The receptor for advanced glycation end-products (RAGE) has increasingly been demonstrated to be an important modulator of inflammation in cases of pulmonary disease. Published reports involving tobacco smoke exposure have demonstrated increased expression of RAGE, its participation in pro-inflammatory signaling and its role in irreversible pulmonary remodeling. The current research evaluated for the first time the in vivo effects of short-term tobacco smoke exposure in RAGE null and control mice compared to identical animals exposed to room air only. Quantitative real time PCR, immunoblotting, and immunohistochemistry revealed elevated RAGE expression in controls after four weeks of exposure and an anticipated absence of RAGE expression in RAGE null mice regardless of smoke exposure. Inflammatory cell behaviors were confirmed by measuring active Ras, NF-κB, and cytokine synthesis and secretion. Furthermore, bronchoalveolar lavage fluid (BALF) was procured from RAGE null and control animals after exposure for the assessment of total protein in order to indirectly measure vascular permeability, inflammatory cells and chemoattractant molecules involved in the inflammatory response. As a general theme, inflammation induced by tobacco smoke exposure was influenced by the availability of RAGE. These data reveal captivating information suggesting a role for RAGE signaling in lungs exposed to tobacco smoke. Furthermore, research may demonstrate RAGE signaling as an important therapeutic target capable of ameliorating cell level inflammation in those coping with exposure.

The tight junctions (TJ), which are located in the apical region between epithelial and endothelial cells, regulate the paracellular diffusion of ions and small molecules and play an important role in maintaining cell polarity, cell-cell integrity, and permeability. In the lung, epithelial cells are attached by TJ structures. They provide a permeable barrier and cell communication. The loss of barrier integrity, which is maintained by the expression of claudins (Cldn), results in cellular permibilization and leads to paracellular diffusion of solutes and harmful molecules. There are 27 known Cldn homologous members in mice and human. Cldn6 is mostly expressed in embryonic stem cells and associated with the programing of epithelial cells during embryo development and lung morphogenesis. In order to test the hypothesis that Cldn6 expression affects lung morphogenesis, we analyzed the expression pattern of Cldn6 during lung ontogenesis to examine cell-specific expression pattern of Cldn6 during each embryonic period in the mouse lung. Also, we assessed transcriptional regulators and control mechanisms that precisely influence Cldn6 expression in pulmonary cells. We discovered that Cldn6 is an important tight junctional component expressed by pulmonary epithelium during lung organogenesis. We found that normal down-regulation of Cldn6 as development proceeds influences differentiation associated with the transition between the embryonic to the alveolar stage. Conditional gain-of-function and loss-of-function experiments in animal models prove to be the most beneficial tool in deciphering the impact of Cldn in organ formation and maintenance. We generated a conditional transgenic mouse that provides the opportunity to genetically up-regulate Cldn6 in distal lung. Our transgenic mouse showed a delay in lung development and down-regulation of transcriptional factors. Cldn6 is both temporally and spatially controlled in the developing lung and its regulation is maintained by critical transcriptional control networks managed by TTF-1. In lung diseases, altered Cldn expression leads to diseases such as COPD, asthma, and ARDS. The tight junctional proteins are differentially regulated by tobacco smoke exposure and Cldn6 is potentially involved as neighboring epithelial cells respond to tobacco smoke. We exposed adult mice to controlled doses of second hand smoke during four days and A-549 cells to 10% CSE for 6 hours. We discovered that mice lungs respond by down-regulating Cldn6 basal levels and impair barrier function. These results reveal that midgestational up-regulation of Cldn6 and its marked down-regulation as development proceeds illustrate the notion that Cldn6 function is important during early programming stages of lung morphogenesis.

No description available.

No description available.

Lung cancer has been a major cause of death among types of cancers in the world. In the early stages, lung nodules can be detected by the aid of imaging modalities such as Computed Tomography (CT). In this stage, radiologists look for irregular rounded-shaped nodules in the lung which are normally less than 3 centimeters in diameter. Recent advancements in image analysis have proven that images contain more information than regular parameters such as intensity, histogram and morphological details. Therefore, in this project we have focused on extracting quantitative, hand-crafted features from nearly 1400 lung CT images to train a variety of classifiers based on them. In the first experiment, in total 424 Radiomics features per image has been used to train classifiers such as: Random Forest (RF), Support Vector Machine (SVM), Decision Tree (DT), Naive Bayes (NB), Linear Discriminant Analysis (LDA) and Multi-Layer Perceptron (MLP). In the second experiment, we evaluate each feature category separately with our classifiers. The third experiment includes wrapper feature selection methods (Forward/Backward/Recursive) and filter-based feature selection methods (Fisher score, Gini Index and Mutual information). They have been implemented to find the most relevant feature set in model construction. Performance of each learning method has been evaluated by accuracy score, wherewe achieved the highest accuracy of 78% with Random Forest classifier (74% in 5-fold average) and 0.82 Area Under the Receiver Operating Characteristics (AUROC) curve. After RF, NB and MLP showed the best average accuracy of 71.4% and 71% respectively.

