Lung Computational Models and the Role of the Small Airways in Asthma.

Rationale: Asthma is characterized by disease within the small airways. Several studies have suggested that forced oscillation technique-derived resistance at 5 Hz (R5) - resistance at 20 Hz (R20) is a measure of small airway disease; however, there has been limited validation of this measurement to date.Objectives: To validate the use of forced oscillation R5 - R20 as a measure of small airway narrowing in asthma, and to investigate the role that small airway narrowing plays in asthma.Methods: Patient-based complete conducting airway models were generated from computed tomography scans to simulate the impact of different degrees of airway narrowing at different levels of the airway tree on forced oscillation R5 - R20 (n = 31). The computational models were coupled with regression models in an asthmatic cohort (n = 177) to simulate the impact of small airway narrowing on asthma control and quality of life. The computational models were used to predict the impact on small airway narrowing of type-2 targeting biologics using pooled data from two similarly design randomized, placebo-controlled biologic trials (n = 137).Measurements and Main Results: Simulations demonstrated that narrowing of the small airways had a greater impact on R5 - R20 than narrowing of the larger airways and was associated (above a threshold of approximately 40% narrowing) with marked deterioration in both asthma control and asthma quality of life, above the minimal clinical important difference. The observed treatment effect on R5 - R20 in the pooled trials equated to a predicted small airway narrowing reversal of approximately 40%.Conclusions: We have demonstrated, using computational modeling, that forced oscillation R5 - R20 is a direct measure of anatomical narrowing in the small airways and that small airway narrowing has a marked impact on both asthma control and quality of life and may be modified by biologics.

Functional CT imaging for identification of the spatial determinants of small-airways disease in adults with asthma.

BACKGROUND: Asthma is a disease characterized by ventilation heterogeneity (VH). A number of studies have demonstrated that VH markers derived by using impulse oscillometry (IOS) or multiple-breath washout (MBW) are associated with key asthmatic patient-related outcome measures and airways hyperresponsiveness. However, the topographical mechanisms of VH in the lung remain poorly understood. OBJECTIVES: We hypothesized that specific regionalization of topographical small-airway disease would best account for IOS- and MBW-measured indices in patients. METHODS: We evaluated the results of paired expiratory/inspiratory computed tomography in a cohort of asthmatic (n = 41) and healthy (n = 11) volunteers to understand the determinants of clinical VH indices commonly reported by using IOS and MBW. Parametric response mapping (PRM) was used to calculate the functional small-airways disease marker PRMfSAD and Hounsfield unit (HU)-based density changes from total lung capacity to functional residual capacity (ΔHU); gradients of ΔHU in gravitationally perpendicular (parallel) inferior-superior (anterior-posterior) axes were quantified. RESULTS: The ΔHU gradient in the inferior-superior axis provided the highest level of discrimination of both acinar VH (measured by using phase 3 slope analysis of multiple-breath washout data) and resistance at 5 Hz minus resistance at 20 Hz measured by using impulse oscillometry (R5-R20) values. Patients with a high inferior-superior ΔHU gradient demonstrated evidence of reduced specific ventilation in the lower lobes of the lungs and high levels of PRMfSAD. A computational small-airway tree model confirmed that constriction of gravitationally dependent, lower-zone, small-airway branches would promote the largest increases in R5-R20 values. Ventilation gradients correlated with asthma control and quality of life but not with exacerbation frequency. CONCLUSIONS: Lower lobe-predominant small-airways disease is a major driver of clinically measured VH in adults with asthma.

Airway pathological heterogeneity in asthma: Visualization of disease microclusters using topological data analysis.

