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1.  Efficacy of an In-home Test Kit in Reducing Dust Mite Allergen Levels: Results of a Randomized Controlled Pilot Study 
Dust mite allergens can induce allergic sensitization and exacerbate asthma symptoms. Although dust mite reduction and control strategies exist, few asthmatics employ them.
We examined whether an in-home test kit, which quantifies dust mite allergen levels, resulted in behavioral changes in implementation and maintenance of mite reduction strategies and helped reduce allergen levels in homes of dust mite-sensitive children.
We enrolled 60 households of children aged 5-15 with parent-reported dust mite allergy into a randomized controlled trial. Intervention homes (N=30) received educational material about reducing dust mites and test kits at 1,2,5, and 8 months. Control homes (N=30) received only educational material. At baseline, 6 and 12 months, study staff visited all homes, collected dust samples from 3 locations and obtained information about parents’ mite reduction behaviors by questionnaire. Allergen concentrations (Der f 2/Der p2) in dust were assessed by immunoassays. After adjusting for visit and location, allergen concentrations in intervention and control homes were compared using mixed effects model analysis.
In the intervention homes, allergen concentrations in the child's bedroom and living room floors were significantly reduced over time compared to control homes. Although not all location-specific differences in allergen concentrations were statistically significant, combining data across locations, there was a differential reduction in allergen concentrations in the intervention group versus the control group (p =0.02).
The use of in-home test kits along with education may beneficially influence behaviors and attitudes towards dust mite reduction strategies and help reduce residential dust mite allergen levels.
PMCID: PMC4769118  PMID: 26308287
dust mites; dust mite allergen; D. pteronyssinus; D. farinae; indoor allergens; intervention trials
2.  Soluble Epoxide Hydrolase Null Mice Exhibit Female and Male Differences In Regulation of Vascular Homeostasis 
Increased CYP epoxygenase activity and consequently up regulation of epoxyeicosatrienoic acids (EETs) levels provides protection against metabolic syndrome and cardiovascular diseases. Conversion of arachidonic acid epoxides to diols by soluble epoxide hydrolase (sEH) diminishes the beneficial cardiovascular properties of these epoxyeicosanoids. We therefore examined the possible biochemical consequences of sEH deletion on vascular responses in male and female mice. Through the use of the sEH KO mouse, we provide evidence of differences in the compensatory response in the balance between nitric oxide (NO), carbon monoxide (CO), EETs and the vasoconstrictor 20-HETE in male and female KO mice. Serum levels of adiponectin, TNFα, IL-1b and MCP1 and protein expression in vascular tissue of p-AMPK, p-AKT and p-eNOS were measured. Deletion of sEH caused a significant (p<0,05) decrease in body weight, and an increase in adiponectin, pAMPK and pAKT levels in female KO mice compared to male KO mice. Gene deletion resulted in a higher production of renal EETs in female KO compared to male KO mice and, concomitantly, we observed an increase in renal 20-HETEs levels and superoxide anion production only in male KO mice. sEH deletion increased p-AKT and p-eNOS protein expression but decreased p-AMPK levels in female KO mice. Increased levels of p-eNOS at Thr-495 were observed only in KO male mice. While p-eNOS at 1177 were not significantly different between male and female. Nitric oxide production was unaltered in male KO mice. These results provide evidence of gender differences in the preservation of vascular homeostasis in response to sEH deletion which involves regulation of phosphorylation of eNOS at the 495 site.
PMCID: PMC4575626  PMID: 25908301
pAKT; pAMPK; adiponectin; EETs
3.  Prevalence of allergic sensitization in the U.S.: Results from the National Health and Nutrition Examination Survey (NHANES) 2005–2006 
Allergic sensitization is an important risk factor for the development of atopic disease. The National Health and Nutrition Examination Survey (NHANES) 2005–2006 provides the most comprehensive information on IgE-mediated sensitization in the general US population.
We investigated clustering, sociodemographic and regional patterns of allergic sensitization and examined risk factors associated with IgE-mediated sensitization.
Data for this cross-sectional analysis were obtained from NHANES 2005–2006. Participants aged ≥1 year (N=9440) were tested for sIgEs to inhalant and food allergens; participants ≥6 years were tested for 19 sIgEs, and children aged 1–5 years for 9 sIgEs. Serum samples were analyzed using the ImmunoCAP System. Information on demographics and participant characteristics was collected by questionnaire.
Of the study population aged 6 and older, 44.6% had detectable sIgEs, while 36.2% of children aged 1–5 years were sensitized to ≥1 allergen. Allergen-specific IgEs clustered into 7 groups that might have largely reflected biological cross-reactivity. Although sensitization to individual allergens and allergen types showed regional variation, the overall prevalence of sensitization did not differ across census regions, except in early childhood. In multivariate modeling, young age, male gender, non-Hispanic black race/ethnicity, geographic location (census region), and reported pet avoidance measures were most consistently associated with IgE-mediated sensitization.