Granular cell tumors (GCTs) are rare neoplasms of neuroectodermal origin characterized by large polygonal cells with abundant eosinophilic and granular cytoplasm. GCTs rarely affect the lungs, with only a few cases reported in the literature. The pathophysiology of this Schwann cell-derived condition is not well understood but is thought to be due to recurring genetic mutations. GCTs have been linked with Noonan syndrome. Here, we report the case of a 17-year-old caucasian male who presented with partial upper airway obstruction due to a GCT. This case promotes awareness among pathologists and clinicians for this condition in the workup of patients presenting with upper airway obstruction.

Chronic lung diseases including Chronic Obstructive Pulmonary Disease (COPD), Idiopathic Pulmonary Fibrosis (IPF) and lung cancer are major causes of morbidity and mortality in the United States due to high incidence and limited therapeutic options. In order to address this critical issue, I have leveraged RNA sequencing and integrative genomics to define disease-associated transcriptomic changes which could be potentially targeted to lead to new therapeutics. We sequenced the lung transcriptome of subjects with IPF (n=19), emphysema (n=19, a subtype of COPD), or neither (n=20). The expression levels of 1770 genes differed between IPF and control lung, and 220 genes differed between emphysema and control lung (p<0.001). Upregulated genes in both emphysema and IPF were enriched for the p53/hypoxia pathway. These results were validated by immunohistochemistry of select p53/hypoxia proteins and by GSEA analysis of independent expression microarray experiments. To identify regulatory events, I constructed an integrative miRNA target prediction and anticorrelation miRNA-mRNA network, which highlighted several miRNA whose expression levels were the opposite of genes differentially expressed in both IPF and emphysema. MiR-96 was a highly connected hub in this network and was subsequently overexpressed in cell lines to validate several potential regulatory connections. Building upon these successful experiments, I next sought to define gene expression changes and the miRNA-mRNA regulatory network in never smoker lung cancer. Large and small RNA was sequenced from matched lung adenocarcinoma tumor and adjacent normal lung tissue obtained from 22 subjects (8 never, 14 current and former smokers). I identified 120 genes whose expression was modified uniquely in never smoker lung tumors. Using a repository of gene-expression profiles associated with small bioactive molecules, several compounds which counter the never smoker tumor signature were identified in silico. Leveraging differential expression information, I again constructed an mRNA-miRNA regulatory network, and subsequently identified a potential never smoker oncomir has-mir-424 and its transcription factor target FOXP2. In this thesis, I have identified genes, pathways and the miRNA-mRNA regulatory network that is altered in COPD, IPF, and lung adenocarcinoma among never smokers. My findings may ultimately lead to improved treatment options by identifying targetable pathways, regulators, and therapeutic drug candidates. / 2017-02-01T00:00:00Z

Chronic obstructive pulmonary disease (COPD) is a major cause of morbidity and mortality worldwide and is a risk factor for lung cancer development. COPD encompasses both emphysema and chronic bronchitis, the pathogenesis of which are unclear. In this dissertation, I leveraged genome-wide gene-expression studies of emphysema and lung cancer to investigate pathogenesis and carcinogenesis in COPD. Tobacco smoke is the primary cause of emphysema. The most severe form is also associated with alpha1-antitrypsin deficiency (AATD) resulting from a mutation. In this study, I leveraged multiple lung samples from patients with emphysema, with or without AATD. While genes involved in tissue repair decreased with emphysema severity, the unfolded protein response (UPR) was uniquely changed in AATD lungs. AATD may play multiple roles in emphysema and UPR activation suggests AAT replacement therapy may be insufficient to treat this form of emphysema. Emphysema is a progressive disease, and the mean linear intercept (Lm) can serve as a surrogate of progression. I evaluated whether Lm increases in non-diseased lungs may represent similar processes to those occurring in emphysema, and could offer insight into early stages of disease or homeostasis. Genes involved in tissue repair increased with Lm in controls but decreased in disease. Tissue repair processes may be active in even the non-insulted lung, suggesting their activity is necessary for lung homeostasis and their deficiency may drive emphysema progression. Finally, COPD patients are at increased lung cancer risk, and transcriptomic changes common to both diseases could explain this risk. In both COPD and lung cancer, I discovered that H3K27Me3 regulated genes are repressed, and that the methyltransferase responsible for H3K27me3, EZH2, is induced. H3K27Me3, an oncogenic histone modification, may drive carcinogenesis and pathogenesis in COPD. Though usual and AATD emphysema share transcriptomic signatures associated with tissue repair, which may be active in the normal homeostatic lung, the UPR changes in AATD emphysema only; successful therapeutic strategies in emphysema will need to account for this difference. In COPD, H3K27Me3 may play a role in both pathogenesis and carcinogenesis, making it an attractive target for therapeutic interventions, but one that would need further augmentation in AATD. / 2019-11-01T00:00:00Z