BACKGROUND: Asthma is a complex chronic disease underpinned by pathological changes within the airway wall. How variations in structural airway pathology and cellular inflammation contribute to the expression and severity of asthma are poorly understood. OBJECTIVES: Therefore we evaluated pathological heterogeneity using topological data analysis (TDA) with the aim of visualizing disease clusters and microclusters. METHODS: A discovery population of 202 adult patients (142 asthmatic patients and 60 healthy subjects) and an external replication population (59 patients with severe asthma) were evaluated. Pathology and gene expression were examined in bronchial biopsy samples. TDA was applied by using pathological variables alone to create pathology-driven visual networks. RESULTS: In the discovery cohort TDA identified 4 groups/networks with multiple microclusters/regions of interest that were masked by group-level statistics. Specifically, TDA group 1 consisted of a high proportion of healthy subjects, with a microcluster representing a topological continuum connecting healthy subjects to patients with mild-to-moderate asthma. Three additional TDA groups with moderate-to-severe asthma (Airway Smooth MuscleHigh, Reticular Basement MembraneHigh, and RemodelingLow groups) were identified and contained numerous microclusters with varying pathological and clinical features. Mutually exclusive TH2 and TH17 tissue gene expression signatures were identified in all pathological groups. Discovery and external replication applied to the severe asthma subgroup identified only highly similar "pathological data shapes" through analyses of persistent homology. CONCLUSIONS: We have identified and replicated novel pathological phenotypes of asthma using TDA. Our methodology is applicable to other complex chronic diseases.

Operable non-small cell lung cancer: correlation of volumetric helical dynamic contrast-enhanced CT parameters with immunohistochemical markers of tumor hypoxia.

PURPOSE: To assess the relationship between helical dynamic contrast material-enhanced (DCE) computed tomographic (CT) parameters and immunohistochemical markers of hypoxia in patients with operable non-small cell lung cancer (NSCLC). MATERIALS AND METHODS: After institutional review board approval was obtained, 20 prospective patients who were suspected of having NSCLC underwent whole-tumor DCE CT with kinetic modeling (Patlak analysis) 24 hours before scheduled surgery. Flow-extraction product (in milliliters per 100 milliliters per minute) and blood volume (in milliliters per 100 milliliters) were derived. After surgery, matched whole-tumor sections were stained for exogenous and endogenous markers of hypoxia (pimonidazole infused intravenously 24 hours before surgery, immediately after DCE CT; glucose transporter protein). Correlation between DCE CT parameters and immunohistochemical markers was assessed by using the Spearman rank correlation. DCE CT parameters and immunohistochemical markers were also compared according to pathologic subtype, grade, stage, and nodal status by using the Mann-Whitney test. P values less than .05 indicated a statistically significant difference. RESULT: Fourteen patients with confirmed primary NSCLC underwent resection. There were negative correlations between blood volume and pimonidazole staining (r = -0.48, P = .004), and between flow-extraction product and glucose transporter protein expression (r = -0.50, P = .002). Flow-extraction product was significantly higher in adenocarcinomas than in squamous cell tumors (17.73 vs 11.46; P = .043). Glucose transporter protein expression was significantly lower for adenocarcinomas than for squamous tumors (14.07 vs 33.03; P < .001) and in node negative than in node positive tumors (15.63 vs 23.85; P = .005). CONCLUSION: Blood volume and flow-extraction product derived at DCE CT correlated negatively with pimonidazole and glucose transporter protein expression, indicating the potential of these CT parameters as imaging biomarkers of hypoxia.

The Role of Diet Compared to Physical Activity on Women's Cancer Mortality: Results From the Third National Health and Nutrition Examination Survey.

Background: Among women in the United States, cancer is the second leading cause of death. Prior studies have examined how lifestyle factors, such as diet and physical activity, influence cancer mortality. However, few have evaluated if diet or physical activity has a stronger protective effect for cancer mortality. Therefore, this study aims to evaluate and compare the impacts of diet and physical activity on women's cancer mortality. Methods: Prospective, cross-sectional data were abstracted from the Third US National Health and Nutrition Examination Survey (NHANES III) on female respondents from 1988 to 1994. Physical activity was derived from the CDC's metabolic equivalent (MET) intensity levels. Dietary classifications were derived from the USDA's healthy eating index (HEI). We utilized the National Death Index to obtain mortality follow-up information on our cohort until December 31, 2015. Chi-squared, multivariable Cox regression, and Kaplan-Meier estimates were employed for statistical analyses. Results: Of 3,590 women (median age: 57, range: 40-89), 30% had an obese BMI (BMI≥30 kg/m2). Additionally, 22% of participants self-reported a healthy diet, 69% needed dietary improvement, and 9% had a poor diet. Furthermore, 21% reported physical inactivity, 44% did not meet physical activity guidelines, and 35% met guidelines. On multivariate analysis, healthy diet (HR: 0.70; 95% CI: 0.51-0.98; p = 0.04), but not physical activity (HR: 0.87; 95% CI: 0.55-1.38; p = 0.55), independently predicted for lower cancer mortality. Participants with a healthy diet but low exercise had decreased cancer mortality compared to participants with an unhealthy diet but high exercise (p = 0.01). Conclusions: A healthful diet was associated with lower cancer mortality in women, even after adjusting for obesity, inflammation, and other covariates. In addition, diet may play a stronger role in reducing cancer mortality in women than physical activity.