The overall prevalence of allergic sensitization does not vary across US census regions, except in early life, although allergen-specific sensitization differs by sociodemographic and regional factors. Biological cross-reactivity may be an important, but not a sole, contributor to the clustering of allergen-specific IgEs.
Clinical implications
IgE-mediated sensitization shows clustering patterns and differs by sociodemographic and regional factors, but the overall prevalence of sensitization may not vary across US census regions.
PMCID: PMC4119838  PMID: 24522093
allergen; allergy; allergic sensitization; serum IgE
4.  Regulation of T Helper Cell Subsets by Cyclooxygenases and Their Metabolites 
Cyclooxygenases and their metabolites are important regulators of inflammatory responses and play critical roles in regulating the differentiation of T helper cell subsets in inflammatory diseases. In this review, we highlight new information on regulation of T helper cell subsets by cyclooxygenases and their metabolites. Prostanoids influence cytokine production on both antigen presenting cells and T cells to regulate the differentiation of naïve CD4+ T cells to Th1, Th2 and Th17 cell phenotypes. Cyclooxygenases and PGE2 generally exacerbate Th2 and Th17 phenotypes, while suppressing Th1 differentiation. Thus, cycloxygenases may play a critical role in diseases that involve immune cell dysfunction. Targeting of cyclooxygenases and their eicosanoid products may represent a new approach for treatment of inflammatory diseases, tumors and autoimmune disorders.
PMCID: PMC3620713  PMID: 23201570
Cyclooxygenases; T helper cells; Prostanoids; PGE2
5.  Cyclooxygenase-2 Inhibits T Helper Cell Type 9 Differentiation during Allergic Lung Inflammation via Down-regulation of IL-17RB 
Rationale: Helper CD4+ T cell subsets, including IL-9– and IL-10–producing T helper cell type 9 (Th9) cells, exist under certain inflammatory conditions. Cyclooxygenase (COX)-1 and COX-2 play important roles in allergic lung inflammation and asthma. It is unknown whether COX-derived eicosanoids regulate Th9 cells during allergic lung inflammation.
Objectives: To determine the role of COX metabolites in regulating Th9 cell differentiation and function during allergic lung inflammation.
Methods: COX-1−/−, COX-2−/−, and wild-type (WT) mice were studied in an in vivo model of ovalbumin-induced allergic inflammation and an in vitro model of Th9 differentiation using flow cytometry, cytokine assays, confocal microscopy, real-time PCR, and immunoblotting. In addition, the role of specific eicosanoids and their receptors was examined using synthetic prostaglandins (PGs), selective inhibitors, and siRNA knockdown.
Measurements and Main Results: Experimental endpoints were not different between COX-1−/− and WT mice; however, the percentage of IL-9+ CD4+ T cells was increased in lung, bronchoalveolar lavage fluid, lymph nodes, and blood of allergic COX-2−/− mice relative to WT. Bronchoalveolar lavage fluid IL-9 and IL-10, serum IL-9, and lung IL-17RB levels were significantly increased in allergic COX-2−/− mice or in WT mice treated with COX-2 inhibitors. IL-9, IL-10, and IL-17RB expression in vivo was inhibited by PGD2 and PGE2, which also reduced Th9 cell differentiation of murine and human naive CD4+ T cells in vitro. Inhibition of protein kinase A significantly increased Th9 cell differentiation of naive CD4+ T cells isolated from WT mice in vitro.
Conclusions: COX-2–derived PGD2 and PGE2 regulate Th9 cell differentiation by suppressing IL-17RB expression via a protein kinase A–dependent mechanism.
PMCID: PMC3707371  PMID: 23449692
T helper cell type 9 cells; cyclooxygenase 2; asthma; prostaglandins; IL-17RB
6.  Cyclooxygenase-2 Regulates Th17 Cell Differentiation during Allergic Lung Inflammation 
Rationale: Th17 cells comprise a distinct lineage of proinflammatory T helper cells that are major contributors to allergic responses. It is unknown whether cyclooxygenase (COX)-derived eicosanoids regulate Th17 cells during allergic lung inflammation.
Objectives: To determine the role of COX metabolites in regulating Th17 cell differentiation and function during allergic lung inflammation.
Methods: COX-1−/−, COX-2−/−, and wild-type mice were studied in an in vivo model of ovalbumin-induced allergic inflammation and an in vitro model of Th17 differentiation using flow cytometry, cytokine assays, confocal microscopy, real-time polymerase chain reaction, and immunoblotting. In addition, the role of specific eicosanoids and their receptors was examined using synthetic prostaglandins (PGs), selective inhibitors, and siRNA knockdown.
