Changes In Lungs Of Mice Research Articles

Gene expression changes in mouse lung induced by subacute inhalation of PM10-rich particulate matter

Introduction Particulate matter (PM) air pollution is associated with an increased incidence of lung diseases, but the underlying mechanisms have not been fully elucidated. In this study, a mouse model of subacute lung inflammation was employed to investigate the cellular responses and gene expression changes induced by exposure to natural ambient air pollution. Methods C57BL/6J mice were exposed to road dust (primarily PM10) at 150 µg/m³ for 21 days (8 h/day) through a nose-only inhalation exposure system. Lung tissues were analyzed for the expression of proinflammatory signaling, oxidative stress, and fibrosis markers. RNA-sequencing analysis was conducted to identify differentially expressed genes (DEGs). A gene ontology over-representation analysis was performed to identify the altered genetic pathways. Results Elevated levels of proinflammatory cytokines, including IL-1β, IL-6, and TNF-α, and an increase in phosphorylated MAPK were determined in the road dust exposure group compared to the control group. Histopathological examinations revealed more severe lung inflammation and damage in the exposed mice, including fibrosis and bronchiolar hyperplasia. Gene expression profiling identified 108 DEGs, with decreases in most except genes such as Krt15 and Reg3g. The protein–protein interaction network analysis together with text-mining identified 18 key hub genes, associated with fatty acid oxidation, lipid metabolism, and peroxisomes. Conclusion This study identified key genes, signaling pathways, and cellular responses in mouse lung affected by road dust exposure. These findings contribute to a deeper understanding of the transcriptional and cellular responses induced by subacute exposure to the PM in road dust.

Molecular Framework of Mouse Endothelial Cell Dysfunction during Inflammation: A Proteomics Approach.

A key aspect of cytokine-induced changes as observed in sepsis is the dysregulated activation of endothelial cells (ECs), initiating a cascade of inflammatory signaling leading to leukocyte adhesion/migration and organ damage. The therapeutic targeting of ECs has been hampered by concerns regarding organ-specific EC heterogeneity and their response to inflammation. Using in vitro and in silico analysis, we present a comprehensive analysis of the proteomic changes in mouse lung, liver and kidney ECs following exposure to a clinically relevant cocktail of proinflammatory cytokines. Mouse lung, liver and kidney ECs were incubated with TNF-α/IL-1β/IFN-γ for 4 or 24 h to model the cytokine-induced changes. Quantitative label-free global proteomics and bioinformatic analysis performed on the ECs provide a molecular framework for the EC response to inflammatory stimuli over time and organ-specific differences. Gene Ontology and PANTHER analysis suggest why some organs are more susceptible to inflammation early on, and show that, as inflammation progresses, some protein expression patterns become more uniform while additional organ-specific proteins are expressed. These findings provide an in-depth understanding of the molecular changes involved in the EC response to inflammation and can support the development of drugs targeting ECs within different organs. Data are available via ProteomeXchange (identifier PXD031804).

Eupatilin Suppresses OVA-Induced Asthma by Inhibiting NF-κB and MAPK and Activating Nrf2 Signaling Pathways in Mice.

To investigate the effect of eupatilin in asthma treatment, we evaluated its therapeutic effect and related signal transduction in OVA-induced asthmatic mice and LPS-stimulated RAW264.7 cells. The BALF was tested for changes in lung inflammatory cells. Th2 cytokines in the BALF and OVA-IgE in the serum were measured by ELISA. H&E and PAS staining were used to evaluate histopathological changes in mouse lungs. The key proteins NF-κB, MAPK, and Nrf2 in lung tissues were quantitatively analyzed by Western blotting. Finally, we evaluated the effect of eupatilin on cytokines and related protein expression in LPS-stimulated RAW 264.7 cells in vitro. In OVA-induced asthmatic mice, eupatilin reduced the numbers of inflammatory cells, especially neutrophils and eosinophils. Eupatilin also decreased the levels of IL-5, IL-13 in the BALF and OVA-IgE in the serum. Furthermore, eupatilin inhibited the activation of NF-κB and MAPK pathways and increased the expression of Nrf2 in OVA-induced asthmatic mice. In vitro, eupatilin significantly reduced LPS-stimulated NO, IL-6, and ROS production. Additionally, the NF-κB, MAPK, and Nrf2 protein expression in LPS-stimulated RAW264.7 cells was consistent with that in OVA-induced asthmatic lung tissues. In summary, eupatilin attenuated OVA-induced asthma by regulating NF-κB, MAPK, and Nrf2 signaling pathways. These results suggest the utility of eupatilin as an anti-inflammatory drug for asthma treatment.

