Resected lobes from patients having pre-operative pulmonary function tests were fixed by inflation with formal saline and cut into 1cm parasagittal slices. Randomly selected tissue, from the lateral two slices, was plastic embedded and sections prepared for microscopic analysis. A semi-automatic image analysis system was used to quantitate bronchiolar calibre and shape and peribronchiolar attachment number, inter-alveolar attachment distance and the amount of macroscopic emphysema. An automatic image analyser (IBAS2) was used to measure alveolar surface area per-unit volume (AWUV). Measured bronchiolar calibre (minimum diameter and measured lumen area) was not related to patient height, lung volume, pulmonary function or other morphometric variables. AWUV, mean inter-alveolar attachment distance, theoretical lumen area and bronchiolar shape were independent of patient size and lung volume, but were inter-related. A combination of low AWUV and loss of attachments profoundly affected bronchiolar shape. However, AWUV and alveolar attachment loss were not always in proportion and demonstrated different functional effects: AWUV affects carbon monoxide transfer factor whereas attachments affect the slope of phase III and forced expiratory volume with bronchiolar shape affecting closing volume. Macroscopic emphysema did not accurately reflect the extent of alveolar wall loss as identified by AWUV and showed poor correlations with pulmonary function tests. Computerised axial tomography (CT scan) exhibited a strong correlation with AWUV and can be used to assess lung density in life.

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Nanomaterials are defined as any material with at least one external dimension less than 100 nm. Recently, nanomaterials have become more common in medicine, technology, and engineering. One reason for their increased interest is due to nanomaterials having unique properties that allow them to interact effectively with biological systems. In terms of drug delivery, the lungs are a highly desirable site to administer therapeutic nanoparticles. Indeed, inflammatory diseases such as asthma and emphysema could potentially benefit from nanoparticle-mediated delivery. However, the lungs are also in constant contact with airborne particulate matter. Thus, harmful nanoparticles can enter the lungs and cause or even exacerbate inflammatory diseases. Our work focused on characterization of both therapeutic and potentially harmful nanoparticles in the lungs. We found that fluorescently-labeled nanoparticles were phagocytosed by macrophages and did not induce apoptosis or inflammation in the lungs, making them potentially useful as a therapeutic for inflammatory diseases. We also characterized a rare form of titanium-based particles called Magnéli phases, which have been shown to be produced via coal burning. We found that while these particles are non-inflammatory in the lungs of mice, they lead to apoptosis of macrophages as well as a change in gene expression associated with increased fibrosis. Ultimately, this was shown to lead to a decrease in lung function parameters and airway hyperresponsiveness, indicating increased lung stiffness after long-term nanoparticle exposure. Our data adds significant contributions to the field by assessing two nanoparticles with vastly different compositions in the lungs. Overall, we found that the unique properties of both particle types allows for interactions with cells and tissues. These interactions can have important outcomes on health, both in terms of disease treatment and exacerbation. / Ph. D. / Over the years, nanoparticles have become more common in medicine, technology, and engineering due to their unique properties. Many of these properties allow for increased interactions with biological materials. Organs such as the lungs are at increased risk of exposure because they naturally encounter microorganisms and airborne particles on a daily basis. However, the lungs are also a highly desirable site for drug delivery using nanoparticles, due to ease of access. Inflammatory diseases such as asthma and emphysema could potentially benefit from nanoparticle-mediated delivery. Additionally, harmful nanoparticles can enter the lungs and cause or even exacerbate these diseases. Unfortunately, there is a lack of knowledge pertaining to this subject. Our work focused on assessing the interactions of nanoparticles in the lungs. First, we looked at nanoparticles that could be used for drug delivery. We found that fluorescentlylabeled nanoparticles were taken up by phagocytic white blood cells called macrophages. Furthermore, these particles did not induce cell death or inflammation in the lungs. Therefore, we found that these particles could be useful for drug delivery in the lungs. Secondly, we investigated potentially harmful nanoparticles and their effects on the lungs. The titanium-based particles called Magnéli phases, have been shown to be produced through coal burning. We found that while these particles are non-inflammatory in the lungs, they do lead to programmed death of macrophages as well as the increase in genes associated with fibrosis. Ultimately these particles led to a decrease in lung function after long-term exposure.

