Osteopontin and cadherin 11 are novel mediators and drug targets for chronic lung diseases

Chronic lung diseases (CLDs) are a considerable source of morbidity and mortality and are thought to arise from dysregulation of normal wound healing processes. An aggressive, feature of many CLDs is pulmonary fibrosis (PF) and is characterized by excess deposition of extracellular matrix (ECM) proteins from myofibroblasts in airways. However, factors regulating myofibroblast biology are incompletely understood. Proteins in the cadherin family contribute epithelial to mesenchymal transition (EMT), a suggested source of myofibroblasts. Cadherin 11 (CDH11) contributes to developmental and pathologic processes that parallel those seen in PF and EMT. Utilizing Cdh11 knockout (Cdh11 -/-) mice, the goal of this study was to characterize the contribution of CDH11 in the bleomycin model of PF and assess the feasibility of treating established PF. We demonstrate CDH11 in macrophages and airway epithelial cells undergoing EMT in lungs of mice given bleomycin and patients with PF. Endpoints consistent with PF including ECM production and myofibroblast formation are reduced in CDH11-targeted mice given bleomycin. Findings suggesting mechanisms of CDH11-dependent fibrosis include the regulation of the profibrotic mediator TGF-â in alveolar macrophages and CDH11-mediated EMT. The results of this study propose CDH11 as a novel drug target for PF. In addition, another CLD, chronic obstructive pulmonary disease (COPD), is characterized by airway inflammation and destruction. Adenosine, a nucleoside signaling molecule generated in response to cell stress is upregulated in patients with COPD and is suggested to contribute to its pathogenesis. An established model of adenosine-mediated lung injury exhibiting features of COPD is the Ada -/- mouse. Previous studies in our lab suggest features of the Ada -/- phenotype may be secondary to adenosine-dependent expression of osteopontin (OPN). OPN is a protein implicated in a variety of human pathology, but its role in COPD has not been examined. To address this, Ada/Opn -/- mice were generated and endpoints consistent with COPD were examined in parallel with Ada -/- mice. Results demonstrate OPN-mediated pulmonary neutrophilia and airway destruction in Ada -/- mice. Furthermore, patients with COPD exhibit increased OPN in airways which correlate with clinical airway obstruction. These results suggest OPN represents a novel biomarker or therapeutic target for the management of patients with COPD. The importance of findings in this thesis is highlighted by the fact that no pharmacologic interventions have been shown to interfere with disease progression or improve survival rates in patients with COPD or PF.

Characterization of Staphylococcus aureus adhesins important for respiratory tract infection

Staphylococcus aureus is a leading cause of lower respiratory tract infections in both adult and pediatric populations. In the past two decades, reports have described emergent incidence of severe necrotizing pneumonia in previously healthy individuals, frequently caused by antibiotic resistant strains. Additionally, S. aureus remains the most common cause of ventilator-associated pneumonia, contributing morbidity and mortality in intensive care units. As treatment of infection is made more difficult by the resistance to multiple antibiotics including vancomycin, there is a pressing need for novel strategies to prevent and treat S. aureus infections. Targeting essential mechanisms that promote infection such as adhesion, colonization, invasion, evasion of immune system and signaling may lead to inhibition of pathogenic surge. Staphylococcal adhesins of the MSCRAMM family (microbial surface components recognizing adherent matrix molecules) represent viable targets for such investigations. Understanding the molecular mechanism of binding is the first step toward the development of such therapies. Analysis of bacterial strains isolated from patients with staphylococcal pneumonia show increased expression of protein A, SdrD, SdrC and ClfB, cell surface proteins members of the MSCRAMM family. In this study the interaction of these MSCRAMMs with candidate ligands has been examined. We found that SdrD mediates S. aureus adherence to the lung epithelial cell line A549. Consistently, bacteria expressing SdrD have increased persistence in the lungs of infected mice after bronchoalveolar lavage in comparison with bacteria lacking this protein. Inhibition studies revealed that bacterial attachment can be abolished using neutralizing antibodies against SdrD. Using phage display, neurexin β isoforms were identified as SdrC binding partners. Previous reports postulated that MSCRAMMS bind their ligands by a 'dock, lock and latch' mechanism of interaction. Our data suggested that ClfB, an MSCRAMM responsible for nasal colonization, binds cytokeratin 10 by a 'dock and lock' variant of this model, in which the 'latching' event is not necessary. In summary, we have characterized aspects of molecular interaction between several MSCRAMMS and host components. We hope that continued delineation of these interactions will lead to identification of novel therapeutic targets or preventive strategies against S. aureus infections. ^

