Lung cancer has the highest mortality rate among all types of malignancies, accounting for approximately 29% of all cancer-related deaths in the United States [1
]. It has been estimated that in 2006 alone, the number of new lung cancer cases will exceed 174,000 and approximately 163,000 people will die of this disease [1
]. Tobacco smoking accounts for 85% of the lung cancers. Former heavy smokers remain at an elevated risk for developing lung cancer even years after they stop smoking [2
]. Fifty percent of newly diagnosed lung cancer patients are former smokers [4
]. It is therefore important to understand the effects of tobacco smoking on the bronchial epithelium in both active and former smokers.
Recently, a large-scale microarray study characterized gene expression differences between current, former, and never smokers [5
], and identified specific genes related to xenobiotic functions, anti-oxidation, cell adhesion and electron transport to be more highly expressed in current smokers relative to never smokers. Genetic regulators of inflammation and putative tumor suppressor genes exhibited decreased expression in current smokers relative to never smokers. Most significantly, a number of genes were identified that exhibited irreversible expression changes upon smoking cessation.
Additional reports have also identified increased expression of various xenobiotic metabolic enzymes including members of the cytochrome P450 (CYP) and glutathione S-transferase (GST) families of proteins in response to cigarette smoke exposure [5
]. CYP enzymes mediate the conversion of benzo (a) pyrene
and other polycyclic aromatic hydrocarbons (PAH) to carcinogenic intermediates that interact with genomic DNA [8
], thus contributing to the formation of DNA adducts in smokers [11
]. Members from both of the CYP
gene families have been implicated as potential susceptibility loci mediated by the presence of single nucleotide polymorphisms (SNPs) leading to aberrant expression in response to smoking [14
Another important process associated with tobacco smoke exposure is the airway mucosal response. In animal models, it has been shown that exposure to cigarette smoke induces goblet cell hyperplasia with accompanied mucus production [16
]. Moreover, mucin 5 (MUC5AC), has been shown to be the most highly expressed mucin in bronchial secretions [18
], induced in response to cigarette smoke through an EGFR-dependent mechanism [19
]. However, beyond this, little is known of the genes that are associated with airway remodeling as a result of tobacco smoking.
Serial analysis of gene expression (SAGE) is a quantitative experimental procedure widely used to determine expression profiles through the enumeration of short sequence tags and their relative abundance [20
]. Although the construction and sequencing of an individual SAGE library is expensive and laborious compared to microarray analysis, SAGE offers the invaluable potential for gene discovery as the analysis is not limited to genes represented on an array. Moreover, comparisons between independent experiments can be performed without sophisticated normalization [21
In this study, we compare the bronchial epithelial transcriptomes of current, former, and never smokers to determine the effect of active smoking on gene expression using bronchial brushings from the peripheral sub-segmental airways. Genes whose expression is reversible upon smoking cessation are expected to differ in abundance between current and former smokers, but are similar between former and never smokers. Conversely, gene expression that is irreversible upon smoking cessation will show similar levels in current and former (ever) smokers but differ between ever and never smokers. Here, we focus on identifying both reversible and irreversible gene expression changes and specifically consider these expression changes in the context of airway mucosal response, and susceptibility to cancer development.