Browsing by Author "Kraft, Monica"
Now showing 1 - 12 of 12
Results Per Page
Sort Options
Item Open Access Airway fibroblasts in asthma manifest an invasive phenotype.(American journal of respiratory and critical care medicine, 2011-06) Ingram, Jennifer L; Huggins, Molly J; Church, Tony D; Li, Yuejuan; Francisco, Dave C; Degan, Simone; Firszt, Rafael; Beaver, Denise M; Lugogo, Njira L; Wang, Ying; Sunday, Mary E; Noble, Paul W; Kraft, MonicaRationale
Invasive cell phenotypes have been demonstrated in malignant transformation, but not in other diseases, such as asthma. Cellular invasiveness is thought to be mediated by transforming growth factor (TGF)-β1 and matrix metalloproteinases (MMPs). IL-13 is a key T(H)2 cytokine that directs many features of airway remodeling through TGF-β1 and MMPs.Objectives
We hypothesized that, in human asthma, IL-13 stimulates increased airway fibroblast invasiveness via TGF-β1 and MMPs in asthma compared with normal controls.Methods
Fibroblasts were cultured from endobronchial biopsies in 20 subjects with mild asthma (FEV(1): 90 ± 3.6% pred) and 17 normal control subjects (FEV(1): 102 ± 2.9% pred) who underwent bronchoscopy. Airway fibroblast invasiveness was investigated using Matrigel chambers. IL-13 or IL-13 with TGF-β1 neutralizing antibody or pan-MMP inhibitor (GM6001) was added to the lower chamber as a chemoattractant. Flow cytometry and immunohistochemistry were performed in a subset of subjects to evaluate IL-13 receptor levels.Measurements and main results
IL-13 significantly stimulated invasion in asthmatic airway fibroblasts, compared with normal control subjects. Inhibitors of both TGF-β1 and MMPs blocked IL-13-induced invasion in asthma, but had no effect in normal control subjects. At baseline, in airway tissue, IL-13 receptors were expressed in significantly higher levels in asthma, compared with normal control subjects. In airway fibroblasts, baseline IL-13Rα2 was reduced in asthma compared with normal control subjects.Conclusions
IL-13 potentiates airway fibroblast invasion through a mechanism involving TGF-β1 and MMPs. IL-13 receptor subunits are differentially expressed in asthma. These effects may result in IL-13-directed airway remodeling in asthma.Item Open Access Effect of the S-nitrosoglutathione reductase inhibitor N6022 on bronchial hyperreactivity in asthma.(Immunity, inflammation and disease, 2018-06) Que, Loretta G; Yang, Zhonghui; Lugogo, Njira L; Katial, Rohit K; Shoemaker, Steven A; Troha, Janice M; Rodman, David M; Tighe, Robert M; Kraft, MonicaRationale
Patients with asthma demonstrate depletion of the endogenous bronchodilator GSNO and upregulation of GSNOR.Objectives
An exploratory proof of concept clinical study of N6022 in mild asthma to determine the potential bronchoprotective effects of GSNOR inhibition. Mechanistic studies aimed to provide translational evidence of effect.Methods
Fourteen mild asthma patients were treated with intravenous N6022 (5 mg) or placebo and observed for 7 days, with repeated assessments of the provocative dose of methacholine causing a 20% fall in FEV1 (methacholine PC20 FEV1), followed by a washout period and crossover treatment and observation. In vitro studies in isolated eosinophils investigated the effect of GSNO and N6022 on apoptosis.Measurements and main results
This was a negative trial as it failed to reach its primary endpoint, which was change from baseline in methacholine PC20 FEV1 at 24 h. However, our exploratory analysis demonstrated significantly more two dose-doubling increases in PC20 FEV1 for N6022 compared with placebo (21% vs 6%, P < 0.05) over the 7-day observation period. Furthermore, a significant treatment effect was observed in the change in PC20 FEV1 from baseline averaged over the 7-day observation period (mean change: +0.82 mg/ml [N6022] from 1.34 mg/ml [baseline] vs -0.18 mg/ml [placebo] from 1.16 mg/ml [baseline], P = 0.023). N6022 was well tolerated in mild asthmatics. In vitro studies demonstrated enhanced eosinophilic apoptosis with N6022.Conclusions