This document only includes an excerpt of the corresponding thesis or dissertation. To request a digital scan of the full text, please contact the Ruth Lilly Medical Library's Interlibrary Loan Department (rlmlill@iu.edu).

Cannabis is widely used for recreational and medicinal purposes. Inhalation of cannabis smoke is the predominant method of drug consumption, exposing the lungs to THC and CBD, as well as a plethora of toxic combustion products. Clinical observations suggest that cannabis smoking contributes to the development of respiratory symptoms and may play a role in the pathogenesis of inflammatory lung disease. However, the association between cannabis smoke, dysregulated pulmonary immunity, and the development of lung disease is inconclusive. To improve our understanding of this relationship, we developed novel mouse models to investigate the effect of cannabis smoke exposure on lung immunity. Using compositionally relevant cannabis strains, we established a mouse model of cannabis smoke exposure and validated that it delivers cannabis smoke by measuring cannabis smoke-associated metabolites in the blood. In our initial lung immune characterization, we demonstrated that acute cannabis smoke exposure induces modest changes to innate immune cellularity in the airways and lung tissue. Specifically, lung macrophage subpopulations were proportionally altered following smoke exposure. As well, we demonstrated that lung disease-associated mediators, including MDC, TARC, and VEGF, were dysregulated in cannabis smoke-exposed lung tissue. In addition to our initial characterization, we established a first-of-its-kind concurrent cannabis smoke exposure and influenza infection model. Using this model, we demonstrated that cannabis smoke exposure exacerbates weight loss following influenza infection. These increases in weight loss corresponded with dysregulated cellular responses and immune mediator expression. Cell types involved in early innate immune signaling, such as macrophages and dendritic cells, were significantly affected by concurrent exposure and infection. Additionally, anti-viral mediators. including IFNγ, IP-10, RANTES, and TNFα, were decreased in cannabis smoke-exposed, infected lung tissue. Collectively, we defined two novel models of cannabis smoke exposure that can be leveraged in future investigations on the inflammatory effects and associated health outcomes of cannabis smoke. / Thesis / Master of Science (MSc) / Cannabis is widely used for recreational and medical purposes. Smoking is the most popular method to consume the drug among users. However, little is understood about the effects of cannabis smoke on lung health, despite evidence suggesting that it may lead to negative health outcomes. To address this gap in knowledge, we developed two unique mouse models of cannabis smoke exposure. Using these models, we explored the effects of cannabis smoke on lung immune responses in healthy and influenza infected mice. Our findings suggested that lung immunity is altered following cannabis smoke exposure. Additionally, we found that overall health was worsened during influenza infection in cannabis smoke-exposed mice. This effect was associated with weakened viral immunity in the lungs. The models we developed and the findings using it thus far create the foundation for future studies on cannabis smoke and lung health.

5-year survival rate of patients with metastasized non-small cell lung cancer (NSCLC) who received chemotherapy was less than 5% (Kathryn C. Arbour, 2019). Our ability to provide survival status of a patient i.e. Alive or death at any time in future is important from at least two standpoints: a) from clinical standpoint it enables clinicians to provide optimal delivery of healthcare and b) from personal standpoint by providing patient’s family with opportunities to plan their life ahead and potentially cope with emotional aspect of loss of life. / Thesis / Master of Applied Science (MASc)