Characterization of the Onset, Progression, and Reversibility of Morphological Changes in Mouse Lung after Pharmacological Inhibition of Leucine-Rich Kinase 2 Kinase Activity.

Gain-of-function mutations in leucine-rich kinase 2 (LRRK2) are associated with increased incidence of Parkinson disease (PD); thus, pharmacological inhibition of LRRK2 kinase activity is postulated as a disease-modifying treatment of PD. Histomorphological changes in lungs of nonhuman primates (NHPs) treated with small-molecule LRRK2 kinase inhibitors have brought the safety of this treatment approach into question. Although it remains unclear how LRRK2 kinase inhibition affects the lung, continued studies in NHPs prove to be both cost- and resource-prohibitive. To develop a tractable alternative animal model platform, we dosed male mice in-diet with the potent, highly selective LRRK2 kinase inhibitor MLi-2 and induced histomorphological changes in lung within 1 week. Oral bolus dosing of MLi-2 at a frequency modeled to provide steady-state exposure equivalent to that achieved with in-diet dosing induced type II pneumocyte vacuolation, suggesting pulmonary changes require sustained LRRK2 kinase inhibition. Treating mice with MLi-2 in-diet for up to 6 months resulted in type II pneumocyte vacuolation that progressed only modestly over time and was fully reversible after withdrawal of MLi-2. Immunohistochemical analysis of lung revealed a significant increase in prosurfactant protein C staining within type II pneumocytes. In the present study, we demonstrated the kinetics for onset, progression, and rapid reversibility of chronic LRRK2 kinase inhibitor effects on lung histomorphology in rodents and provide further evidence for the derisking of safety and tolerability concerns for chronic LRRK2 kinase inhibition in PD. SIGNIFICANCE STATEMENT: We have defined a mouse model by which the on-target lung effects of leucine-rich kinase 2 (LRRK2) kinase inhibition can be monitored, whereas previous in vivo testing relied solely on nonhuman primates. Data serve to derisk long-term treatment with LRRK2 kinase inhibitors, as all lung changes were mild and readily reversible.

Using Bronchoalveolar Lavage to Evaluate Changes in Pulmonary Diseases.

Bronchoalveolar lavage (BAL) is a procedure that can be used to collect samples from human and animal lungs to efficiently evaluate the immune response and the potentially pathological changes by examining both the compositions of cells and fluid from lavage. There are observable changes including inflammatory response in human and animal lungs exposed to environmental exposures such as toxic chemicals and microorganisms, or under pathophysiological conditions in respiratory system. The profile of inflammatory cells in BAL provides a qualitative description of inflammatory response, and the secretion in BAL fluid contains secreted proteins of inflammatory mediators and albumin as a quantitative measurement of inflammation and tissue injury in the lungs. Mouse is the most common model system being used for pulmonary disease-related research. A consistent experimental approach on how to lavage mouse lungs and collect samples from mouse lungs is important for a reproducible evaluation of pathological and physiological changes in mouse lung especially for the analysis of inflammation.

Similar and Differential Canonical Pathways and Biological Processes Associated With Multiwalled Carbon Nanotube and Asbestos-Induced Pulmonary Fibrosis: A 1-Year Postexposure Study.

Respiratory exposure to multiwalled carbon nanotubes (MWCNT) or asbestos results in fibrosis; however, the mechanisms to reach this end point may be different. A previous study by our group identified pulmonary effects and significantly altered messenger RNA (mRNA) signaling pathways following exposure to 1, 10, 40, and 80 µg MWCNT and 120 µg crocidolite asbestos on mouse lungs over time at 1-month, 6-month, and 1-year postexposure following pulmonary aspiration. As a continuation to the above study, this current study took an in-depth look at the signaling pathways involved in fibrosis development at a single time point, 1 year, and exposure, 40 µg MWCNT, the lowest exposure at which fibrosis was pathologically evident. The 120 µg asbestos exposure was included to compare MWCNT-induced fibrosis with asbestos-induced fibrosis. A previously validated computational model was used to identify mRNAs with expression profiles matching the fibrosis pathology patterns from exposed mouse lungs. mRNAs that matched the pathology patterns were then input into ingenuity pathway analysis to determine potential signaling pathways and physiological disease functions inherent to MWCNT and asbestos exposure. Both MWCNT and asbestos exposure induced changes in mouse lungs regarding gene expression, cell proliferation, and survival, while MWCNT uniquely induced alterations in pathways involved in oxidative phosphorylation, mitochondrial dysfunction, and transcription. Asbestos exposure produced unique alterations in pathways involved in sustained inflammation. Although typically considered similar due to scale and fiber-like appearance, the different compositional properties inherent to either MWCNT or asbestos may play a role in their ability to induce fibrosis after pulmonary exposure.