In this early phase exploratory proof of concept trial in asthma, N6022 did not significantly alter methacholine PC20 FEV1 at 24 h, but did have a treatment effect at 7 days compared to baseline. Further investigation of the efficacy of S-nitrosoglutathione reductase inhibition in a patient population with eosinophilic asthma is warranted.Item Restricted Hyperpolarized Xe MR imaging of alveolar gas uptake in humans.(PLoS One, 2010-08-16) Cleveland, Zackary I; Cofer, Gary P; Metz, Gregory; Beaver, Denise; Nouls, John; Kaushik, S Sivaram; Kraft, Monica; Wolber, Jan; Kelly, Kevin T; McAdams, H Page; Driehuys, BastiaanBACKGROUND: One of the central physiological functions of the lungs is to transfer inhaled gases from the alveoli to pulmonary capillary blood. However, current measures of alveolar gas uptake provide only global information and thus lack the sensitivity and specificity needed to account for regional variations in gas exchange. METHODS AND PRINCIPAL FINDINGS: Here we exploit the solubility, high magnetic resonance (MR) signal intensity, and large chemical shift of hyperpolarized (HP) (129)Xe to probe the regional uptake of alveolar gases by directly imaging HP (129)Xe dissolved in the gas exchange tissues and pulmonary capillary blood of human subjects. The resulting single breath-hold, three-dimensional MR images are optimized using millisecond repetition times and high flip angle radio-frequency pulses, because the dissolved HP (129)Xe magnetization is rapidly replenished by diffusive exchange with alveolar (129)Xe. The dissolved HP (129)Xe MR images display significant, directional heterogeneity, with increased signal intensity observed from the gravity-dependent portions of the lungs. CONCLUSIONS: The features observed in dissolved-phase (129)Xe MR images are consistent with gravity-dependent lung deformation, which produces increased ventilation, reduced alveolar size (i.e., higher surface-to-volume ratios), higher tissue densities, and increased perfusion in the dependent portions of the lungs. Thus, these results suggest that dissolved HP (129)Xe imaging reports on pulmonary function at a fundamental level.Item Open Access IL-13 in asthma and allergic disease: asthma phenotypes and targeted therapies.(The Journal of allergy and clinical immunology, 2012-10) Ingram, Jennifer L; Kraft, MonicaDecades of research in animal models have provided abundant evidence to show that IL-13 is a key T(H)2 cytokine that directs many of the important features of airway inflammation and remodeling in patients with allergic asthma. Several promising focused therapies for asthma that target the IL-13/IL-4/signal transducer and activator of transcription 6 pathway are in development, including anti-IL-13 mAbs and IL-4 receptor antagonists. The efficacy of these new potential asthma therapies depends on the responsiveness of patients. However, an understanding of how IL-13-directed therapies might benefit asthmatic patients is confounded by the complex heterogeneity of the disease. Recent efforts to classify subphenotypes of asthma have focused on sputum cellular inflammation profiles, as well as cluster analyses of clinical variables and molecular and genetic signatures. Researchers and clinicians can now evaluate biomarkers of T(H)2-driven airway inflammation in asthmatic patients, such as serum IgE levels, sputum eosinophil counts, fraction of exhaled nitric oxide levels, and serum periostin levels, to aid decision making in clinical trials and drug development and to identify subsets of patients who might benefit from therapies. Although it is unlikely that these therapies will benefit all asthmatic patients with this heterogeneous disease, advances in understanding asthma subphenotypes in relation to clinical variables and T(H)2 cytokine responses offer the opportunity to improve the efficacy and safety of proposed therapies for asthma.Item Open Access Interleukin-13 induces collagen type-1 expression through matrix metalloproteinase-2 and transforming growth factor-β1 in airway fibroblasts in asthma.