Measurements and Main Results: Th17 cell differentiation in lung, lymph nodes, and bronchoalveolar lavage fluid was significantly lower in COX-2−/− mice after ovalbumin sensitization and exposure in vivo. In vitro studies revealed significantly impaired Th17 cell differentiation of COX-2−/− naive CD4+ T cells with decreased Stat3 phosphorylation and RORγt expression. Synthetic PGF2α and PGI2 enhanced Th17 cell differentiation of COX-2−/− CD4+ T cells in vitro. The selective COX-2 inhibitor, NS-398, and PGF2α receptor and PGI2 receptor siRNA knockdown significantly decreased Th17 cell differentiation in vitro. Administration of synthetic PGs restored accumulation of Th17 cells in lungs of allergic COX-2−/− mice in vivo.
Conclusions: COX-2 is a critical regulator of Th17 cell differentiation during allergic lung inflammation via autocrine signaling of PGI2 and PGF2α through their respective cell surface receptors.
PMCID: PMC3172888  PMID: 21474648
Th17 cell; COX-2; asthma; prostaglandins; IL-17
7.  Ventilation Defects Observed with Hyperpolarized 3He Magnetic Resonance Imaging in a Mouse Model of Acute Lung Injury 
Regions of diminished ventilation are often evident during functional pulmonary imaging studies, including hyperpolarized gas magnetic resonance imaging (MRI), positron emission tomography, and computed tomography (CT). The objective of this study was to characterize the hypointense regions observed via 3He MRI in a murine model of acute lung injury. LPS at doses ranging from 15–50 μg was intratracheally administered to C57BL/6 mice under anesthesia. Four hours after exposure to either LPS or saline vehicle, mice were imaged via hyperpolarized 3He MRI. All images were evaluated to identify regions of hypointense signals. Lungs were then characterized by conventional histology, or used to obtain tissue samples from regions of normal and hypointense 3He signals and analyzed for cytokine content. The characterization of 3He MRI images identified three distinct types of hypointense patterns: persistent defects, atelectatic defects, and dorsal lucencies. Persistent defects were associated with the administration of LPS. The number of persistent defects depended on the dose of LPS, with a significant increase in mean number of defects in 30–50-μg LPS-dosed mice versus saline-treated control mice. Atelectatic defects predominated in LPS-dosed mice under conditions of low-volume ventilation, and could be reversed with deep inspiration. Dorsal lucencies were present in nearly all mice studied, regardless of the experimental conditions, including control animals that did not receive LPS. A comparison of 3He MRI with histopathology did not identify tissue abnormalities in regions of low 3He signal, with the exception of a single region of atelectasis in one mouse. Furthermore, no statistically significant differences were evident in concentrations of IL-1β, IL-6, macrophage inflammatory protein (MIP)-1α, MIP-2, chemokine (C-X-C motif) ligand 1 (KC), TNFα, and monocyte chemotactic protein (MCP)-1 between hypointense and normally ventilated lung regions in LPS-dosed mice. Thus, this study defines the anatomic, functional, and biochemical characteristics of ventilation defects associated with the administration of LPS in a murine model of acute lung injury.
PMCID: PMC3095984  PMID: 20595465
ventilation defect; lipopolysaccharide; lung inflammation; magnetic resonance imaging; cytokines
8.  Allergy-related outcomes in relation to serum IgE: Results from the National Health and Nutrition Examination Survey 2005–2006 
The National Health and Nutrition Examination Survey (NHANES) 2005–2006 was the first population-based study to investigate levels of serum total and allergen-specific immunoglobulin E (IgE) in the general US population.
We estimated prevalence of allergy-related outcomes and examined relationships between serum IgE levels and these outcomes in a representative sample of the US population.
Data for this cross-sectional analysis were obtained from the NHANES 2005–2006. Study subjects aged 6 years and older (N=8086) had blood taken for measurement of total IgE and 19 specific IgEs against common aeroallergens, including Alternaria alternata, Aspergillus fumigatus, Bermuda grass, birch, oak, ragweed, Russian thistle, rye grass, cat dander, cockroach, dog dander, dust mite (Dermatophagoides farinae and D. pteronyssinus), mouse and rat urine proteins; and selected foods (egg white, cow’s milk, peanut, and shrimp). Serum samples were analyzed for total and allergen-specific IgEs using the Pharmacia CAP System. Information on allergy-related outcomes and demographics was collected by questionnaire.
In the NHANES 2005–2006, 6.6% reported current hay fever and 23.5% suffered from current allergies. Allergy-related outcomes increased with increasing total IgE (adjusted ORs for a 10-fold increase in total IgE =1.86, 95% CI:1.44–2.41 for hay fever and 1.64, 95% CI: 1.41–1.91 for allergies). Elevated levels of plant-, pet-, and mold-specific IgEs contributed independently to allergy-related symptoms. The greatest increase in odds was observed for hay fever and plant-specific IgEs (adjusted OR=4.75, 95% CI:3.83–5.88).
In the US population, self-reported allergy symptoms are most consistently associated with elevated levels of plant-, pet-, and mold-specific IgEs.