Epithelial cell-derived cytokines CST3 and GDF15 as potential therapeutics for pulmonary fibrosis

While wound healing is completed, the epithelium functions to normalize the interstitial context by eliminating fibroblasts excited during matrix reconstruction. If not, tissues undergo pathologic fibrosis. Pulmonary fibrosis is a fatal and hardly curable disorder. We here tried to identify epithelium-derived cytokines capable of ameliorating pulmonary fibrosis. Human lung fibroblasts were inactivated in epithelial cell-conditioned media. Cystatin C (CST3) and growth differentiation factor 15 (GDF15) were found to be enriched in the conditioned media and to inhibit the growth and activation of lung fibroblasts by inactivating the TGF–Smad pathway. In mouse and human lungs with interstitial fibrosis, CST3 and GDF15 expressions were markedly reduced, and the restoration of these cytokines alleviated the fibrotic changes in mouse lungs. These results suggest that CST3 and GDF15 are bona fide regulators to prevent excessive proliferation and activation of fibroblasts in injured lungs. These cytokines could be potential therapeutics for ameliorating interstitial lung fibrosis.

Inhaled resveratrol treatments slow ageing-related degenerative changes in mouse lung.

Lung ageing, a significant risk factor for chronic human lung diseases such as COPD and emphysema, is characterised by airspace enlargement and decreasing lung function. Likewise, in prematurely ageing telomerase null (terc-/-) mice, p53 stabilisation within diminishing numbers of alveolar epithelial type 2 cells (AEC2) accompanies reduced lung function. Resveratrol (RSL) is a plant phytoalexin that has previously showed efficacy in enhancing invertebrate longevity and supporting mammalian muscle metabolism when delivered orally. Here, we tested whether inhaled RSL could protect young, terc-/- mice from accelerated ageing of the lung. terc-/- mice aged 2 months inhaled 1 mg/kg RSL that was instilled intratracheally once per month for 3 months. One month after the last inhalation, whole lung function, structure and cellular DNA damage were evaluated and AEC2 survival was assessed by western blotting for survival pathway gene expression. RSL treatments delayed the loss of lung compliance (p<0.05), maintained lung structure (p<0.001) and blocked parenchymal cell DNA damage as measured by TdT Nick-End Labeling (TUNEL). RSL, a known agonist of deacetylase SIRT1, supported AEC2 survival by stimulating SIRT1 expression, promoting p53 destabilisation and decreasing Bax expression and by maintaining expression levels of Peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), activated p-Akt and p-Mdm2 and inactivated Phospho-Phosphatase and tensin homolog (p-PTEN). RSL prophylaxis by inhalation is a potential approach for slowing ageing-related deterioration of lung function and structure by maintaining AEC2 integrity.

Identification of paraquat-induced lung injury and fibrosis in mice using intraperitoneal injection in different dose and exposure time