(The European respiratory journal, 2014-02) Firszt, Rafael; Francisco, Dave; Church, Tony D; Thomas, Joseph M; Ingram, Jennifer L; Kraft, MonicaAirway remodelling is a feature of asthma that contributes to loss of lung function. One of the central components of airway remodelling is subepithelial fibrosis. Interleukin (IL)-13 is a key T-helper 2 cytokine and is believed to be the central mediator of allergic asthma including remodelling, but the mechanism driving the latter has not been elucidated in human asthma. We hypothesised that IL-13 stimulates collagen type-1 production by the airway fibroblast in a matrix metalloproteinase (MMP)- and transforming growth factor (TGF)-β1-dependent manner in human asthma as compared to healthy controls. Fibroblasts were cultured from endobronchial biopsies in 14 subjects with mild asthma and 13 normal controls that underwent bronchoscopy. Airway fibroblasts were treated with various mediators including IL-13 and specific MMP-inhibitors. IL-13 significantly stimulated collagen type-1 production in asthma compared to normal controls. Inhibitors of MMP-2 significantly attenuated collagen production in asthma but had no effect in normal controls. IL-13 significantly increased total and active forms of TGF-β1, and this activation was blocked using an MMP-2 inhibitor. IL-13 activated endogenous MMP-2 in asthma patients as compared to normal controls. In an ex vivo model, IL-13 potentiates airway remodelling through a mechanism involving TGF-β1 and MMP-2. These effects provide insights into the mechanism involved in IL-13-directed airway remodelling in asthma.Item Open Access Microfluidic platform versus conventional real-time polymerase chain reaction for the detection of Mycoplasma pneumoniae in respiratory specimens.(Diagn Microbiol Infect Dis, 2010-05) Wulff-Burchfield, Elizabeth; Schell, Wiley A; Eckhardt, Allen E; Pollack, Michael G; Hua, Zhishan; Rouse, Jeremy L; Pamula, Vamsee K; Srinivasan, Vijay; Benton, Jonathan L; Alexander, Barbara D; Wilfret, David A; Kraft, Monica; Cairns, Charles B; Perfect, John R; Mitchell, Thomas GRapid, accurate diagnosis of community-acquired pneumonia (CAP) due to Mycoplasma pneumoniae is compromised by low sensitivity of culture and serology. Polymerase chain reaction (PCR) has emerged as a sensitive method to detect M. pneumoniae DNA in clinical specimens. However, conventional real-time PCR is not cost-effective for routine or outpatient implementation. Here, we evaluate a novel microfluidic real-time PCR platform (Advanced Liquid Logic, Research Triangle Park, NC) that is rapid, portable, and fully automated. We enrolled patients with CAP and extracted DNA from nasopharyngeal wash (NPW) specimens using a biotinylated capture probe and streptavidin-coupled magnetic beads. Each extract was tested for M. pneumoniae-specific DNA by real-time PCR on both conventional and microfluidic platforms using Taqman probe and primers. Three of 59 (5.0%) NPWs were positive, and agreement between the methods was 98%. The microfluidic platform was equally sensitive but 3 times faster and offers an inexpensive and convenient diagnostic test for microbial DNA.Item Open Access Obese asthmatic patients have decreased surfactant protein A levels: Mechanisms and implications.(The Journal of allergy and clinical immunology, 2018-03) Lugogo, Njira; Francisco, Dave; Addison, Kenneth J; Manne, Akarsh; Pederson, William; Ingram, Jennifer L; Green, Cynthia L; Suratt, Benjamin T; Lee, James J; Sunday, Mary E; Kraft, Monica; Ledford, Julie GBackground
Eosinophils are prominent in some patients with asthma and are increased in the submucosa in a subgroup of obese patients with asthma (OAs). Surfactant protein A (SP-A) modulates host responses to infectious and environmental insults.Objective
