a-chronic-cardiac-ischemia-model-in-swine-using-an-ameroid-constrictor

A Chronic Cardiac Ischemia Model in Swine Using an Ameroid Constrictor

Cardiovascular disease remains the number one cause of mortality in the United States. There are numerous approaches to treating these diseases, but ...

National Institutes of Health

Distinct sets of corticosteroid-unresponsive genes modulate disease severity in asthma.

Apolipoprotein E negatively regulates house dust mite-induced asthma via a low-density lipoprotein receptor-mediated pathway.

New treatment approaches are needed for patients with asthma. Apolipoprotein A-I (apoA-I), the major structural protein of high-density lipoproteins, mediates reverse cholesterol transport and has atheroprotective and anti-inflammatory effects. In this study, we hypothesized that an apoA-I mimetic peptide might be effective at inhibiting asthmatic airway inflammation. A 5A peptide, which is a synthetic, bihelical apoA-I mimetic, was administered to wild-type A/J mice via osmotic mini-pump prior to the induction of house dust mite (HDM)-induced asthma. HDM-challenged mice that received the 5A apoA-I mimetic peptide had significant reductions in the number of bronchoalveolar lavage fluid eosinophils, lymphocytes, and neutrophils, as well as in histopathological evidence of airway inflammation. The reduction in airway inflammation was mediated by a reduction in the expression of Th2- and Th17-type cytokines, as well as in chemokines that promote T cell and eosinophil chemotaxis, including CCL7, CCL17, CCL11, and CCL24. Furthermore, the 5A apoA-I mimetic peptide inhibited the alternative activation of pulmonary macrophages in the lungs of HDM-challenged mice. It also abrogated the development of airway hyperresponsiveness and reduced several key features of airway remodeling, including goblet cell hyperplasia and the expression of collagen genes (Col1a1 and Col3a1). Our results demonstrate that the 5A apoA-I mimetic peptide attenuates the development of airway inflammation and airway hyperresponsiveness in an experimental murine model of HDM-induced asthma. These data support the conclusion that strategies using apoA-I mimetic peptides, such as 5A, might be developed further as a possible new treatment approach for asthma.

5A, an apolipoprotein A-I mimetic peptide, attenuates the induction of house dust mite-induced asthma.

Apolipoprotein E (apoE) is an endogenous negative regulator of airway hyperreactivity (AHR) and mucous cell metaplasia in experimental models of house dust mite (HDM)-induced airway disease. The gene encoding human apoE is polymorphic, with three common alleles (ε2, ε3, and ε4) reflecting single amino acid substitutions at amino acids 112 and 158. The objective of this study was to assess whether the human apoE alleles modify airway responses to repeated nasal HDM challenges. Mice expressing the human apoE ε2 (huApoE2), ε3 (huApoE3), or ε4 (huApoE4) alleles received nasal HDM challenges, and airway responses were compared with mice expressing the endogenous murine apoE gene (muApoE). huApoE3 mice displayed significant reductions in AHR, mucous cell metaplasia, and airway inflammation compared with muApoE mice. The attenuated severity of airway inflammation in huApoE3 mice was associated with reductions in lung mRNA levels of Th2 and Th17 cytokines, as well as chemokines (CCL7, CCL11, CCL24). huApoE4 mice had an intermediate phenotype, with attenuated AHR and IgE production, compared with muApoE mice, whereas airway inflammation and mucous cell metaplasia were not reduced. In contrast, HDM-induced airway responses were not modified in mice expressing the huApoE2 allele. We conclude that the polymorphic huApoE alleles differentially modulate HDM-induced airway disease, which can be stratified, in rank order of increasing disease severity, ε3 < ε4 < ε2. These results raise the possibility that the polymorphic apoE alleles may modify disease severity in human asthma.

Human apolipoprotein E genotypes differentially modify house dust mite-induced airway disease in mice.

Apolipoprotein A-I (apoA-I) is a key component of high-density lipoproteins that mediates reverse cholesterol transport from cells and reduces vascular inflammation. We investigated whether endogenous apoA-I modulates ovalbumin (OVA)-induced airway inflammation in mice. We found that apoA-I expression was significantly reduced in the lungs of OVA-challenged, compared with saline-challenged, wild-type (WT) mice. Next, to investigate the role of endogenous apoA-I in the pathogenesis of OVA-induced airway inflammation, WT and apoA-I(-/-) mice were sensitized by intraperitoneal injections of OVA and aluminum hydroxide, followed by multiple nasal OVA challenges for 4 weeks. OVA-challenged apoA-I(-/-) mice exhibited a phenotype of increased airway neutrophils compared with WT mice, which could be rescued by an administration of a 5A apoA-I mimetic peptide. Multiple pathways promoted neutrophilic inflammation in OVA-challenged apoA-I(-/-) mice, including the up-regulated expression of (1) proinflammatory cytokines (IL-17A and TNF-α), (2) CXC chemokines (CXCL5), (3) vascular adhesion molecules (i.e., vascular cell adhesion molecule-1), and (4) granulocyte colony-stimulating factors (G-CSF). Because concentrations of G-CSF in bronchoalveolar lavage fluid (BALF) were markedly increased in OVA-challenged apoA-I(-/-) mice, we hypothesized that enhanced G-CSF expression may represent the predominant pathway mediating increased neutrophilic inflammation. This was confirmed by the intranasal administration of a neutralizing anti-G-CSF antibody, which significantly reduced BALF neutrophilia by 72% in OVA-challenged apoA-I(-/-) mice, compared with mice that received a control antibody. We conclude that endogenous apoA-I negatively regulates OVA-induced neutrophilic airway inflammation, primarily via a G-CSF-dependent mechanism. Furthermore, these findings suggest that apoA-I may play an important role in modulating the severity of neutrophilic airway inflammation in asthma.