Scanned Ion Beam Therapy for Thoracic Tumors

Although frequently cured of Hodgkin lymphoma, adolescents and young adults can develop radiation induced second cancers. These patients could potentially benefit from scanned ion radiotherapy yet likely would require motion mitigation strategies. In theory, four-dimensional (4D) optimization of ion beam fields for individual motion states of respiration can enable superior sparing of healthy tissue near moving targets, compared to other motion mitigation strategies. Furthermore, carbon-ion therapy can sometimes provide greater relative biological effectiveness (RBE) for cell sterilization in a target but nearly equivalent RBE in tissue upstream of the target, compared to proton therapy. Thus, we expected that for some patients with Hodgkin lymphoma, carbon-ion therapy would reduce the predicted risk of second cancer incidence in the breast compared with proton therapy. The purpose of this work was to determine whether 4D-optimized carbon-ion therapy would significantly reduce the predicted risk of radiation induced second cancers in the breast for female Hodgkin lymphoma patients while preserving tumor control compared with proton therapy. To achieve our goals, we first investigated whether 4D-optimized carbon beam tracking could reduce dose to volumes outside a moving target compared with 3D-optimized carbon beam tracking while preserving target dose coverage. To understand the reliability of scanned carbon beam tracking, we studied the robustness of dose distributions in thoracic targets to uncertainties in patient motion. Finally, we investigated whether using carbon-ion therapy instead of proton therapy would significantly reduce the predicted risk of second cancer in the breast for a sample of Hodgkin lymphoma patients. We found that 4D-optimized ion beam tracking therapy can reduce the maximum dose to critical structures near a moving target by as much as 53%, compared to 3D-optimized ion beam tracking therapy. We validated these findings experimentally using a scanned carbon ion synchrotron and a motion phantom. We found scanned carbon beam tracking to be sensitive to a number of motion uncertainties, most notably phase delays in tracking, systematic spatial errors, and interfractional motion changes. Our findings indicate that a lower risk of second cancer in the breast might be expected for some Hodgkin lymphoma patients using carbon-ion therapy instead of proton therapy. For our reference scenario, we found the ratio of risk to be 0.77 ± 0.35 for radiogenic breast cancer after carbon-ion therapy versus proton therapy. Our findings were dependent on the RBE values for tumor induction and the radiosensitivity of breast tissue, as well as the physical dose distribution.