PMCID: PMC3108140  PMID: 21320720
allergen; allergy; allergic sensitization; serum IgE
9.  Epoxyeicosanoids stimulate multiorgan metastasis and tumor dormancy escape in mice 
Epoxyeicosatrienoic acids (EETs) are small molecules produced by cytochrome P450 epoxygenases. They are lipid mediators that act as autocrine or paracrine factors to regulate inflammation and vascular tone. As a result, drugs that raise EET levels are in clinical trials for the treatment of hypertension and many other diseases. However, despite their pleiotropic effects on cells, little is known about the role of these epoxyeicosanoids in cancer. Here, using genetic and pharmacological manipulation of endogenous EET levels, we demonstrate that EETs are critical for primary tumor growth and metastasis in a variety of mouse models of cancer. Remarkably, we found that EETs stimulated extensive multiorgan metastasis and escape from tumor dormancy in several tumor models. This systemic metastasis was not caused by excessive primary tumor growth but depended on endothelium-derived EETs at the site of metastasis. Administration of synthetic EETs recapitulated these results, while EET antagonists suppressed tumor growth and metastasis, demonstrating in vivo that pharmacological modulation of EETs can affect cancer growth. Furthermore, inhibitors of soluble epoxide hydrolase (sEH), the enzyme that metabolizes EETs, elevated endogenous EET levels and promoted primary tumor growth and metastasis. Thus, our data indicate a central role for EETs in tumorigenesis, offering a mechanistic link between lipid signaling and cancer and emphasizing the critical importance of considering possible effects of EET-modulating drugs on cancer.
PMCID: PMC3248288  PMID: 22182838
10.  Gene Delivery of Cytochrome P450 Epoxygenase Ameliorates Monocrotaline-Induced Pulmonary Artery Hypertension in Rats 
Pulmonary arterial hypertension (PAH) is a life-threatening disease that leads to progressive pulmonary hypertension, right heart failure, and death. Endothelial dysfunction and inflammation were implicated in the pathogenesis of PAH. Epoxyeicosatrienoic acids (EETs), products of the cytochrome P450 epoxygenase metabolism of arachidonic acid, are potent vasodilators that possess anti-inflammatory and other protective properties in endothelial cells. We investigated whether gene delivery with the human cytochrome P450 epoxygenase 2J2 (CYP2J2) ameliorates monocrotaline (MCT)-induced pulmonary hypertension in rats. Significant pulmonary hypertension developed 3 weeks after the administration of MCT, but gene therapy with CYP2J2 significantly attenuated the development of pulmonary hypertension and pulmonary vascular remodeling, without causing changes in systemic arterial pressure or heart rate. These effects were associated with increased pulmonary endothelial NO synthase (eNOS) expression and its activity, inhibition of inflammation in the lungs, and transforming growth factor (TGF)-β/type II bone morphogenetic protein receptor (BMPRII)-drosophila mothers against decapentaplegic proteins (Smads) signaling. Collectively, these data suggest that gene therapy with CYP2J2 may have potential as a novel therapeutic approach to this progressive and oftentimes lethal disorder.
PMCID: PMC2993093  PMID: 20118222
arachidonic acids; cytochrome P450 epoxygenase; gene therapy; monocrotaline
11.  Hormonal Influences on Lung Function and Response to Environmental Agents 
Numerous studies in humans and experimental animals have identified considerable sex differences in respiratory physiology and in the response of the lung to environmental agents. These differences appear to be mediated, at least in part, by sex hormones and their nuclear receptors. Moreover, animal models are increasingly used to study pathogenic mechanisms and test potential therapies for a variety of human lung diseases, many of which appear to be influenced by sex and sex hormones. In this article, data are summarized from studies of lung function and disease in which sex differences have been observed. Specific attention is paid to animal models of acute lung injury, nonallergic and allergic lung inflammation, and lung fibrosis. It is anticipated that continued investigation of the role of sex and sex hormones in animal models will provide valuable insight into the pathogenesis and potential treatments for a variety of acute and chronic human lung diseases.
PMCID: PMC3266051  PMID: 19934354
sex; sex hormones; respiratory mechanics; inflammation; airway
13.  Indoor Allergens in School and Daycare Environments 
Most studies that have examined exposure to indoor allergens have focused on home environments. However, allergen exposures can be encountered in environments other than the home. For example, many children spend a large part of their time in schools and daycare facilities. Over the past two decades, a large number of studies have been conducted in school and daycare environments. However, the role of indoor exposures in allergy and asthma development or morbidity in these settings is not well characterized. The purpose of this review is to evaluate the importance of indoor allergen exposures in school and daycare settings. We summarize the key findings from recent scientific literature, describe exposure characteristics, discuss the role of these exposures in relation to asthma and allergy symptoms, and provide information on the effectiveness of published interventions.
PMCID: PMC2749571  PMID: 19577284
allergen; indoor; exposure; asthma; allergy; school; daycare
14.  Nanoparticle-Mediated Drug Delivery and Pulmonary Hypertension 
Hypertension  2009;53(5):751-753.