Induction of heart valve lesions by small-molecule ALK5 inhibitors.

Aberrant signaling by transforming growth factor-ß (TGF-ß) and its type I (ALK5) receptor has been implicated in a number of human diseases and this pathway is considered a potential target for therapeutic intervention. Transforming growth factor-ß signaling via ALK5 plays a critical role during heart development, but the role of ALK5 in the adult heart is poorly understood. In the current study, the preclinical toxicology of ALK5 inhibitors from two different chemistry scaffolds was explored. Ten-week-old female Han Wistar rats received test compounds by the oral route for three to seven days. Both compounds induced histopathologic heart valve lesions characterized by hemorrhage, inflammation, degeneration, and proliferation of valvular interstitial cells. The pathology was observed in all animals, at all doses tested, and occurred in all four heart valves. Immunohistochemical analysis of ALK5 in rat hearts revealed expression in the valves, but not in the myocardium. Compared to control animals, protein levels of ALK5 were unchanged in the heart valves of treated animals. We also observed a physeal dysplasia in the femoro-tibial joint of rats treated with ALK5 inhibitors, a finding consistent with a pharmacological effect described previously with ALK5 inhibitors. Overall, these findings suggest that TGF-ß signaling via ALK5 plays a critical role in maintaining heart valve integrity.

Recombinant expression of aryl hydrocarbon receptor for quantitative ligand-binding analysis.

Recombinant expression of the aryl hydrocarbon receptor (AhR) yields small amounts of ligand-binding-competent AhR. Therefore, Spodoptera frugiperda (Sf9) cells and baculovirus have been evaluated for high-level and functional expression of AhR. Rat and human AhR were expressed as soluble protein in significant amounts. Expression of ligand-binding-competent AhR was sensitive to the protein concentration of Sf9 extract, and coexpression of the chaperone p23 failed to affect the yield of functional ligand-binding AhR. The expression system yielded high levels of functional protein, with the ligand-binding capacity (Bmax) typically 20-fold higher than that obtained with rat liver cytosol. Quantitative estimates of the ligand-binding affinity of human and rat AhR were obtained; the Kd for recombinant rat AhR was indistinguishable from that of native rat AhR, thereby validating the expression system as a faithful model for native AhR. The human AhR bound TCDD with significantly lower affinity than the rat AhR. These findings demonstrate high-level expression of ligand-binding-competent AhR, and sufficient AhR for quantitative analysis of ligand binding.

APPL1 associates with TrkA and GIPC1 and is required for nerve growth factor-mediated signal transduction.

The neurotrophin receptor TrkA plays critical roles in the nervous system by recruiting signaling molecules that activate pathways required for the growth and survival of neurons. Here, we report APPL1 as a TrkA-associated protein. APPL1 and TrkA co-immunoprecipitated in sympathetic neurons. We have identified two routes through which this association can occur. APPL1 was isolated as a binding partner for the TrkA-interacting protein GIPC1 from rat brain lysate by mass spectrometry. The PDZ domain of GIPC1 directly engaged the C-terminal sequence of APPL1. This interaction provides a means through which APPL1 may be recruited to TrkA. In addition, the APPL1 PTB domain bound to TrkA, indicating that APPL1 may associate with TrkA independently of GIPC1. Isolation of endosomal fractions by high-resolution centrifugation determined that APPL1, GIPC1, and phosphorylated TrkA are enriched in the same fractions. Reduction of APPL1 or GIPC1 protein levels suppressed nerve growth factor (NGF)-dependent MEK, extracellular signal-regulated kinase, and Akt activation and neurite outgrowth in PC12 cells. Together, these results indicate that GIPC1 and APPL1 play a role in TrkA function and suggest that a population of endosomes bearing a complex of APPL1, GIPC1, and activated TrkA may transmit NGF signals.

Assessing small airways dysfunction in asthma, asthma remission and healthy controls using particles in exhaled air.