NANOPARTICLE AGGLOMERATES FOR PULMONARY DRUG DELIVERY

The Pulmonary route has been traditionally used to treat diseases of the respiratory tract. However, important research within the last two decades have shown that in addition to treating local diseases, a wide range of systemic diseases can be treated by delivering drugs to the lungs. The recent FDA approval to market Exubera, an inhalable form of insulin developed by Pfizer, to treat Diabetes, may just be the stepping stone that the pharmaceutical industry needs to market other drugs to treat systemic diseases via the lungs. However, this technology still needs repeated drug doses to control glucose levels, as the inhaled drug is cleared rapidly. Technologies have been developed where inhaled particles are capable of controlled release of drug from the lungs. An important feature of these technologies is the large geometric size of the particles that makes it difficult for the lung macrophages to clear these particles, which results in longer residence times for the particles in the lungs. Owing to the porosity, these particles have lower densities making them deliverable to the deep lungs. However, no modulation of drug release can be achieved with these technologies when more drug release may be required. This additional requirement can only be assuaged by additional dosing of the drug formulation, which can have undesirable effects due to excess loading of excipients in the lungs. In an attempt to bring about modulation of release from long residence time particles, a novel concept was developed in our laboratory that has been termed as the Agglomerated Vesicle Technology (AVT). Liposomes with encapsulated drug were agglomerated using well known cross linking chemistries to form agglomerates in the micron sized range. The large particles exhibited aerodynamic sizes in the respirable size range with minimal damage to the particles upon nebulization. By breaking the cross links between the liposomes with a cleaving agent, it was anticipated that triggered release of drug from the AVT particles could be achieved. In vivo studies done in healthy rabbits showed that post-administration modulation of drug release is possible from the AVT particles after the introduction of the cleaving agent. This study has important implications for the future development of this technology, where the AVT particles can be made “sensitive” to the product of disease. It is envisaged that a single dose of AVT containing the appropriate drug when administered to the lungs would maintain drug levels at a controlled rate over an extended period of time. When the need for more drug arises, the product of the disease would trigger the AVT particles to release more drug as needed to control the condition, thus eliminating the need for repeated drug doses and improved compliance amongst patients.

Respiratory syncytial virus infection in hematopoietic stem cell transplant patients

Respiratory Syncytial Virus (RSV) is a major cause of respiratory tract infections in immunocompromised patients such as children less than 2 years, premature infants with congenital heart disease and chronic lung disease, elderly patients and patients who have undergone hematopoietic stem cell transplant (HSCT). HSCT patients are at high risk of RSV infection, at increased risk of developing pneumonia, and RSV-related mortality. Immunodeficiency can be a major risk factor for severe infection & mortality. Therapy of RSV infection with Ribavirin, Palivizumab and Immunoglobulin has shown to reduce the risk of progression to LRI and mortality, especially if initiated early in the disease. Data on RSV infection in HSCT patients is limited, especially at various levels of immunodeficiency. 323 RSV infections in HSCT patients have been identified between 1/1995 and 8/2009 at University of Texas M D Anderson Cancer Center (UTMDACC). In this proposed study, we attempted to analyze a de-identified database of these cases and describe the epidemiologic characteristics of RSV infection in HSCT patients, the course of the infection, rate of development of pneumonia and RSV-related mortality in HSCT patients at UTMDACC.^ Key words: RSV infections, HSCT patients ^

Water to Thrive health needs assessment for Robit, Ethiopia

Developing countries suffer from an array of diseases, of which the developed world is unfamiliar. In order to facilitate the development of community interventions and streamline NGO partnership, needs assessments in targeted areas are conducted. The purpose of this assessment was to attain baseline descriptive data to further understand the needs of the village of Robit, Ethiopia. A trained team collected data from Austin based non-profit Water to Thrive (W2T) on June 1st and 2nd, 2011 through focus groups, key informant interviews, and individual surveys. Qualitative and quantitative data were paired to affirm the results of one another through triangulation. The results identified an apparent need for health intervention and education. Malaria, water-borne disease, respiratory issues (asthma, Upper respiratory tract infections), and maternal and child health were among the evident problems in Robit. There was a clear need for midwife training as well as water sanitation, latrine building, and general illness treatment. Poor road conditions and annual flooding of Robit plays an important role in the poor health and lack of food security of the village. While some evidence of social desirability and recall bias was found in the interview and survey data, the triangulation of findings provided important insights and validity to the needs assessment. ^

CREB mediates prostaglandin F2alpha-induced MUC5AC overexpression.