PMCID: PMC2694215  PMID: 19307468
15.  Cockroach Allergen Reduction by Cockroach Control Alone in Low-Income, Urban Homes-A Randomized Control Trial 
We previously reported significant reductions in cockroach allergen concentrations in urban homes by reducing cockroach infestations.
Determine the effectiveness of pest control performed by professional entomologists, compared to commercial companies, in reducing cockroach allergen.
This 3-arm randomized controlled trial enrolled 60 cockroach-infested homes in North Carolina. Homes were randomly assigned to a control group or one of two treatment groups. Treatment 1 had insecticide baits placed by entomologists from North Carolina State University. Treatment 2 received pest control from a randomly assigned commercial company. Vacuumed dust sampling and cockroach trapping were conducted at 0, 6 and 12 months. Dust samples were analyzed by ELISA.
In Treatment 1 homes, there were significant reductions in geometric mean trap counts compared to Control and Treatment 2 homes at 12 months. Relative to control, significant 12-month reductions in Bla g 1 were evident in Treatment 1 homes at all sampled sites, except bedroom floor. From baseline to month 12, geometric mean Bla g 1 concentrations (U/g) decreased from 64.2 to 5.6 in kitchen, 10.6 to 1.1 in living room, 10.7 to 1.9 in bedroom floor and 3.6 to 2.3 in bed. Treatment 2 homes showed no significant 12-month allergen reductions versus control.
Reductions in Bla g 1 in cockroach-infested homes can be achieved by reducing infestations; however, the magnitude of allergen reduction is dependent on the thoroughness and effectiveness of cockroach eradication efforts.
PMCID: PMC2804464  PMID: 17825893
cockroaches; cockroach allergen; Bla g 1; Bla g 2; indoor allergens; intervention trial
16.  Male Sex Hormones Exacerbate Lung Function Impairment after Bleomycin-Induced Pulmonary Fibrosis 
The roles of sex hormones as modulators of lung function and disease have received significant attention as differential sex responses to various lung insults have been recently reported. The present study used a bleomycin-induced pulmonary fibrosis model in C57BL/6 mice to examine potential sex differences in physiological and pathological outcomes. Endpoints measured included invasive lung function assessment, immunological response, lung collagen deposition, and a quantitative histological analysis of pulmonary fibrosis. Male mice had significantly higher basal static lung compliance than female mice (P < 0.05) and a more pronounced decline in static compliance after bleomycin administration when expressed as overall change or percentage of baseline change (P < 0.05). In contrast, there were no significant differences between the sexes in immune cell infiltration into the lung or in total lung collagen content after bleomycin. Total lung histopathology scores measured using the Ashcroft method did not differ between the sexes, while a quantitative histopathology scoring system designed to determine where within the lung the fibrosis occurred indicated a tendency toward more fibrosis immediately adjacent to airways in bleomycin-treated male versus female mice. Furthermore, castrated male mice exhibited a female-like response to bleomycin while female mice given exogenous androgen exhibited a male-like response. These data indicate that androgens play an exacerbating role in decreased lung function after bleomycin administration, and traditional measures of fibrosis may miss critical differences in lung function between the sexes. Sex differences should be carefully considered when designing and interpreting experimental models of pulmonary fibrosis in mice.
PMCID: PMC2438447  PMID: 18276795
fibrosis; bleomycin; sex; respiratory mechanics
17.  Cardiomyocyte-specific expression of CYP2J2 prevents development of cardiac remodelling induced by angiotensin II 
Cardiovascular Research  2015;105(3):304-317.
Cardiac remodelling is one of the key pathological changes that occur with cardiovascular disease. Previous studies have demonstrated the beneficial effects of CYP2J2 expression on cardiac injury. In the present study, we investigated the effects of cardiomyocyte-specific CYP2J2 expression and EET treatment on angiotensin II-induced cardiac remodelling and sought to determine the underlying molecular mechanisms involved in this process.
Methods and results
Eight-week-old mice with cardiomyocyte-specific CYP2J2 expression (αMHC-CYP2J2-Tr) and wild-type (WT) control mice were treated with Ang-II. Ang-II treatment of WT mice induced changes in heart morphology, cardiac hypertrophy and dysfunction, as well as collagen accumulation; however, cardiomyocyte-specific expression of CYP2J2 attenuated these effects. The cardioprotective effects observed in α-MHC-CYP2J2-Tr mice were associated with peroxisome proliferator-activated receptor (PPAR)-γ activation, reduced oxidative stress, reduced NF-κB p65 nuclear translocation, and inhibition of TGF-β1/smad pathway. The effects seen with cardiomyocyte-specific expression of CYP2J2 were partially blocked by treatment with PPAR-γ antagonist GW9662. In in vitro studies, 11,12-EET(1 μmol/L) treatment attenuated cardiomyocyte hypertrophy and remodelling-related protein (collagen I, TGF-β1, TIMP1) expression by inhibiting the oxidative stress-mediated NF-κB pathway via PPAR-γ activation. Furthermore, conditioned media from neonatal cardiomyocytes treated with 11,12-EET inhibited activation of cardiac fibroblasts and TGF-β1/smad pathway.