We sought to determine whether SP-A levels are altered in OAs compared with a control group and to determine the implications of these alterations in SP-A levels in asthmatic patients.Methods
Bronchoalveolar lavage fluid from 23 lean, 12 overweight, and 20 obese subjects were examined for SP-A. Mouse tracheal epithelial cells grown at an air-liquid interface were used for mechanistic studies. SP-A-/- mice were challenged in allergen models, and exogenous SP-A therapy was given after the last challenge. Eosinophils were visualized and quantitated in lung parenchyma by means of immunostaining.Results
Significantly less SP-A (P = .002) was detected in samples from OAs compared with those from control subjects. A univariable regression model found SP-A levels were significantly negatively correlated with body mass index (r = -0.33, P = .014), whereas multivariable modeling demonstrated that the correlation depended both on asthma status (P = .017) and the interaction of asthma and body mass index (P = .008). Addition of exogenous TNF-α to mouse tracheal epithelial cells was sufficient to attenuate SP-A and eotaxin secretion. Allergen-challenged SP-A-/- mice that received SP-A therapy had significantly less tissue eosinophilia compared with mice receiving vehicle.Conclusions
SP-A functions as an important mediator in resolving tissue and lavage fluid eosinophilia in allergic mouse models. Decreased levels of SP-A in OAs, which could be due to increased local TNF-α levels, might lead to impaired eosinophil resolution and could contribute to the eosinophilic asthma phenotype.Item Open Access Role of hyaluronan and hyaluronan-binding proteins in human asthma.(The Journal of allergy and clinical immunology, 2011-08) Liang, Jiurong; Jiang, Dianhua; Jung, Yoosun; Xie, Ting; Ingram, Jennifer; Church, Tony; Degan, Simone; Leonard, Maura; Kraft, Monica; Noble, Paul WBackground
The characteristics of human asthma are chronic inflammation and airway remodeling. Hyaluronan, a major extracellular matrix component, accumulates during inflammatory lung diseases, including asthma. Hyaluronan fragments stimulate macrophages to produce inflammatory cytokines. We hypothesized that hyaluronan and its receptors would play a role in human asthma.Objective
To investigate the role of hyaluronan and hyaluronan-binding proteins in human asthma.Methods
Twenty-one subjects with asthma and 25 healthy control subjects underwent bronchoscopy with endobronchial biopsy and bronchoalveolar lavage. Fibroblasts were cultured, and hyaluronan and hyaluronan synthase expression was determined at baseline and after exposure to several mediators relevant to asthma pathobiology. The expression of hyaluronan-binding proteins CD44, TLR (Toll-like receptor)-2, and TLR4 on bronchoalveolar lavage macrophages was determined by flow cytometry. IL-8 production by macrophages in response to hyaluronan fragment stimulation was compared.Results
Airway fibroblasts from patients with asthma produced significantly increased concentrations of lower-molecular-weight hyaluronan compared with those of normal fibroblasts. Hyaluronan synthase 2 mRNA was markedly increased in asthmatic fibroblasts. Asthmatic macrophages showed a decrease in cell surface CD44 expression and an increase in TLR2 and TLR4 expression. Macrophages from subjects with asthma showed an increase in responsiveness to low-molecular-weight hyaluronan stimulation, as demonstrated by increased IL-8 production.Conclusion