Mucus secretion is an important protective mechanism for the luminal lining of open tubular organs, but mucin overproduction in the respiratory tract can exacerbate the inflammatory process and cause airway obstruction. Production of MUC5AC, a predominant gel-forming mucin secreted by airway epithelia, can be induced by various inflammatory mediators such as prostaglandins. The two major prostaglandins involved in inflammation are PGE(2) and PGF(2alpha). PGE(2)-induced mucin production has been well studied, but the effect of PGF(2alpha) on mucin production remains poorly understood. To elucidate the effect and underlying mechanism of PGF(2alpha) on MUC5AC production, we investigated the signal transduction of PGF(2alpha) associated with this effect using normal human tracheobronchial epithelial cells. Our results demonstrated that PGF(2alpha) induces MUC5AC overproduction via a signaling cascade involving protein kinase C, ERK, p90 ribosomal S6 protein kinase, and CREB. The regulation of PGF(2alpha)-induced MUC5AC expression by CREB was further confirmed by cAMP response element-dependent MUC5AC promoter activity and by interaction between CREB and MUC5AC promoter. The abrogation of all downstream signaling activities via suppression of each signaling molecule along the pathway indicates that a single pathway from PGF(2alpha) receptor to CREB is responsible for inducing MUC5AC overproduction. As CREB also mediates mucin overproduction induced by PGE(2) and other inflammatory mediators, our findings have important clinical implications for the management of airway mucus hypersecretion.

Does exhaled nitric oxide predict asthma exacerbation in children?

Asthma is a chronic complex disorder of the respiratory tract that affects millions of people globally, a large percentage of which are children. Triggered by a host of factors such as allergens and changes in temperature, the pathophysiologic and clinical indices vary among patients and have contributed to difficulties in overall management of asthma. Shortly after exhaled nitric oxide (eNO) was discovered in higher concentrations in asthma patients, it was shown to be superior to other markers such as PEFR, FEV1 and sputum eosinophils in screening asthma patients. Studies have also noted promising results regarding the use of eNO to predict asthma exacerbation in adults while in children, asthma symptoms have been observed to be good predictors of asthma exacerbation. Currently however, the potential of eNO as a predictor of asthma exacerbation in children is yet to be examined. The objective of this study was to assess eNO potential to predict asthma exacerbation in children by examining the relationship between eNO and changes in pulmonary function, asthma symptoms and rescue medication use.^ The primary study "Air Toxics and Asthma in Children" (ATAC), recruited children aged 9 to 14 years with labile persistent asthma diagnosed at least one year earlier. The data obtained from 30 study participants, included exhaled nitric oxide concentration, PEFR, FEV1, asthma symptoms and frequency of emergency medication use.^ Descriptive statistics, Pearson's and Spearman's correlation tests were followed by a simple linear regression in which eNO was the independent (predictor) variable while FEV1, PEFR, asthma symptoms and frequency of emergency medication use were the dependent (outcome) variables.^ Results showed that eNO was associated with percent change in FEV1, day time wheeze, night time shortness of breath, but correlated only weakly with PEFR, amplitude percent of mean PEFR, FEV1, percent change in FEV1 and asthma symptoms.^ Further research is imperative to better define the role of eNO and understand intrinsic pathologic mechanisms towards asthma management in children.^

Occupational respiratory disease in the services industry

Objective. To describe the spectrum and occurrence of occupational exposures of relevance to the respiratory system and their subsequent adverse effects within the service industries and occupations, as outlined by the U.S. Department of Labor Bureau of Labor Statistics Occupational Outlook Handbook, 2007. ^ Design. Systematic review of the literature from an Ovid search including years 1950 to 2008. Initially, occupational respiratory disease categories were searched, and then combined with each of the different occupations for a comprehensive review of the literature. ^ Results. Ten groups within the U.S. Department of Labor Bureau of Labor Statistics Occupational Outlook Handbook, 2007 were identified as having exposures leading to occupational respiratory disease. These include janitors/cleaners, dental personnel, cosmetology professionals, traffic police, veterinary personnel, firefighters, healthcare workers, bakers, and bar/restaurant workers. The most common respiratory disorder affecting this population was occupational asthma caused by many different exposures in each occupation. The biggest limitation was the absence of a uniform reporting method for occupational respiratory diseases. ^ Conclusion. There is evidence that there are risks for occupational respiratory disease in the services industry. ^ Key Words: occupational and respiratory disease and service industries ^

Glioma-specific alternative RNA splicing: A role for polypyrimidine tract binding protein-1