Cardiomyocyte-specific expression of CYP2J2 or treatment with EETs protects against cardiac remodelling by attenuating oxidative stress-mediated NF-κBp65 nuclear translocation via PPAR-γ activation.
PMCID: PMC4351370  PMID: 25618409
Epoxyeicosatrienoic acids; Cardiac hypertrophy; CYP2J2; Cardiac remodelling; Oxidative stress
18.  Cyclooxygenase-2 Deficiency Exacerbates Bleomycin-Induced Lung Dysfunction but Not Fibrosis 
Cyclooxygenase (COX)-derived eicosanoids have been implicated in the pathogenesis of pulmonary fibrosis. Uncertainty regarding the influence of COX-2 on experimental pulmonary fibrosis prompted us to clarify the fibrotic and functional effects of intratracheal bleomycin administration in mice genetically deficient in COX-2. Further, the effects of airway-specific COX-1 overexpression on fibrotic and functional outcomes in wild-type and COX-2 knockout mice were assessed. Equivalent increases in airway cell influx, lung collagen content, and histopathologic evidence of fibrosis were observed in wild-type and COX-2 knockout mice 21 d after bleomycin treatment, suggesting that COX-2 deficiency did not alter the extent or severity of fibrosis in this model. However, bleomycin-induced alterations in respiratory mechanics were more severe in COX-2 knockout mice than in wild-type mice, as illustrated by a greater decrease in static compliance compared with genotype-matched, saline-treated control mice (26 ± 3% versus 11 ± 4% decreases for COX-2 knockout and wild-type mice, respectively; P < 0.05). The influence of COX-1 overexpression in airway Clara cells was also examined. Whereas the fibrotic effects of bleomycin were not altered in wild-type or COX-2 knockout mice overexpressing COX-1, the exaggerated lung function decrement in bleomycin-treated COX-2 knockout mice was prevented by COX-1 overexpression and coincided with decreased airway cysteinyl leukotriene levels. Collectively, these data suggest an important regulatory role for COX-2 in the maintenance of lung function in the setting of lung fibrosis, but not in the progression of the fibrotic process per se.
PMCID: PMC1994226  PMID: 17496151
cyclooxygenase; fibrosis; respiratory mechanics; prostaglandin; transgenic
19.  Exposure to multiple indoor allergens in US homes and relationship to asthma 
The National Survey of Lead and Allergens in Housing was the first population-based study to measure indoor allergen levels in US homes.
We characterized the overall burden to multiple allergens and examined whether elevated allergen levels were associated with occupants’ asthma status.
This cross-sectional study surveyed a nationally representative sample of 831 housing units in 75 different locations throughout the US. Information was collected by questionnaire and environmental assessments. Allergen concentrations in dust samples were assessed by immunoassays. The following cut points were used to define elevated allergen levels: 10 μg/g for Der p 1, Der f 1, and Can f 1; 8 μg/g for Fel d 1; 8 U/g Bla g 1; 1.6 μg/g for mouse urinary protein; and 7 μg/g for Alternaria antigens. Allergen burden was considered high when 4 or more allergens exceeded elevated levels in any of the sampling locations.
Exposure to multiple allergens was common in US homes. Of the surveyed homes, 51.5% had at least 6 detectable allergens and 45.8% had at least 3 allergens exceeding elevated levels. Occupants’ race, income, housing type, absence of children, and presence of smokers, pets, cockroaches, rodents and mold/moisture related problems were independent predictors of high allergen burden. Among atopics, high allergen burden increased the odds of having asthma symptoms (OR=1.81, 95% CI: 1.04-3.15).
Elevated allergen levels in the home are associated with asthma symptoms in allergic individuals.
Clinical implication
In allergic asthma, indoor allergen exposures play an important role in asthma exacerbations.
PMCID: PMC2376121  PMID: 18255132
allergen; indoor; exposure; asthma; allergy
20.  G-protein Pathway Suppressor 2 (GPS2) Interacts with the Regulatory Factor X4 Variant 3 (RFX4_v3) and Functions as a Transcriptional Co-activator* 
The Journal of biological chemistry  2008;283(13):8580-8590.