Apolipoprotein A-I attenuates ovalbumin-induced neutrophilic airway inflammation via a granulocyte colony-stimulating factor-dependent mechanism.

The mammalian target of rapamycin (mTOR) modulates immune responses and cellular proliferation. The objective of this study was to assess whether inhibition of mTOR with rapamycin modifies disease severity in two experimental murine models of house dust mite (HDM)-induced asthma. In an induction model, rapamycin was administered to BALB/c mice coincident with nasal HDM challenges for 3 weeks. In a treatment model, nasal HDM challenges were performed for 6 weeks and rapamycin treatment was administered during weeks 4 through 6. In the induction model, rapamycin significantly attenuated airway inflammation, airway hyperreactivity (AHR) and goblet cell hyperplasia. In contrast, treatment of established HDM-induced asthma with rapamycin exacerbated AHR and airway inflammation, whereas goblet cell hyperplasia was not modified. Phosphorylation of the S6 ribosomal protein, which is downstream of mTORC1, was increased after 3 weeks, but not 6 weeks of HDM-challenge. Rapamycin reduced S6 phosphorylation in HDM-challenged mice in both the induction and treatment models. Thus, the paradoxical effects of rapamycin on asthma severity paralleled the activation of mTOR signaling. Lastly, mediastinal lymph node re-stimulation experiments showed that treatment of rapamycin-naive T cells with ex vivo rapamycin decreased antigen-specific Th2 cytokine production, whereas prior exposure to in vivo rapamycin rendered T cells refractory to the suppressive effects of ex vivo rapamycin. We conclude that rapamycin had paradoxical effects on the pathogenesis of experimental HDM-induced asthma. Thus, consistent with the context-dependent effects of rapamycin on inflammation, the timing of mTOR inhibition may be an important determinant of efficacy and toxicity in HDM-induced asthma.

Paradoxical effects of rapamycin on experimental house dust mite-induced asthma.

Peptidoglycan recognition protein (Pglyrp) 1 is a pattern-recognition protein that mediates antibacterial host defense. Because we had previously shown that Pglyrp1 expression is increased in the lungs of house dust mite (HDM)-challenged mice, we hypothesized that it might modulate the pathogenesis of asthma. Wild-type and Pglyrp1(-/-) mice on a BALB/c background received intranasal HDM or saline, 5 days/week for 3 weeks. HDM-challenged Pglyrp1(-/-) mice showed decreases in bronchoalveolar lavage fluid eosinophils and lymphocytes, serum IgE, and mucous cell metaplasia, whereas airway hyperresponsiveness was not changed when compared with wild-type mice. T helper type 2 (Th2) cytokines were reduced in the lungs of HDM-challenged Pglyrp1(-/-) mice, which reflected a decreased number of CD4(+) Th2 cells. There was also a reduction in C-C chemokines in bronchoalveolar lavage fluid and lung homogenates from HDM-challenged Pglyrp1(-/-) mice. Furthermore, secretion of CCL17, CCL22, and CCL24 by alveolar macrophages from HDM-challenged Pglyrp1(-/-) mice was markedly reduced. As both inflammatory cells and airway epithelial cells express Pglyrp1, bone marrow transplantation was performed to generate chimeric mice and assess which cell type promotes HDM-induced airway inflammation. Chimeric mice lacking Pglyrp1 on hematopoietic cells, not structural cells, showed a reduction in HDM-induced eosinophilic and lymphocytic airway inflammation. We conclude that Pglyrp1 expressed by hematopoietic cells, such as alveolar macrophages, mediates HDM-induced airway inflammation by up-regulating the production of C-C chemokines that recruit eosinophils and Th2 cells to the lung. This identifies a new family of innate immune response proteins that promotes HDM-induced airway inflammation in asthma.

Peptidoglycan recognition protein 1 promotes house dust mite-induced airway inflammation in mice.

