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1.
J Allergy Clin Immunol ; 142(6): 1808-1817.e3, 2018 12.
Artículo en Inglés | MEDLINE | ID: mdl-29522849

RESUMEN

BACKGROUND: Altered epithelial physical and functional barrier properties along with TH1/TH2 immune dysregulation are features of allergic asthma. Regulation of junction proteins to improve barrier function of airway epithelial cells has the potential for alleviation of allergic airway inflammation. OBJECTIVE: We sought to determine the immunomodulatory effect of knob protein of the adenoviral capsid on allergic asthma and to investigate its mechanism of action on airway epithelial junction proteins and barrier function. METHODS: Airway inflammation, including junction protein expression, was evaluated in allergen-challenged mice with and without treatment with knob. Human bronchial epithelial cells were exposed to knob, and its effects on expression of junction proteins and barrier integrity were determined. RESULTS: Administration of knob to allergen-challenged mice suppressed airway inflammation (eosinophilia, airway hyperresponsiveness, and IL-5 levels) and prevented allergen-induced loss of airway epithelial occludin and E-cadherin expression. Additionally, knob decreased expression of TH2-promoting inflammatory mediators, specifically IL-33, by murine lung epithelial cells. At a cellular level, treatment of human bronchial epithelial cells with knob activated c-Jun N-terminal kinase, increased expression of occludin and E-cadherin, and enhanced epithelial barrier integrity. CONCLUSION: Increased expression of junction proteins mediated by knob leading to enhanced epithelial barrier function might mitigate the allergen-induced airway inflammatory response, including asthma.


Asunto(s)
Proteínas de la Cápside/farmacología , Proteínas de la Cápside/uso terapéutico , Células Epiteliales/efectos de los fármacos , Adenoviridae , Anciano , Animales , Bronquios/citología , Líquido del Lavado Bronquioalveolar/inmunología , Cadherinas/metabolismo , Línea Celular , Citocinas/inmunología , Eosinofilia/inmunología , Células Epiteliales/metabolismo , Femenino , Humanos , Masculino , Ratones Endogámicos BALB C , Persona de Mediana Edad , Ocludina/metabolismo , Proteínas Recombinantes/farmacología , Proteínas Recombinantes/uso terapéutico , Hipersensibilidad Respiratoria/tratamiento farmacológico , Hipersensibilidad Respiratoria/inmunología
2.
Blood ; 120(8): 1742-51, 2012 Aug 23.
Artículo en Inglés | MEDLINE | ID: mdl-22791291

RESUMEN

Neutrophil recruitment and extravasation at sites of inflammation provide a mechanism for host defense. We showed previously that heparan sulfate, a type of sulfated glycosaminoglycan, facilitates neutrophil recruitment based on the reduction of neutrophil infiltration in mice in which the overall sulfation of the chains was reduced by selective inactivation of N-acetylglucosamine N-deacetylase-N-sulfotransferase (Ndst1) in endothelial cells. Here we show that inactivation of uronyl 2-O-sulfotransferase in endothelial cells (Hs2st), an enzyme that acts downstream from Ndst1, results in enhanced neutrophil recruitment in several models of acute inflammation. Enhanced neutrophil infiltration resulted in part from reduced rolling velocity under flow both in vivo and in vitro, which correlated with stronger binding of neutrophil L-selectin to mutant endothelial cells. Hs2st-deficient endothelial cells also displayed a striking increase in binding of IL-8 and macrophage inflammatory protein-2. The enhanced binding of these mediators of neutrophil recruitment resulted from a change in heparan sulfate structure caused by increased N-sulfation and 6-O-sulfation of glucosamine units in response to the decrease in 2-O-sulfation of uronic acid residues. This gain-of-function phenotype provides formidable evidence demonstrating the importance of endothelial heparan sulfate in inflammation and suggests a novel enzyme target for enhancing the innate immune response.