Objective To explore the effect of paraquat (PQ) on lung injury and fibrosis in mice by using intra-peritoneal administration of different doses and different lengths of exposure time, as well as to establish a more appropriate animal model of lung injury and related fibrosis in a simple operation with favorable reproducibility. Methods Male C57BL/6J mice of 8-10 weeks in the model group were intra-peritoneally administered with different paraquat doses for different lengths of exposure time. There were three fashions of giving PQ: single dosing with different doses (40 mg/kg, 50 mg/kg, or 60 mg/kg), successive dosing every day for 5 d with different doses (10 mg/kg, 12.5 mg/kg, or 15 mg/kg) or intermittent dosing every 4 d for 4 occasions with different doses (20 mg/kg, 22.5 mg/kg, or 25 mg/kg), respectively. Control animals (randomly selected) received an equal volume of saline instead at the same time. Mice were sacrificed at 1 week or 2 weeks after the last exposure. The general physical condition of mice including body weight as well as mortality was monitored daily. Lung static compliance measurement, bronchoalveolar lavage (BAL) fluid samples for cytospin analysis, hydroxyproline assay, and whole left lung sections stained for histological analysis were obtained. Results Both single dosing method and intermittent dosing method with different doses of paraquat could induce acute lung injury in some of mice. The inflammation changes became more serious as the dose increased during the early observation. While during the late observation, the pulmonary fibrosis was found in only a few survival mice. The pathological section showed the fibrotic lesions were mainly found in posterior portion, sub-pleural and peripheral portions of lung. However, the mortality was high in these two methods during early lung injury stage, and lower morbidity of pulmonary fibrosis in late stage. The successive dosing strategy didn’t produced significant differences in pathological changes in lung of mice between different intervals of observation. Conclusion Single dosing with high dosage or intermittent dosing with moderate dosage as a systemic administration strategy was necessary for intra-peritoneal PQ to induce mice acute lung injury. However, the model with low morbidity of pulmonary fibrosis and high mortality during acute injury stage was unsatisfactory one. PQ administration with successive lower doses strategy was not sufficient to induce lung injury as well as pulmonary fibrosis in C57BL/6J mice. Key words: Paraquat; Intraperitoneal injection; Lung injury; Pulmonary fibrosis; Animal model

A mouse model of chronic idiopathic pulmonary fibrosis

Chronic idiopathic pulmonary fibrosis (IPF) is a progressive, fatal, and untreatable disease with unclear etiology. There are few models of this chronic pathology, and although delivery of bleomycin to induce acute lung injury is the most common animal model of pulmonary fibrosis, there is considerable uncertainty about whether this acute injury resolves in those animals that survive. In this report, we have systematically followed groups of mice for up to 6 months following a single insult of bleomycin. We assessed changes in lung function and pathology over this time course, with measurements of the diffusion capacity for carbon monoxide, lung mechanics, quantitative stereology, and collagen. Our results show that, while there is some repair over this extended time course, the injury in the lung never fully resolves. This persistent degree of fibrosis may have similarities to many features of human IPF. Thus, these chronic fibrotic changes in mouse lungs could be a useful model to evaluate potential therapeutic interventions to accelerate repair and possible treat this debilitating disease.

Cotinine Concentration in Serum Correlates with Tobacco Smoke-Induced Emphysema in Mice

Secondhand smoke (SHS) has been associated with a variety of adverse health outcomes in nonsmokers, including emphysema (a chronic obstructive pulmonary disease). One way to detect SHS exposure is to measure the concentration of cotinine, the primary metabolite of nicotine, in bodily fluids. We have developed a method for cotinine analysis by combining micellar electrokinetic chromatography with enrichment techniques. We employed the method to measure cotinine concentrations in serum samples of mice exposed to tobacco smoke for 12 or 24 weeks and found that it was 3.1-fold or 4.8-fold higher than those exposed to room air for the same period. Further, we investigated the morphological changes in lungs of mice and observed tobacco smoke induced emphysema. Our results indicate that the method can be used to measure cotinine and there is an association between the serum cotinine concentration and tobacco smoke-induced emphysema in mice.

Morphological and Structural Changes in Lung Tissue Infected by Paracoccidioides brasiliensis: FTIR Photoacoustic Spectroscopy and Histological Analysis

This study evaluated physical, chemical and morphological changes in lungs of mice infected with Paracoccidioides brasiliensis. The animals were inoculated with 0.1mL of fungal suspension of the P. brasiliensis 18 isolate and were euthanized 1, 2, 4 and 8weeks after inoculation. The upper left lobe of the lung was isolated, fixed and processed for paraffin embedding. The sections were stained with H&E for histopathological study, with Gomori-Grocott to locate and identify the fungus, and with TUNEL immunostaining to detect the occurrence of programmed cell death. The lower and middle right lobes were analyzed by Fourier Transform Infrared Photoacoustic Spectrocopy (FTIR-PAS) to investigate physical and chemical features of the infected lungs. The results showed that lungs infected by P. brasiliensis underwent structural changes that varied according to the time period analyzed, and that changes in the absorption bands of different chemical groups resulted from these morphological changes. The results suggest that the combination of FTIR-PAS spectroscopy with morphological evaluation is an effective procedure for the study of paracoccidioidomycosis, one of the most important systemic mycoses that can damage the lung architecture and consequently impair the respiratory function.