The regulatory factor X4 variant 3 (RFX4_v3) is a brain-specific isoform of the transcription factor RFX4. Insertional mutagenesis in mice demonstrates that Rfx4_v3 is crucial for normal brain development. Many genes involved in critical processes during brain morphogenesis are dysregulated in Rfx4_v3 mutant brains. For example, Cx3cl1 is a CX3C-type chemokine that is abundant in brain and is a direct transcriptional target of RFX4_v3 through a specific promoter X-box (X-box 1), the responsive element for RFX proteins. To identify potential interacting partners for RFX4_v3, we performed yeast two-hybrid analysis. Nine candidate interactors were identified, including G-protein pathway suppressor 2 (GPS2). Indirect immunofluorescence demonstrated that GPS2 and RFX4_v3 co-localized to the nucleus. Both GPS2 and RFX4_v3 mRNAs were also present in most portions of the adult mouse brain, as well as in brains at different ages, suggesting that the two proteins could bind to each other. Co-immunoprecipitation assays indicated that physical interactions between GPS2 and RFX4_v3 did indeed occur. Furthermore, GPS2 was recruited to the Cx3cl1 promoter by RFX4_v3 and potentiated RFX4_v3 transactivation on this promoter through X-box 1, suggesting that the protein-protein interaction was functionally relevant. GPS2 bound to both the carboxyl-terminal region (amino acids 575-735) and the middle region (amino acids 250-574) of the RFX4_v3 protein. RFX4_v3 amino acids 1-574 stimulated the Cx3cl1 promoter to the similar extent as the full-length RFX4_v3 protein; however, deletion of the carboxyl-terminal region of RFX4_v3 impaired the co-activating abilities of GPS2. Based on these data, we conclude that GPS2 interacts with RFX4_v3 to modulate transactivation of genes involved in brain morphogenesis, including Cx3Cl1.
PMCID: PMC2365754  PMID: 18218630
21.  Epoxyeicosatrienoic Acids Enhance Embryonic Haematopoiesis and Adult Marrow Engraftment 
Nature  2015;523(7561):468-471.
Haematopoietic stem and progenitor cell (HSPC) transplant is a widely used treatment for life-threatening conditions including leukemia; however, the molecular mechanisms regulating HSPC engraftment of the recipient niche remain incompletely understood. Here, we developed a competitive HSPC transplant method in adult zebrafish, using in vivo imaging as a non-invasive readout. We used this system to conduct a chemical screen and identified epoxyeicosatrienoic acids (EET) as a family of lipids1,2 that enhance HSPC engraftment. EETs’ pro-haematopoietic effects were conserved in the developing zebrafish embryo, where 11,12-EET promoted HSPC specification by activating a unique AP-1/runx1 transcription program autonomous to the haemogenic endothelium. This effect required the activation of the PI3K pathway, specifically PI3Kγ. In adult HSPCs, 11,12-EET induced transcriptional programs, including AP-1 activation, which modulate multiple cellular processes, such as migration, to promote engraftment. Finally, we demonstrated that the EET effects on enhancing HSPC homing and engraftment are conserved in mammals. Our study established a novel method to explore the molecular mechanisms of HSPC engraftment, and discovered a previously unrecognized, evolutionarily conserved pathway regulating multiple haematopoietic generation and regeneration processes. EETs may have clinical application in marrow or cord blood transplantation.
PMCID: PMC4754787  PMID: 26201599
22.  Asthma Cases Attributable to Atopy: Results from the Third National Health and Nutrition Examination Survey 
The percentage of asthma cases attributable to atopy is the subject of debate.
The objectives were to estimate the percentage of asthma cases in the U.S. population attributable to atopy and to examine associations between allergen-specific skin tests and asthma.
Data were obtained from NHANES III in which subjects aged 6–59 years were skin tested with 10 allergens. Atopy was defined as at least one positive allergen-specific test. Doctor-diagnosed current asthma was assessed by questionnaire.
In the U.S., 56.3% of the asthma cases were attributable to atopy, and that percentage was greater among males than females, among persons in the highest education category than in lower education categories, and among persons living in highly populated metropolitan areas than in all other areas. Each allergen-specific test was strongly associated with asthma before adjustment (odds ratios varied from 2.1 to 4.5); however, after adjustment by all the allergens, only tests to cat, Alternaria, white oak, and perennial rye were independently associated with asthma. Perennial rye was inversely associated with asthma. Of the 10 allergens, a positive response to cat accounted for the highest percentage of asthma cases (29.3%).
About half of the current asthma cases in the U.S. population represented by NHANES III were attributable to atopy. Some allergen-specific skin tests were not independently associated with asthma.
Clinical Implications
If atopy could be prevented or reversed, or its effect on asthma blocked, then a large percentage of asthma cases in the U.S. population could be prevented.