PExA mass can distinguish asthmatics from healthy individuals. Subjects with complete, but not clinical, asthma remission exhale more PExA mass compared to asthma. Higher PExA mass was associated with better function of both the small and large airways. http://bit.ly/2znHABg.

Limited utility of endomyocardial biopsy in the first year after heart transplantation.

BACKGROUND: Surveillance endomyocardial biopsies (EMBs) are used for the early diagnosis of acute cardiac allograft rejection. Protocols became standardized in an earlier era and their utility with contemporary immunosuppression has not been investigated. METHODS: We studied 258 patients after orthotopic heart transplantation comparing 135 patients immunosuppressed by mycophenolate mofetil (MMF) with 123 patients treated by azathioprine (AZA); both with cyclosporine and corticosteroids after induction therapy with rabbit antithymocyte globulin. Fifteen EMBs were scheduled in the first year. Additional EMBs were performed for suspected rejection, after treatment, or for inadequate samples. The MMF group had 1875 EMBs vs. 1854 in the AZA group. RESULTS: The yield of International Society for Heart and Lung Transplantation (ISHLT) grade> or =3A biopsy-proven acute rejection (BPAR) was 1.87% per biopsy (35 of 1875) with MMF vs. 3.13% (58 of 1854) with AZA P=0.024. The number of clinically silent BPAR ISHLT grade > or =3A (the true yield of surveillance EMBs) was 1.39% (26 of 1875) of biopsies MMF vs. 2.1% (39 of 1854) AZA, P=0.48. There were five serious complications requiring intervention or causing long-term sequelae; 0.13% (5 of 3729) per biopsy and 1.94% (5 of 258) per patient. The incidence of all definite and potential complications was 1.42% (53 of 3729) per biopsy and 20.5% (53 of 258) per patient. There was no biopsy-related mortality. CONCLUSION: The yield of BPAR was low in the AZA group and very low in the MMF group. The incidence of complications was also low, but repeated biopsies led to a higher rate per patient. Routine surveillance EMBs and the frequency of such biopsies should be reevaluated in the light of their low yield with current immunosuppression.

Sarcoid heart disease.

To this day the aetiology of sarcoidosis continues to elude definition. Partially as a consequence of this, little in the way of new therapies has evolved. The enigma of this condition is that, unusually for a disease with the potential for devastating consequences, many patients show spontaneous resolution and recover. Cardiac involvement can affect individuals of any age, gender or race and has a predilection for the conduction system of the heart. Heart involvement can also cause a dilated cardiomyopathy with consequent progressive heart failure. The most common presentation of this systemic disease is with pulmonary infiltration, but many cases will be asymptomatic and are detected on routine chest radiography revealing lymphadenopathy. Current advances lie in the newer methods of imaging and diagnosing this unusual heart disease. This review describes the pathology and diagnosis of this condition and the newer imaging techniques that have developed for determining cardiac involvement.

A pilot evaluation of the use of tissue microarrays for quantitation of target distribution in drug discovery pathology.

The use of tissue microarrays (TMAs) in the determination of novel target molecule distribution in organs is an expanding area of discovery pathology. This pilot study was carried out to assess the Chromavision automated cellular imaging system (ACIS) for quantitation of both mRNA and protein distribution in rat and dog TMAs. The targets chosen were a protein kinase, P-CIP2, for mRNA assessment and its downsteam target, peptidylglycine amidating monoxygenase (PAM), for immunohistochemistry (IHC). Oligonucleotide probes produced against P-CIP2, together with an antibody against PAM, were evaluated on rat and dog TMAs. A method for evaluation of target distribution using the ACIS was developed and involved a two-tier approach. Firstly, an initial scanning of the labelled slides identified which tissues expressed the target. Secondly, a more comprehensive analysis was made. This required operator interaction to select specific regions of interest within selected tissue cores and exclude any background labelling from the final assessment. This exacted the level of expression of P-CIP2 or PAM in different cellular populations in tissue cores. A comparative semi-quantitative analysis of the same arrays was concomitantly made by the pathologist in order to assess the relative benefits of a potentially time-consuming detailed morphological evaluation. This involved the histological identification by the pathologist of specific cell populations expressing P-CIP2 or PAM. In this study, we demonstrate the power of an image analysing system to provide quantitative data on target distribution by in situ hybridisation and IHC on normal TMAs. This methodology, together with detailed histological analysis by a pathologist, forms a guideline for future target distribution evaluation within discovery pathology.