Abstract 2810: Evaluating MMP contribution to pre-metastatic niche formation.

Abstract Metastasis, the fatal progression of solid malignancies, consists of a complex and multistep process characterized by a series of ordered biological events. One step in this process occurs when stromal microenvironmental changes take place at secondary sites before the arrival of tumor cells. These modified tissue-specific sites are induced by signals from the primary tumor and further supported by recruitment of progenitor bone marrow derived cells and are commonly referred to as “pre-metastatic niches.” Matrix metalloproteinases (MMPs) are crucially involved in many pathological processes including tumor invasion and metastasis. Our goal is to determine the roles of MMPs in generation of a pre-metastatic niche in different target organs, specifically the lung and liver. Using an MMP proteolytic beacon capable of detecting MMP activity in situ, we observed increased signal in lungs of mice carrying pre-metastatic mammary tumors. In a LLC-implantation model, qPCR was used to profile MMP expression in pre-metastatic tissues versus controls and indicated that in the lung, MMP9 (gelatinase B) was increased. Further, we have developed staining and quantitation techniques to define pre-metastatic niches. These were also based on molecules identified by Kaplan et al such as VEGFR-1, VLA-4 and fibronectin. Immunofluorescent analysis of mouse lungs subcutaneously implanted with LLC cells demonstrated clusters of cells that positively co-stained for VEGFR-1, VLA-4 and Fibronectin. We also tested whether soluble factors from metastatic cell lines could induce changes in lungs of mice. Analysis of MMP-9 levels as well as localization of the known markers both failed to show any significant changes in the lung, leading us to believe that the presence of a primary tumor in vivo is required for pre-metastatic niche formation. We are now working with the well-characterized metastatic mammary tumor line 4T1 to test whether the changes we have seen with LLC cells are reproducible in a different cancer type and mouse strain. Additionally we are testing whether the absence of host MMP-9 affects the formation of a pre-metastatic niche in either model. The increased presence of MMP-9 and mentioned factors before tumor cell arrival could indicate potential roles and markers for metastatic spread, possibly leading to potential therapeutics for preventing metastasis and increasing overall cancer survival. Egeblad et al. Journal of Biochemistry and Molecular Biology, Vol. 36, No. 1, January 2002, pp. 128-137 Kaplan et el. Nature, Vol. 438, No. 7069, December 2005, pp 820-827 Citation Format: Elizabeth A. Dozier, J. Oliver McIntyre, Conor Lynch, Lynn Matrisian, Barbara Fingleton. Evaluating MMP contribution to pre-metastatic niche formation. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2810. doi:10.1158/1538-7445.AM2013-2810

Detection and Early Phase Assessment of Radiation-Induced Lung Injury in Mice Using Micro-CT

Radiation therapy is an important therapeutic modality for thoracic malignancies. However, radiation-induced pulmonary injuries such as radiation pneumonitis and fibrosis are major dose-limiting factors. Previous research shows that micro-computed tomography (micro-CT) can detect radiation-induced lung injuries a few months following irradiation, but studies to assess the early response of lung tissue are lacking. The aim of this study was to determine if micro-CT could be used to detect and assess early-phase radiation–induced lung injury in mice. Twenty-one animals were divided into three groups: normal (n = 7), one day after x-ray exposure (n = 7), and at four days after x-ray exposure (n = 7). The x-ray-exposed groups received a single dose of 20 Gy, to the whole lung. Histology showed enlargements of the air space (Lm: mean chord length) following irradiation. 40.5±3.8 µm and 60.0±6.9 µm were observed after one and four days, respectively, compared to 26.5±3.1 µm in normal mice. Three-dimensional micro-CT images were constructed and histograms of radiodensity - Hounsfield Units (HU) - were used to assess changes in mouse lungs. Radiation-induced lung injury was observed in irradiated mice, by the use of two parameters which were defined as shifts in peak HU between −200 to −800 HU (PeakHU) and increase in the number of pixels at −1000 HU (Number-1000). These parameters were correlated with histological changes. The results demonstrate that micro-CT can be used for the early detection and assessment of structural and histopathological changes resulting from radiation-induced lung injury in mice. Micro-CT has the advantage, over traditional histological techniques, of allowing longitudinal studies of lung disease progression and assessment of the entire lung, while reducing the number of animals required for such studies.