Lung cancer (LC) is the leading cause of cancer death in the US, claiming over 160,000 lives annually. Although CT screening has been shown to be efficacious in reducing mortality, the limited access to screening programs among high-risk individuals and the high number of false positives contribute to low survival rates and increased healthcare costs. As a result, there is an urgent need for preventative therapeutics and novel interception biomarkers that would enhance current methods for detection of early-stage LC. This thesis addresses this challenge by examining the hypothesis that transcriptomic changes preceding the onset of LC can be identified by studying bronchial premalignant lesions (PMLs) and the normal-appearing airway epithelial cells altered in their presence (i.e., the PML-associated airway field of injury). PMLs are the presumed precursors of lung squamous cell carcinoma (SCC) whose presence indicates an increased risk of developing SCC and other subtypes of LC. Here, I leverage high-throughput mRNA and miRNA sequencing data from bronchial brushings and lesion biopsies to develop biomarkers of PML presence and progression, and to understand regulatory mechanisms driving early carcinogenesis. First, I utilized mRNA sequencing data from normal-appearing airway brushings to build a biomarker predictive of PML presence. After verifying the power of the 200-gene biomarker to detect the presence of PMLs, I evaluated its capacity to predict PML progression and detect presence of LC (Aim 1). Next, I identified likely regulatory mechanisms associated with PML severity and progression, by evaluating miRNA expression and gene coexpression modules containing their targets in bronchial lesion biopsies (Aim2). Lastly, I investigated the preservation of the PML-associated miRNAs and gene modules in the airway field of injury, highlighting an emergent link between the airway field and the PMLs (Aim 3). Overall, this thesis suggests a multi-faceted utility of PML-associated genomic signatures as markers for stratification of high-risk smokers in chemoprevention trials, markers for early detection of lung cancer, and novel chemopreventive targets, and yields valuable insights into early lung carcinogenesis by characterizing mRNA and miRNA expression alterations that contribute to premalignant disease progression towards LC. / 2020-01-25

Development of tyrosine kinase inhibitors (TKIs) against oncogenic drivers has significantly improved survival of patients with oncogene-mutated non-small cell lung cancer (NSCLC). However, acquired resistance to TKIs emerges over time in essentially all patients who initially respond. Recent evidence suggests that drug-tolerant persister (DTP) cells, which survive and adapt to targeted therapies during an early phase of treatment, play an important role in the emergence of drug resistance. A previous study reported that cluster of differentiation 74 (CD74) expression is upregulated in epidermal growth factor receptor (EGFR)-mutated lung cancer after treatment with EGFR-TKIs and that CD74 can be one of the DTP cell markers. However, both the mechanism underlying CD74 expression and the role of CD74 in DTP cells remain unclear. In the current study, an attempt was made to identify the mechanism using cell culture systems and transgenic mouse models. The results confirmed CD74 upregulation at the messenger RNA (mRNA) level after treatments with TKIs in various oncogene-mutated cell lines, including those with EGFR mutations, ROS1 fusions, and ALK fusions. The class II transactivator (CIITA), upstream of CD74, and tumor necrosis factor (TNF)-α expression were induced by treatments with TKIs in tumor cells, leading to an increase in CD74 expression. In addition, the results showed that treatments with TKIs enhance the autocrine secretion of macrophage migration inhibitory factor (MIF), a ligand of CD74, from tumor cells. This implied that autocrine stimulation of CD74 signaling blocks apoptosis and causes emergence of DTP cells. To examine whether CD74 plays an important role in the emergence of resistance to TKIs in vivo, experiments were completed in which lung-specific EGFR-L858R-T790M transgenic mice were crossed with Cd74 knockout mice. The results showed that complete deletion of CD74 overcomes or delays resistance to TKIs. Taken together, the results of this study suggest that the MIF-CD74 axis can be a novel target to overcome resistance in driver-mutated NSCLC. / 2026-02-28T00:00:00Z