Hyaluronan homeostasis is deranged in asthma, with increased production by fibroblasts and decreased CD44 expression on alveolar macrophages. Upregulation of TLR2 and TLR4 on macrophages with increased sensitivity to hyaluronan fragments suggests a novel proinflammatory mechanism by which persistence of hyaluronan fragments could contribute to chronic inflammation and airway remodeling in asthma.Item Open Access Role of Matrix Metalloproteinases-1 and -2 in Interleukin-13-Suppressed Elastin in Airway Fibroblasts in Asthma.(American journal of respiratory cell and molecular biology, 2016-01) Ingram, Jennifer L; Slade, David; Church, Tony D; Francisco, Dave; Heck, Karissa; Sigmon, R Wesley; Ghio, Michael; Murillo, Anays; Firszt, Rafael; Lugogo, Njira L; Que, Loretta; Sunday, Mary E; Kraft, MonicaElastin synthesis and degradation in the airway and lung parenchyma contribute to airway mechanics, including airway patency and elastic recoil. IL-13 mediates many features of asthma pathobiology, including airway remodeling, but the effects of IL-13 on elastin architecture in the airway wall are not known. We hypothesized that IL-13 modulates elastin expression in airway fibroblasts from subjects with allergic asthma. Twenty-five subjects with mild asthma (FEV1, 89 ± 3% predicted) and 30 normal control subjects (FEV1, 102 ± 2% predicted) underwent bronchoscopy with endobronchial biopsy. Elastic fibers were visualized in airway biopsy specimens using Weigert's resorcin-fuchsin elastic stain. Airway fibroblasts were exposed to IL-13; a pan-matrix metalloproteinase (MMP) inhibitor (GM6001); specific inhibitors to MMP-1, -2, -3, and -8; and combinations of IL-13 with MMP inhibitors in separate conditions in serum-free media for 48 hours. Elastin (ELN) expression as well as MMP secretion and activity were quantified. Results of this study show that elastic fiber staining of airway biopsy tissue was significantly associated with methacholine PC20 (i.e., the provocative concentration of methacholine resulting in a 20% fall in FEV1 levels) in patients with asthma. IL-13 significantly suppressed ELN expression in asthmatic airway fibroblasts as compared with normal control fibroblasts. The effect of IL-13 on ELN expression was significantly correlated with postbronchodilator FEV1/FVC in patients with asthma. MMP inhibition significantly stimulated ELN expression in patients with asthma as compared with normal control subjects. Specific inhibition of MMP-1 and MMP-2, but not MMP-3 or MMP-8, reversed the IL-13-induced suppression of ELN expression. In asthma, MMP-1 and MMP-2 mediate IL-13-induced suppression of ELN expression in airway fibroblasts.Item Open Access SHP-1 as a critical regulator of Mycoplasma pneumoniae-induced inflammation in human asthmatic airway epithelial cells.(Journal of immunology (Baltimore, Md. : 1950), 2012-04) Wang, Ying; Zhu, Zhou; Church, Tony D; Lugogo, Njira L; Que, Loretta G; Francisco, Dave; Ingram, Jennifer L; Huggins, Molly; Beaver, Denise M; Wright, Jo Rae; Kraft, MonicaAsthma is a chronic inflammatory disease in which airway epithelial cells are the first line of defense against exposure of the airway to infectious agents. Src homology protein (SHP)-1, a protein tyrosine phosphatase, is a negative regulator of signaling pathways that are critical to the development of asthma and host defense. We hypothesize that SHP-1 function is defective in asthma, contributing to the increased inflammatory response induced by Mycoplasma pneumoniae, a pathogen known to exacerbate asthma. M. pneumoniae significantly activated SHP-1 in airway epithelial cells collected from nonasthmatic subjects by bronchoscopy with airway brushing but not in cells from asthmatic subjects. In asthmatic airway epithelial cells, M. pneumoniae induced significant PI3K/Akt phosphorylation, NF-κB activation, and IL-8 production compared with nonasthmatic cells, which were reversed by SHP-1 overexpression. Conversely, SHP-1 knockdown significantly increased IL-8 production and PI3K/Akt and NF-κB activation in the setting of M. pneumoniae infection in nonasthmatic cells, but it did not exacerbate these three parameters already activated in asthmatic cells. Thus, SHP-1 plays a critical role in abrogating M. pneumoniae-induced IL-8 production in nonasthmatic airway epithelial cells through inhibition of PI3K/Akt and NF-κB activity, but it is defective in asthma, resulting in an enhanced inflammatory response to infection.Item Open Access Surfactant protein A is defective in abrogating inflammation in asthma.