Neutrophil dysplasia (acquired pseudo-pelger anomaly) caused by ganciclovir.

Two thoracic organ-transplant recipients with persistent cytomegalovirus infection developed morphologic abnormalities in their circulating neutrophils while receiving either ganciclovir or its prodrug valganciclovir. Neither patient was receiving concomitant treatment with mycophenolate mofetil or azathioprine. In both patients, the morphologic abnormalities preceded the development of neutropenia and bone-marrow hypoplasia. The morphologic changes most likely reflected interference of DNA polymerization within bone marrow progenitor cells. The changes resolved completely after the drug was withdrawn.

Multiphoton crosslinking for biocompatible 3D printing of type I collagen.

Multiphoton fabrication is a powerful technique for three-dimensional (3D) printing of structures at the microscale. Many polymers and proteins have been successfully structured and patterned using this method. Type I collagen comprises a large part of the extracellular matrix for most tissue types and is a widely used cellular scaffold material for tissue engineering. Current methods for creating collagen tissue scaffolds do not allow control of local geometry on a cellular scale. This means the environment experienced by cells may be made up of the native material but unrelated to native cellular-scale structure. In this study, we present a novel method to allow multiphoton crosslinking of type I collagen with flavin mononucleotide photosensitizer. The method detailed allows full 3D printing of crosslinked structures made from unmodified type I collagen and uses only demonstrated biocompatible materials. Resolution of 1 µm for both standing lines and high-aspect ratio gaps between structures is demonstrated and complex 3D structures are fabricated. This study demonstrates a means for 3D printing with one of the most widely used tissue scaffold materials. High-resolution, 3D control of the fabrication of collagen scaffolds will facilitate higher fidelity recreation of the native extracellular environment for engineered tissues.

Cardiotoxicity associated with targeting kinase pathways in cancer.

Cardiotoxicity, also referred to as drug-induced cardiac injury, is an issue associated with the use of some small-molecule kinase inhibitors and antibody-based therapies targeting signaling pathways in cancer. Although these drugs have had a major impact on cancer patient survival, data have implicated kinase-targeting agents such as sunitinib, imatinib, trastuzumab, and sorafenib in adversely affecting cardiac function in a subset of treated individuals. In many cases, adverse cardiac events in the clinic were not anticipated based on preclinical safety evaluation of the molecule. In order to support the development of efficacious and safe kinase inhibitors for the treatment of cancer and other indications, new preclinical approaches and screens are required to predict clinical cardiotoxicity. Laboratory investigations into the underlying molecular mechanisms of heart toxicity induced by these molecules have identified potentially common themes including mitochondrial perturbation and modulation of adenosine monophosphate-activated protein kinase activity. Studies characterizing cardiac-specific kinase knockout mouse models have developed our understanding of the homeostatic role of some of these signaling mediators in the heart. Therefore, when considering kinases as potential future targets or when examining secondary pharmacological interactions of novel kinase inhibitors, these models may help to inform us of the potential adverse cardiac effects in the clinic.

Improved united-atom force field for 1-alkyl-3-methylimidazolium chloride.

We have developed a united atom (UA) nonpolarizable force field for 1-alkyl-3-methyl-imidazolium chloride ([C(n)mim][Cl], n = 1, 2, 4, 6, 8), a potential solvent for the pretreatment of lignocellulosic biomass. The charges were assigned by fitting the electrostatic potential surface (ESP) of the ion pair dimers. The Lennard-Jones parameters of the hydrogen atoms on the imidazolium ring were adjusted to agree with the ab initio optimized geometries of isolated ion pairs. Molecular dynamics (MD) simulations were performed for a wide range of temperatures to validate the force field. Substantial improvements were found in both the dynamical properties and the fluid structures, as compared to those predicted using our previously developed UA force field (UA2006) (Phys. Chem. Chem. Phys. 2006, 8, 1096). Liquid densities were found to lie within 2% experimental data. The simulated heats of vaporization decreased about 30% relative to that predicted using the UA2006 force field. The site-site radial distribution functions between the hydrogen atoms on the imidazolium ring and the chloride anions were in good agreement with those determined by ab initio molecular dynamics. The newly developed force field gives a much better description of the self-diffusion coefficients and shear viscosities, which usually deviate by 1 order of magnitude when determined using other force fields.