There is increasing evidence indicating that exposure to air pollutants may be associated with the onset of several respiratory diseases such as allergic airway disease and chronic obstructive pulmonary disorder (COPD). Many lung diseases demonstrate an outgrowth of pathogenic bacteria belonging to the Proteobacteria phylum, and the incidence of occurrence of these diseases is higher in heavily polluted regions. Within the human body, the lungs are among the first to be exposed to the harmful effects of inhaled pollutants and microbes. Research in the past few decades have expounded on the air-pollution-induced local and systemic inflammatory responses, but the involvement of the lung microbial communities has not yet been well-characterized. Lungs were historically considered to be sterile, but recent advances have demonstrated that the lower respiratory tract is replete with a wide variety of microorganisms - both in health and disease. Recent studies show that these lung microbes may play a significant role in modulating the immune environment by inducing IgA and mucus production. Air pollutants have previously been shown to alter intestinal bacterial populations that increase susceptibility to inflammatory diseases; however, to date, the effects of traffic-generated air pollutants on the resident microbial communities on the lungs have not been explored. The microbiome is influenced by several factors, including diet and environmental exposures. A large percentage of the Western world population consumes a high-fat (HF) diet which has resulted in the epidemic of obesity. Consumption of an HF diet has been shown to alter the intestinal microflora and increase baseline inflammation. We aimed to understand whether diet might also contribute to the alteration of the commensal lung microbiome, either alone or related to exposure. Thus, we investigated the hypothesis that exposure to air pollutants can alter the commensal lung microbiota, thereby promoting alterations in the lung's immune and inflammatory responses; in addition to determining whether these outcomes are exacerbated by a high fat-diet. We performed two studies with exposures to different components of air pollutant mixtures on C57Bl/6 mice placed on either a control (LF) diet or a high-fat (HF) diet. Our first exposure study was performed on C57Bl/6 mice with a mixture of gasoline and diesel engine emissions (ME: 30 µg PM/m3 gasoline engine emissions + 70 µg PM/m3 diesel engine emissions) or filtered air (FA) for 6h/d, 7 d/wk for 30 days. The ME study investigated the alterations in immunoglobulin A (IgA), IgG and IgM, and lung microbiota abundance and diversity. Our results revealed ME exposures alongside the HF diet causes a decrease in IgA and IgG when compared to FA controls, thereby decreasing airway barrier protection. This was accompanied by the expansion of bacteria within the Proteobacteria phylum and a decrease in the overall bacterial diversity and richness in the exposed vs. control groups. In our second study, we exposed C57Bl/6 mice to only the diesel exhaust particle component (35µg DEP, suspended in 35µl 0.9% sterile saline) or sterile saline only (control) twice a week for 30 days. We investigated immunoglobulin profiles by ELISA that revealed a significant increase in IgA and IgG in response to DEP. We also observed an increase in inflammatory tumor necrosis factor (TNF) - α, Interleukin (IL) -10, Toll-like receptors (TLR) - 2,4, nuclear factor kappa B (NF-κB) histologically and by RT-qPCR. Mucus production and collagen deposition within the lungs were also significantly elevated with DEP exposures. Microbial abundance determined quantitatively from the bronchoalveolar lavage fluid (BALF) by qPCR revealed an expansion of bacteria belonging to the Proteobacteria phylum in the DEP exposed groups on the HF diet. We also observed an increase in reactive oxygen and nitrogen species (ROS-RNS) products (nitrates), within the groups that revealed an expansion of Proteobacteria. These observations are most likely due to the unique metabolic capabilities of Proteobacteria to proliferate in inflammatory environments with excess nitrates. We assessed if treatments with probiotics could attenuate the DEP-induced inflammation by supplementing a separate group of study animals on the HF diet with 0.3 g/day of Winclove Ecologic® Barrier probiotics in their drinking water throughout the study. With probiotic treatments, we observed a significant decrease in ROS-RNS that was accompanied by complete elimination of Proteobacteria suggesting that in the absence of nitrates, the expansion of Proteobacteria is curbed effectively. We also observed a decrease in proinflammatory TNF-α and collagen deposition with probiotic treatments, and an increase in IgA levels within the BALF, suggesting that probiotics aid in balancing proinflammatory responses and enhance beneficial immune responses to efficiently mediate the DEP-induced inflammation. Both studies showed that air pollutants alter the immune defenses and contribute to lung microbial alterations with an expansion of Proteobacteria. The immunoglobulin profiles discordant between the two studies can be explained by the route and/or duration and composition of air pollutant exposure. Collectively these studies suggest that exposure to air pollutants alter immune responses and/or increase the availability of inflammatory by-products within the lungs that can enable the selective outgrowth of pathogenic bacteria. The observed detrimental outcomes are further exacerbated when coupled with the consumption of an HF diet. Importantly, these results may shed light on the missing link between air pollution-induced inflammation and bacterial expansion and also point to therapeutic alternatives to curb bacterial outgrowth in lung disease exacerbations observed in patient populations living and/or working in heavily polluted regions.

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