Asunto(s)
Silenciador del Gen , Infiltración Neutrófila , Neutrófilos/inmunología , Peritonitis/inmunología , Sulfotransferasas/genética , Sulfotransferasas/inmunología , Animales , Células Cultivadas , Quimiocinas/inmunología , Células Endoteliales/inmunología , Células Endoteliales/metabolismo , Heparitina Sulfato/química , Heparitina Sulfato/inmunología , Inflamación/inducido químicamente , Inflamación/genética , Inflamación/inmunología , Selectina L/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Neutrófilos/citología , Peritonitis/inducido químicamente , Peritonitis/genética , Tioglicolatos
3.
J Immunol ; 188(3): 1479-90, 2012 Feb 01.
Artículo en Inglés | MEDLINE | ID: mdl-22210919

RESUMEN

Eosinophils are the predominant inflammatory cells recruited to allergic airways. In this article, we show that human and murine eosinophils express SWAP-70, an intracellular RAC-binding signaling protein, and examine its role in mediating eosinophil trafficking and pulmonary recruitment in a murine model of allergic airway inflammation. Compared with wild-type eosinophils, SWAP-70-deficient (Swap-70(-/-)) eosinophils revealed altered adhesive interactions within inflamed postcapillary venules under conditions of blood flow by intravital microscopy, exhibiting enhanced slow rolling but decreased firm adhesion. In static adhesion assays, Swap-70(-/-) eosinophils adhered poorly to VCAM-1 and ICAM-1 and exhibited inefficient leading edge and uropod formation. Adherent Swap-70(-/-) eosinophils failed to translocate RAC1 to leading edges and displayed aberrant cell surface localization/distribution of α4 and Mac-1. Chemokine-induced migration of Swap-70(-/-) eosinophils was significantly decreased, correlating with reduced intracellular calcium levels, defective actin polymerization/depolymerization, and altered cytoskeletal rearrangement. In vivo, recruitment of eosinophils to the lungs of allergen-challenged Swap-70(-/-) mice, compared with wild-type mice, was significantly reduced, along with considerable attenuation of airway inflammation, indicated by diminished IL-5, IL-13, and TNF-α levels; reduced mucus secretion; and improved airway function. These findings suggest that regulation of eosinophil trafficking and migration by SWAP-70 is important for the development of eosinophilic inflammation after allergen exposure.


Asunto(s)
Movimiento Celular , Proteínas de Unión al ADN , Eosinófilos/patología , Factores de Intercambio de Guanina Nucleótido , Proteínas Nucleares , Hipersensibilidad Respiratoria/etiología , Alérgenos , Animales , Adhesión Celular , Humanos , Inflamación , Ratones , Antígenos de Histocompatibilidad Menor , Hipersensibilidad Respiratoria/patología , Vénulas/patología
4.
Sci Transl Med ; 16(739): eadd8936, 2024 Mar 20.
Artículo en Inglés | MEDLINE | ID: mdl-38507467

RESUMEN

Glucocorticoids (GCs) are efficacious drugs used for treating many inflammatory diseases, but the dose and duration of administration are limited because of severe side effects. We therefore sought to identify an approach to selectively target GCs to inflamed tissue. Previous work identified that anti-tumor necrosis factor (TNF) antibodies that bind to transmembrane TNF undergo internalization; therefore, an anti-TNF antibody-drug conjugate (ADC) would be mechanistically similar, where lysosomal catabolism could release a GC receptor modulator (GRM) payload to dampen immune cell activity. Consequently, we have generated an anti-TNF-GRM ADC with the aim of inhibiting pro-inflammatory cytokine production from stimulated human immune cells. In an acute mouse model of contact hypersensitivity, a murine surrogate anti-TNF-GRM ADC inhibited inflammatory responses with minimal effect on systemic GC biomarkers. In addition, in a mouse model of collagen-induced arthritis, single-dose administration of the ADC, delivered at disease onset, was able to completely inhibit arthritis for greater than 30 days, whereas an anti-TNF monoclonal antibody only partially inhibited disease. ADC treatment at the peak of disease was also able to attenuate the arthritic phenotype. Clinical data for a human anti-TNF-GRM ADC (ABBV-3373) from a single ascending dose phase 1 study in healthy volunteers demonstrated antibody-like pharmacokinetic profiles and a lack of impact on serum cortisol concentrations at predicted therapeutic doses. These data suggest that an anti-TNF-GRM ADC may provide improved efficacy beyond anti-TNF alone in immune mediated diseases while minimizing systemic side effects associated with standard GC treatment.