Global Transcriptome Analysis in Influenza-Infected Mouse Lungs Reveals the Kinetics of Innate and Adaptive Host Immune Responses

An infection represents a highly dynamic process involving complex biological responses of the host at many levels. To describe such processes at a global level, we recorded gene expression changes in mouse lungs after a non-lethal infection with influenza A virus over a period of 60 days. Global analysis of the large data set identified distinct phases of the host response. The increase in interferon genes and up-regulation of a defined NK-specific gene set revealed the initiation of the early innate immune response phase. Subsequently, infiltration and activation of T and B cells could be observed by an augmentation of T and B cell specific signature gene expression. The changes in B cell gene expression and preceding chemokine subsets were associated with the formation of bronchus-associated lymphoid tissue. In addition, we compared the gene expression profiles from wild type mice with Rag2 mutant mice. This analysis readily demonstrated that the deficiency in the T and B cell responses in Rag2 mutants could be detected by changes in the global gene expression patterns of the whole lung. In conclusion, our comprehensive gene expression study describes for the first time the entire host response and its kinetics to an acute influenza A infection at the transcriptome level.

Anthrax Lethal Toxin-Induced Gene Expression Changes in Mouse Lung

A major virulence factor of Bacillus anthracis is the anthrax Lethal Toxin (LeTx), a bipartite toxin composed of Protective Antigen and Lethal Factor. Systemic administration of LeTx to laboratory animals leads to death associated with vascular leakage and pulmonary edema. In this study, we investigated whether systemic exposure of mice to LeTx would induce gene expression changes associated with vascular/capillary leakage in lung tissue. We observed enhanced susceptibility of A/J mice to death by systemic LeTx administration compared to the C57BL/6 strain. LeTx-induced groups of both up- and down-regulated genes were observed in mouse lungs 6 h after systemic administration of wild type toxin compared to lungs of mice exposed to an inactive mutant form of the toxin. Lungs of the less susceptible C57BL/6 strain showed 80% fewer differentially expressed genes compared to lungs of the more sensitive A/J strain. Expression of genes known to regulate vascular permeability was modulated by LeTx in the lungs of the more susceptible A/J strain. Unexpectedly, the largest set of genes with altered expression was immune specific, characterized by the up-regulation of lymphoid genes and the down-regulation of myeloid genes. Transcripts encoding neutrophil chemoattractants, modulators of tumor regulation and angiogenesis were also differentially expressed in both mouse strains. These studies provide new directions for the investigation of vascular leakage and pulmonary edema induced by anthrax LeTx.

Serial MRI characterization of the functional and morphological changes in mouse lung in response to cardiac remodeling following myocardial infarction

The temporal evolution of heart failure and associated pulmonary congestion in rodent heart failure models has not yet been characterized simultaneously and noninvasively. In this study, MRI was used to assess the serial progression of left-ventricular dysfunction and lung congestion in mice following myocardial infarction (MI). Cardiac and lung (1) H MRI was performed at baseline and every 3 days up to 13 days postsurgery in sham and MI mice. Respiratory parameters and terminal lung mechanics were assessed followed by histological analysis. MRI revealed that the MI induced significant pulmonary congestion/edema as detected by increased MRI signal intensity and was associated with increased lung volume and reduced cardiac contractility. Pulmonary function was also depressed in MI-mice, reflected by a reduced tidal volume and a low minute ventilation rate. Additionally, MI significantly increased lung resistance, markedly reduced lung compliance and total lung capacity and significantly increased lung weights by 57%. Significant correlations were observed between the MRI measured lung congestion, lung volume, ejection fraction, and lung wet-weight parameters. This study demonstrates that MRI may be of significant value in evaluating therapies aimed at primary intervention for lung congestion and secondary prevention of unfavorable cardiac remodeling.

Chronic LPS inhalation causes emphysema-like changes in mouse lung that are associated with apoptosis.