The very low density lipoprotein receptor (VLDLR) is a member of the low-density lipoprotein receptor family that binds multiple ligands and plays a key role in brain development. Although the VLDLR mediates pleiotropic biological processes, only a limited amount of information is available regarding its role in adaptive immunity. In this study, we identify an important role for the VLDLR in attenuating house dust mite (HDM)-induced airway inflammation in experimental murine asthma. We show that HDM-challenged Vldlr(-/-) mice have augmented eosinophilic and lymphocytic airway inflammation with increases in Th2 cytokines, C-C chemokines, IgE production, and mucous cell metaplasia. A genome-wide analysis of the lung transcriptome identified that mRNA levels of CD209e (DC-SIGNR4), a murine homolog of DC-SIGN, were increased in the lungs of HDM-challenged Vldlr(-/-) mice, which suggested that the VLDLR might modify dendritic cell (DC) function. Consistent with this, VLDLR expression by human monocyte-derived DCs was increased by HDM stimulation. In addition, 55% of peripheral blood CD11c(+) DCs from individuals with allergy expressed VLDLR under basal conditions. Lastly, the adoptive transfer of HDM-pulsed, CD11c(+) bone marrow-derived DCs (BMDCs) from Vldlr(-/-) mice to the airways of wild type recipient mice induced augmented eosinophilic and lymphocytic airway inflammation upon HDM challenge with increases in Th2 cytokines, C-C chemokines, IgE production, and mucous cell metaplasia, as compared with the adoptive transfer of HDM-pulsed, CD11c(+) BMDCs from wild type mice. Collectively, these results identify a novel role for the VLDLR as a negative regulator of DC-mediated adaptive immune responses in HDM-induced allergic airway inflammation.

The very low density lipoprotein receptor attenuates house dust mite-induced airway inflammation by suppressing dendritic cell-mediated adaptive immune responses.

Apolipoprotein A-I (apoA-I) is an important component of high-density lipoprotein particles that mediates reverse cholesterol transport out of cells by interacting with the ATP-binding cassette transporter 1 (ABCA1). apoA-I has also been shown to attenuate neutrophilic airway inflammation in experimental ovalbumin (OVA)-induced asthma by reducing the expression of granulocyte colony-stimulating factor (G-CSF). Here, we hypothesized that overexpression of the ABCA1 transporter might similarly attenuate OVA-induced neutrophilic airway inflammation. Tie2-human ABCA1 (hABCA1) mice expressing human ABCA1 under the control of the Tie2 promoter, which is primarily expressed by vascular endothelial cells, but can also be expressed by macrophages, received daily intranasal OVA challenges, 5 d/wk for 5 weeks. OVA-challenged Tie2-hABCA1 mice had significant reductions in total bronchoalveolar lavage fluid (BALF) cells that reflected a decrease in neutrophils, as well as reductions in peribronchial inflammation, OVA-specific IgE levels, and airway epithelial thickness. The reduced airway neutrophilia in OVA-challenged Tie2-hABCA1 mice was associated with significant decreases in G-CSF protein levels in pulmonary vascular endothelial cells, alveolar macrophages, and BALF. Intranasal administration of recombinant murine G-CSF to OVA-challenged Tie2-hABCA1 mice for 5 days increased BALF neutrophils to a level comparable to that of OVA-challenged wild-type mice. We conclude that ABCA1 suppresses OVA-induced airway neutrophilia by reducing G-CSF production by vascular endothelial cells and alveolar macrophages. These findings suggest that ABCA1 expressed by vascular endothelial cells and alveolar macrophages may play important roles in attenuating the severity of neutrophilic airway inflammation in asthma.

ATP-binding cassette transporter 1 attenuates ovalbumin-induced neutrophilic airway inflammation.

DNA-dependent protein kinase (DNA-PK) mediates double-stranded DNA break repair, V(D)J recombination and immunoglobulin class switch recombination, as well as innate immune and pro-inflammatory responses. However, there is limited information regarding the role of DNA-PK in adaptive immunity mediated by dendritic cells (DCs), which are the primary antigen-presenting cells in allergic asthma. Here we show that house dust mite induces DNA-PK phosphorylation, which is a marker of DNA-PK activation, in DCs via the generation of intracellular reactive oxygen species. We also demonstrate that pharmacological inhibition of DNA-PK, as well as the specific deletion of DNA-PK in DCs, attenuates the induction of allergic sensitization and Th2 immunity via a mechanism that involves the impaired presentation of mite antigens. Furthermore, pharmacological inhibition of DNA-PK following antigen priming similarly reduces the manifestations of mite-induced airway disease. Collectively, these findings suggest that DNA-PK may be a potential target for treatment of allergic asthma.

Karen J. Keeran

Animal Surgery and Resources Core,
National Heart,
Lung,
and Blood Institute,
Animal Surgery and Resources Core, National Heart, Lung, and Blood Institute

National Institutes of Health

A Chronic Cardiac Ischemia Model in Swine Using an Ameroid Constrictor

Karen J. Keeran

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National Institutes of Health

A Chronic Cardiac Ischemia Model in Swine Using an Ameroid Constrictor

Animal Surgery and Resources Core,
National Heart,
Lung,
and Blood Institute,
Animal Surgery and Resources Core, National Heart, Lung, and Blood Institute