Alternative RNA splicing plays an integral role in cell fate determination and function, especially in the cells of the brain. Errors in RNA processing contribute to diseases such as cancer, where it leads to the production of oncogenic proteins or the loss of tumor suppressors. In silica mining suggests that hundreds of splice isoforms are misexpressed in the glial cell-derived glioma. However, there is little experimental evidence of the prevalence and contribution of these changes and whether they contribute to the formation and progression of this devastating malignancy. To determine the frequency of these aberrant events, global profiling of alternative RNA splice patterns in glioma and nontumor brain was conducted using an exon array. Most splicing changes were less than 5-fold in magnitude and 14 cassette exon events were validated, including 7 previously published events. To determine the possible causes of missplicing, the differential expression levels of splicing factors in these two tissues were also analyzed. Six RNA splicing factors had greater than 2-fold changes in expression. The highest differentially expressed factor was polypyrimidine tract binding protein-1 (PTB). Evaluation by immunohistochemistry determined that this factor was elevated in both early and late stages of glioma. Glial cell-specific PTB expression in the adult brain led me to examine the role of PTB in gliomagenesis. Downregulation of PTB slowed glioma cell proliferation and migration and enhanced cell adhesion to fibronectin and vitronectin. To determine whether PTB was affecting these processes through splicing, genome-wide exon expression levels were correlated with PTB levels. Surprisingly, previously reported PTB target transcripts were insensitive to changes in PTB levels in both patient samples and PTB-depleted glioma cells. Only one validated glioma-specific splice target, RTN4/Nogo, had a significant PTB-mediated splicing change. Downregulation of PTB enhanced inclusion of its alternative exon 3, which encodes an auxiliary domain within a neurite inhibitor protein. Overexpression of this splice isoform in glioma cells slowed proliferation in a manner similar to that observed in PTB knockdown cells. In summary, aberrant expression of splicing factors such as PTB in glioma may elicit changes in splicing patterns that enhance tumorigenesis. ^

Occupational respiratory disease in the services industry

Objective. To describe the spectrum and occurrence of occupational exposures of relevance to the respiratory system and their subsequent adverse effects within the service industries and occupations, as outlined by the U.S. Department of Labor Bureau of Labor Statistics Occupational Outlook Handbook, 2007. ^ Design. Systematic review of the literature from an Ovid search including years 1950 to 2008. Initially, occupational respiratory disease categories were searched, and then combined with each of the different occupations for a comprehensive review of the literature. ^ Results. Ten groups within the U.S. Department of Labor Bureau of Labor Statistics Occupational Outlook Handbook, 2007 were identified as having exposures leading to occupational respiratory disease. These include janitors/cleaners, dental personnel, cosmetology professionals, traffic police, veterinary personnel, firefighters, healthcare workers, bakers, and bar/restaurant workers. The most common respiratory disorder affecting this population was occupational asthma caused by many different exposures in each occupation. The biggest limitation was the absence of a uniform reporting method for occupational respiratory diseases. ^ Conclusion. There is evidence that there are risks for occupational respiratory disease in the services industry. ^ Key Words. occupational and respiratory disease and service industries ^

RESPIRATORY COMPENSATION MECHANISMS IN THE UPPER AND LOWER RESPIRATORY TRACTS OF AN ANIMAL MODEL AFTER SUBCHRONIC INHALATION EXPOSURE TO FORMALDEHYDE: IMPLICATIONS FOR TOXICOLOGY RISK ASSESSMENT (DEPOSITION, DOSE RECEIVED, RESPONSE)