Lipopolysaccharide (LPS) is ubiquitous in the environment. Recent epidemiologic data suggest that occupational exposure to inhaled LPS can contribute to the progression of chronic obstructive pulmonary disease. To address the hypothesis that inhaled LPS can cause emphysema-like changes in mouse pulmonary parenchyma, we exposed C57BL/6 mice to aerosolized LPS daily for 4 weeks. By 3 days after the end of the 4-week exposure, LPS-exposed mice developed enlarged airspaces that persisted in the 4-week recovered mice. These architectural alterations in the lung are associated with enhanced type I, III, and IV procollagen mRNA as well as elevated levels of matrix metalloproteinase (MMP)-9 mRNA, all of which have been previously associated with human emphysema. Interestingly, MMP-9-deficient mice were not protected from the development of LPS-induced emphysema. However, we demonstrate that LPS-induced airspace enlargement was associated with apoptosis within the lung parenchyma, as shown by prominent TUNEL staining and elevated cleaved caspase 3 immunoreactivity. Antineutrophil antiserum-treated mice were partially protected from the lung destruction caused by chronic inhalation of LPS. Taken together, these findings demonstrate that inhaled LPS can cause neutrophil-dependent emphysematous changes in lung architecture that are associated with apoptosis and that these changes may be occurring through mechanisms different than those induced by cigarette smoke.

Comparative Proteomic Analysis of Radiation-Induced Changes in Mouse Lung: Fibrosis-Sensitive and -Resistant Strains

To determine whether comparative proteomics could detect differential protein expression after lung irradiation in two mouse strains with different radiation responses, lung proteins were subjected to two-dimensional orthogonal liquid-phase separations, with chromatofocusing in the first dimension and nonporous silica reverse-phase high-performance liquid chromatography (NPS-RP-HPLC) in the second. Five weeks after 12 Gy whole-lung irradiation, 15 and 31 proteins had significantly altered expression levels in C3H/HeJ (less likely to develop lung fibrosis) and C57BL/6J mice (more likely to develop lung fibrosis), respectively. These proteins were analyzed by HPLC-electrospray ionization tandem mass spectrometry (HPLC-ESI-MS/MS) and identified by matching sequences in a peptide database. The proteins are associated with redox, energy consumption, glycolysis, or chromatin/ RNA structure formation. Five of the six redox-related proteins, including superoxide dismutase 1 (SOD1), cytochrome c oxidase, glutamate dehydrogenase, biliverdin reductase, peroxiredoxin and carbonyl reductase, were down-regulated in the irradiated C57BL/6J mice, whereas SOD1, sulfurtransferase and carbonyl reductase increased in the irradiated C3H/ HeJ mice. Thus decreased antioxidant proteins in the irradiated C57BL/6J mice may be correlated with increased early lung toxicity. Changes in SOD1 and 8-hydroxydeoxy-guanosine (8-OHdG, an oxidative stress marker) were further confirmed by immunohistochemistry and/or Western blot analysis. These data suggest that a proteomics approach has the potential to detect protein changes relevant to early lung toxicity after irradiation.

In Utero Environmental Tobacco Smoke Exposure Alters Gene Expression in Lungs of Adult BALB/c Mice

BackgroundIn utero environmental tobacco smoke (ETS) exposure exacerbates initial lung responses of adult mice to ovalbumin (OVA), a common allergen in rodent models of allergic asthma.ObjectiveWe tested the hypothesis that in utero ETS exposure alters expression of genes (including asthma-related and inflammatory genes) in the lungs of adult mice and that this differential expression is reflected in differential respiratory and immune responses to nontobacco allergens.MethodsUsing Affymetrix Mouse Genome 430 2.0 arrays, we examined gene expression changes in lungs of BALB/c mice exposed to ETS in utero, OVA, or saline aerosol at weeks 7–8, and OVA sensitization and challenge at weeks 11–15. Data sets were filtered by transcript p-value (≤ 0.05), false discovery rate (≤ 0.05), and fold change (≥ 1.5). Differential expression of selected genes was confirmed by polymerase chain reaction (PCR).ResultsGenes differentially expressed as a result of in utero ETS exposure are involved in regulation of biological processes (immune response, cell proliferation, apoptosis, cell metabolism) through altered cytoskeleton, adhesion, transcription, and enzyme molecules. A number of genes prominent in lung inflammation were differentially expressed on PCR but did not pass selection criteria for microarray, including arginase (Arg1), chitinases (Chia, Chi3l3, Chi3l4), eotaxins (Ccl11, Ccl24), small proline-rich protein 2a (Sprr2a), and cytokines (Il4, Il6, Il10, Il13, Tnfa) .ConclusionThe differential lung gene expression reported here is consistent with previously reported functional changes in lungs of mice exposed in utero to ETS and as adults to the nontobacco allergen OVA.