PMCID: PMC2291202  PMID: 17889931
allergens; allergic sensitization; allergy skin test; asthma; atopy; epidemiology; NHANES III; skin prick test; survey
23.  Gender Differences in Murine Airway Responsiveness and Lipopolysaccharide-Induced Inflammation1 
The roles of gender and sex hormones in lung function and disease are complex and not completely understood. The present study examined the influence of gender on lung function and respiratory mechanics in naive mice and on acute airway inflammation and hyperresponsiveness induced by intratracheal LPS administration. Basal lung function characteristics did not differ between naive males and females, but males demonstrated significantly greater airway responsiveness than females following aerosolized methacholine challenge as evidenced by increased respiratory system resistance and elastance (p < 0.05). Following LPS administration, males developed more severe hypothermia and greater airway hyperresponsiveness than females (p < 0.05). Inflammatory indices including bronchoalveolar lavage fluid total cells, neutrophils, and TNF-α content were greater in males than in females 6 h following LPS administration (p < 0.05), whereas whole-lung TLR-4 protein levels did not differ among treatment groups, suggesting that differential expression of TLR-4 before or after LPS exposure did not underlie the observed inflammatory outcomes. Gonadectomy decreased airway inflammation in males but did not alter inflammation in females, whereas administration of exogenous testosterone to intact females increased their inflammatory responses to levels observed in intact males. LPS-induced airway hyperresponsiveness was also decreased in castrated males and was increased in females administered exogenous testosterone. Collectively, these data indicate that airway responsiveness in naive mice is influenced by gender, and that male mice have exaggerated airway inflammatory and functional responses to LPS compared with females. These gender differences are mediated, at least in part, by effects of androgens.
PMCID: PMC2262913  PMID: 16785560
24.  Electrophysiological Properties of Cardiomyocytes Isolated from CYP2J2 Transgenic Mice 
Molecular pharmacology  2007;72(4):1063-1073.
CYP2J2 is abundant in cardiac tissue and active in the biosynthesis of eicosanoids such as epoxyeicosatrienoic acids (EETs). To determine the effects of CYP2J2 and its eicosanoid products in the heart, we characterized the electrophysiology of single cardiomyocytes isolated from adult transgenic (Tr) mice with cardiac-specific overexpression of CYP2J2. CYP2J2 Tr cardiomyocytes had a shortened action potential. At 90% repolarization, the action potential duration (APD) was 30.6 ± 3.0 ms (n = 22) in wild-type (Wt) cells and 20.2 ± 2.3 ms (n = 19) in CYP2J2 Tr cells (p < 0.005). This shortening was probably due to enhanced maximal peak transient outward K+ currents (Ito,peak), which were 38.6 ± 2.8 and 54.4 ± 4.9 pA/pF in Wt and CYP2J2 Tr cells, respectively (p < 0.05). In contrast, the late portion of the transient outward K+ current (Ito,280ms), the slowly inactivating outward K+ current (IK,slow), and the voltage-gated Na+ current (INa) were not significantly altered in CYP2J2 Tr cells. N-Methylsulphonyl-6-(2-proparglyloxy-phenyl)hexanamide (MS-PPOH), a specific inhibitor of EET biosynthesis, significantly reduced Ito,peak and increased APD in CYP2J2 Tr cardiomyocytes but not in Wt cells. Intracellular dialysis with a monoclonal antibody against CYP2J2 also significantly reduced Ito,peak and increased APD in CYP2J2 Tr cardiomyocytes. Addition of 11,12-EET or 8-bromo-cAMP significantly reversed the MS-PPOH- or monoclonal antibody-induced changes in Ito,peak and APD in CYP2J2 Tr cells. Together, our data demonstrate that shortening of the action potential in CYP2J2 Tr cardiomyocytes is associated with enhanced Ito,peak via an EET-dependent, cAMP-mediated mechanism.
PMCID: PMC2243182  PMID: 17652182
25.  Cyclooxygenase-1 Overexpression Decreases Basal Airway Responsiveness but Not Allergic Inflammation1 
Pharmacological inhibition or genetic disruption of cyclooxygenase (COX)-1 or COX-2 exacerbates the inflammatory and functional responses of the lung to environmentally relevant stimuli. To further examine the contribution of COX-derived eicosanoids to basal lung function and to allergic lung inflammation, transgenic (Tr) mice were generated in which overexpression of human COX-1 was targeted to airway epithelium. Although no differences in basal respiratory or lung mechanical parameters were observed, COX-1 Tr mice had increased bronchoalveolar lavage fluid PGE2 content compared with wild-type littermates (23.0 ± 3.6 vs 8.4 ± 1.4 pg/ml; p < 0.05) and exhibited decreased airway responsiveness to inhaled methacholine. In an OVA-induced allergic airway inflammation model, comparable up-regulation of COX-2 protein was observed in the lungs of allergic wild-type and COX-1 Tr mice. Furthermore, no genotype differences were observed in allergic mice in total cell number, eosinophil content (70 vs 76% of total cells, respectively), and inflammatory cytokine content of bronchoalveolar lavage fluid, or in airway responsiveness to inhaled methacholine (p > 0.05). To eliminate the presumed confounding effects of COX-2 up-regulation, COX-1 Tr mice were bred into a COX-2 null background. In these mice, the presence of the COX-1 transgene did not alter allergen-induced inflammation but significantly attenuated allergen-induced airway hyperresponsiveness, coincident with reduced airway leuko-triene levels. Collectively, these data indicate that COX-1 overexpression attenuates airway responsiveness under basal conditions but does not influence allergic airway inflammation.
PMCID: PMC2222891  PMID: 16982919

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