(American journal of physiology. Lung cellular and molecular physiology, 2011-10) Wang, Ying; Voelker, Dennis R; Lugogo, Njira L; Wang, Guirong; Floros, Joanna; Ingram, Jennifer L; Chu, Hong Wei; Church, Tony D; Kandasamy, Pitchaimani; Fertel, Daniel; Wright, Jo Rae; Kraft, MonicaSurfactant protein A (SP-A) regulates a variety of immune cell functions. We determined the ability of SP-A derived from normal and asthmatic subjects to modulate the inflammatory response elicited by Mycoplasma pneumoniae, a pathogen known to exacerbate asthma. Fourteen asthmatic and 10 normal control subjects underwent bronchoscopy with airway brushing and bronchoalveolar lavage (BAL). Total SP-A was extracted from BAL. The ratio of SP-A1 to total SP-A (SP-A1/SP-A) and the binding of total SP-A to M. pneumoniae membranes were determined. Airway epithelial cells from subjects were exposed to either normal or asthmatic SP-A before exposure to M. pneumoniae. IL-8 protein and MUC5AC mRNA were measured. Total BAL SP-A concentration did not differ between groups, but the percentage SP-A1 was significantly increased in BAL of asthmatic compared with normal subjects. SP-A1/SP-A significantly correlated with maximum binding of total SP-A to M. pneumoniae, but only in asthma. SP-A derived from asthmatic subjects did not significantly attenuate IL-8 and MUC5AC in the setting of M. pneumoniae infection compared with SP-A derived from normal subjects. We conclude that SP-A derived from asthmatic subjects does not abrogate inflammation effectively, and this dysfunction may be modulated by SP-A1/SP-A.Item Open Access Targeted HAS2 Expression Lessens Airway Responsiveness in Chronic Murine Allergic Airway Disease.(American journal of respiratory cell and molecular biology, 2017-12) Walker, Julia KL; Theriot, Barbara S; Ghio, Michael; Trempus, Carol S; Wong, Jordan E; McQuade, Victoria L; Liang, Jiurong; Jiang, Dianhua; Noble, Paul W; Garantziotis, Stavros; Kraft, Monica; Ingram, Jennifer LHyaluronan (HA), a major component of the extracellular matrix, is secreted by airway structural cells. Airway fibroblasts in allergic asthma secrete elevated levels of HA in association with increased HA synthase 2 (HAS2) expression. Thus, we hypothesized that HA accumulation in the airway wall may contribute to airway remodeling and hyperresponsiveness in allergic airways disease. To examine this hypothesis, transgenic mice in which the α-smooth muscle actin (α-SMA) promoter drives HAS2 expression were generated. Mixed male and female α-SMA-HAS2 mice (HAS2+ mice, n = 16; HAS2- mice, n = 13) were sensitized via intraperitoneal injection and then chronically challenged with aerosolized ovalbumin (OVA) for 6 weeks. To test airway responsiveness, increasing doses of methacholine were delivered intravenously and airway resistance was measured using the forced oscillation technique. HA, cytokines, and cell types were analyzed in bronchoalveolar lavage fluid, serum, and whole lung homogenates. Lung sections were stained using antibodies specific for HA-binding protein (HABP) and α-SMA, as well as Masson's trichrome stain. Staining of lung tissue demonstrated significantly increased peribronchial HA, α-SMA, and collagen deposition in OVA-challenged α-SMA-HAS2+ mice compared with α-SMA-HAS2- mice. Unexpectedly, OVA-challenged α-SMA-HAS2+ mice displayed significantly reduced airway responsiveness to methacholine compared with similarly treated α-SMA-HAS2- mice. The total numbers of inflammatory cell types in the bronchoalveolar lavage fluid did not differ significantly between OVA-challenged α-SMA-HAS2+ mice and α-SMA-HAS2- mice. We conclude that allergen-challenged mice that overexpress HAS2 in myofibroblasts and smooth muscle cells develop increased airway fibrosis, which lessens airway hyperresponsiveness to bronchoconstrictors.