonary infection and the airway epithelium of smokers with chronic obstructive pulmonary disease (COPD) is characterized by bacterial colonization and acute exacerbation of airway infections, we assessed whether smoking alters the expression of TLRs in human small airway epithelium, the primary site of smoking-induced disease. Microarrays were used to survey the TLR family gene expression in small airway (10th-12th order) epithelium from healthy nonsmokers (n=60), healthy smokers (n=73) and smokers with COPD (n=36). Using the criteria of detection call of present in ≥50%, 6 of 10 TLRs (1, 2, 3, 4, 5 and 8) were expressed. Compared to nonsmokers, the most strikingly changed gene is TLR5, which down-regulated in healthy smokers (1.4-fold decrease, p<10-13) and in smokers with COPD (1.6-fold, p<10-14). TaqMan RT-PCR confirmed these observations. Bronchial biopsies immunofluorescence showed that TLR5 protein was expressed mainly on the apical side of the human airway epithelium and decreased in healthy smokers and smokers with COPD. In vitro studies showed that the level of TLR5 downstream genes, IL-6 and IL-8 were highly induced in TLR5 high-expressing cells compared to TLR5 low-expressing cells after flagellin exposure. In the context that TLR5 functions to recognize pathogens and activate innate immune responses, the smoking-induced down-regulation of TLR5 likely contributes to smoking-related susceptibility to airway infection. The toll-like receptors (TLRs) are important components of the respiratory epithelium host innate defense. Microarrays were used to survey the TLR family gene expression in small airway (10th-12th order) epithelium from healthy nonsmokers (n=60), healthy smokers (n=73) and smokers with COPD (n=36). Using the criteria of detection call of present in ≥50%, 6 of 10 TLRs (1, 2, 3, 4, 5 and 8) were expressed. Compared to nonsmokers, the most strikingly changed gene is TLR5, which down-regulated in healthy smokers (1.4-fold decrease, p<10-13) and in smokers with COPD (1.6-fold, p<10-14). In the context that TLR5 functions to recognize pathogens and activate innate immune responses, the smoking-induced down-regulation of TLR5 likely contributes to smoking-related susceptibility to airway infection. *** Processed data not provided for all gene expression records. ***

Project description:Lectins are proteins present on cell surfaces or as shed extracellular proteins that function in innate immune defense as phagocytic receptors to recognize specific bacterial cell wall components. Based on the knowledge that cigarette smoking is associated with increased risk of bacterial infection, we hypothesized that cigarette smoking may modulate the expression of lectin genes in the airway epithelium. Affymetrix HG U133 Plus 2.0 microarrays were used to survey expression of lectin genes in large (3rd to 4th order bronchi) airway epithelium from 9 normal nonsmokers and 20 phenotypic normal smokers and small (10th to 12th order bronchi) airway epithelium from 13 normal nonsmokers and 20 phenotypic normal smokers. From the 72 lectin genes that were surveyed, there were no changes (>2-fold change, p<0.05) in gene expression in either large or small airway epithelium among normal smokers compared to nonsmokers except for a striking down regulation in both large and small airway epithelium of normal smokers of intelectin 1, a recently described lectin that participates in the innate immune response by recognizing and binding to galactofuranosyl residues in the cell walls of bacteria (large airway epithelium, p<0.003; small airway epithelium, p<0.002). TaqMan RT-PCR confirmed the observation that intelectin 1 was down-regulated in both large (p<0.05) and small airway epithelium (p<0.02) of normal smokers compared to normal nonsmokers. Immunohistochemistry assessment of biopsies of the large airway epithelium of normal nonsmokers demonstrated intelectin 1 was expressed in secretory cells, with qualitatively decreased expression in biopsies from normal smokers. Western analysis confirmed the decreased expression of intelectin 1 in airway epithelium of normal smokers compared to normal nonsmokers (p<0.02). Finally, compared to normal nonsmokers, intelectin 1 expression was decreased in small airway epithelium of smokers with early COPD (n= 13, p<0.001) and smokers with established COPD (n= 14, p<0.001), in a fashion similar to that of normal smokers. In the context that intelectin 1 is an epithelial molecule that likely plays a role in defense against bacteria, the down regulation of expression of intelectin 1 in response to cigarette smoking may contribute to the increase in susceptibility to infections observed in smokers, including those with COPD. Keywords: COPD Comparison of gene expression in airway epithelial cells of normal non-smokers, phenotypic normal smokers, smokers with early COPD, and smokers with COPD.

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