The potential for significant human populations to experience long-term inhalation of formaldehyde and reports of symptomatology due to this exposure has led to a considerable interest in the toxicologic assessment of risk from subchronic formaldehyde exposures using animal models. Since formaldehyde inhalation depresses certain respiratory parameters in addition to its other forms of toxicity, there is a potential for the alteration of the actual dose received by the exposed individual (and the resulting toxicity) due to this respiratory effect. The respiratory responses to formaldehyde inhalation and the subsequent pattern of deposition were therefore investigated in animals that had received subchronic exposure to the compound, and the potential for changes in the formaldehyde dose received due to long-term inhalation evaluated. Male Sprague-Dawley rats were exposed to either 0, 0.5, 3, or 15 ppm formaldehyde for 6 hours/day, 5 days/week for up to 6 months. The patterns of respiratory response, deposition and the compensation mechanisms involved were then determined in a series of formaldehyde test challenges to both the upper and to the lower respiratory tracts in separate groups of subchronically exposed animals and age-specific controls (four concentration groups, two time points). In both the control and pre-exposed animals, there was a characteristic recovery of respiratory parameters initially depressed by formaldehyde inhalation to at or approaching pre-exposure levels within 10 minutes of the initiation of exposure. Also, formaldehyde deposition was found to remain very high in the upper and lower tracts after long-term exposure. Therefore, there was probably little subsequent effect on the dose received by the exposed individual that was attributable to the repeated exposures. There was a diminished initial minute volume response in test challenges of both the upper and lower tracts of animals that had received at least 16 weeks of exposure to 15 ppm, with compensatory increases in tidal volume in the upper tract and respiratory rate in the lower tract. However, this dose-related effect was probably not relevant to human risk estimation because this formaldehyde dose is in excess of that experienced by human populations. ^

Epidemiology of Acute Lung Injury and Acute Respiratory Distress Syndrome in children in Vietnam

Acute Lung Injury (ALI) and Acute Respiratory Distress Syndrome (ARDS) are life- threatening disorders that can result from many severe conditions and diseases. Since the American European Consensus Conference established the internationally accepted definition of ALI and ARDS, the epidemiology of pediatric ALI/ARDS has been described in some developed countries. In the developing world, however, there are very few data available regarding the burden, etiologies, management, outcome, and factors associated with outcomes of ALI/ARDS in children. ^ Therefore, we conducted this observational, clinical study to estimate the prevalence and case mortality rate of ALI/ARDS among a cohort of patients admitted to the pediatric intensive care unit (PICU) of the National Hospital of Pediatrics in Hanoi, the largest children's hospital in Vietnam. Etiologies and predisposing factors, and management strategies for pediatric ALI/ARDS were described. In addition, we determined the prevalence of HIV infection among children with ALI/ARDS in Vietnam. We also identified the causes of mortality and predictors of mortality and prolonged mechanical ventilation of children with ALI/ARDS. ^ A total of 1,051 patients consecutively admitted to the pediatric intensive care unit from January 2011 to January 2012 were screened daily for development of ALI/ARDS using the American-European Consensus Conference Guidelines. All identified patients with ALI/ARDS were followed until hospital discharge or death in the hospital. Patients' demographic and clinical data were collected. Multivariable logistic regression models were developed to identify independent predictors of mortality and other adverse outcome of ALI/ARDS. ^ Prevalence of ALI and ARDS was 9.6% (95% confidence interval, 7.8% to 11.4%) and 8.8% (95% confidence interval, 7.0% to 10.5%) of total PICU admissions, respectively. Infectious pneumonia and sepsis were the most common causes of ALI/ARDS accounting for 60.4% and 26.7% of cases, respectively. Prevalence of HIV infection among children with ALI/ARDS was 3.0%. The case fatality rate of ALI/ARDS was 63.4% (95% confidence interval, 53.8% to 72.9%). Multiple organ failure and refractory hypoxemia were the main causes of death. Independent predictors of mortality and prolonged mechanical ventilation were male gender, duration of intensive care stay prior to ALI/ARDS diagnosis, level of oxygenation defect measured by PaO2/FiO2 ratio at ALI/ARDS diagnosis, presence of non-pulmonary organ dysfunction at day one and day three after ALI/ARDS diagnosis, and presence of hospital acquired infection. ^ The results of this study demonstrated that ALI/ARDS was a common and severe condition in children in Vietnam. The level of both pulmonary and non-pulmonary organ damage influenced survival of patients with ALI/ARDS. Strategies for preventing ALI/ARDS and for clinical management of the disease are necessary to reduce the associated risks.^