Asunto(s)
Anticuerpos , Artritis Experimental , Inmunoconjugados , Esteroides , Humanos , Animales , Ratones , Preparaciones Farmacéuticas , Receptores de Glucocorticoides/uso terapéutico , Inhibidores del Factor de Necrosis Tumoral/uso terapéutico , Glucocorticoides/farmacología , Glucocorticoides/uso terapéutico , Factor de Necrosis Tumoral alfa/metabolismo , Modelos Animales de Enfermedad , Inmunoconjugados/farmacología , Inmunoconjugados/uso terapéutico
5.
Am J Respir Cell Mol Biol ; 47(4): 427-35, 2012 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-22556157

RESUMEN

The ADP-ribosyl cyclase activity of CD38 generates cyclic ADP-ribose, a Ca(2+)-mobilizing agent. In human airway smooth muscle (HASM) cells, TNF-α mediates CD38 expression through mitogen-activated protein kinases and NF-κB and AP-1. The phosphatidylinositol-3 kinase/Akt (PI3K/Akt) pathway is involved in TNF-α signaling and contributes to airway hyperresponsiveness and airway remodeling. We hypothesized that PI3Ks mediate CD38 expression and are involved in the differential induction of CD38 by TNF-α in asthmatic HASM cells. HASM cells were treated with pan-PI3K inhibitors (LY294002 or wortmannin) or class I-selective (GDC0941) or isoform-selective PI3K inhibitors (p110α-PIK-75 and p110ß-TGX-221) with or without TNF-α. HASM cells were transfected with a catalytically active form of PI3K or phosphatase and tensin homolog (PTEN) or nontargeting or p110 isoform-targeting siRNAs before TNF-α exposure. CD38 expression and activation of Akt, NF-κB, and AP-1 were determined. LY294002 and wortmannin inhibited TNF-α-induced Akt activation, whereas only LY294002 inhibited CD38 expression. P110 expression caused Akt activation and basal and TNF-α-induced CD38 expression, whereas PTEN expression attenuated Akt activation and CD38 expression. Expression levels of p110 isoforms α, ß, and δ were comparable in nonasthmatic and asthmatic HASM cells. Silencing of p110α or -δ, but not p110ß, resulted in comparable attenuation of TNF-α-induced CD38 expression in asthmatic and nonasthmatic cells. NF-κB and AP-1 activation were unaltered by the PI3K inhibitors. In HASM cells, regulation of CD38 expression occurs by specific class I PI3K isoforms, independent of NF-κB or AP-1 activation, and PI3K signaling may not be involved in the differential elevation of CD38 in asthmatic HASM cells.


Asunto(s)
ADP-Ribosil Ciclasa 1/metabolismo , Fosfatidilinositol 3-Quinasa Clase Ia/metabolismo , Regulación de la Expresión Génica , Glicoproteínas de Membrana/metabolismo , Miocitos del Músculo Liso/enzimología , Sistema Respiratorio/patología , ADP-Ribosil Ciclasa 1/genética , Asma/enzimología , Asma/metabolismo , Asma/patología , Células Cultivadas , Cromonas/farmacología , Fosfatidilinositol 3-Quinasa Clase Ia/genética , Activación Enzimática , Técnicas de Silenciamiento del Gen , Humanos , Isoenzimas/antagonistas & inhibidores , Isoenzimas/genética , Isoenzimas/metabolismo , Glicoproteínas de Membrana/genética , Morfolinas/farmacología , Miocitos del Músculo Liso/metabolismo , FN-kappa B/metabolismo , Fosfohidrolasa PTEN/metabolismo , Inhibidores de las Quinasa Fosfoinosítidos-3 , Cultivo Primario de Células , Proteínas Proto-Oncogénicas c-akt , Pirimidinonas/farmacología , Interferencia de ARN , Transducción de Señal , Factor de Transcripción AP-1/metabolismo , Factor de Necrosis Tumoral alfa/fisiología
6.
J Biol Chem ; 286(44): 38231-38241, 2011 Nov 04.
Artículo en Inglés | MEDLINE | ID: mdl-21911487

RESUMEN

Allergic airway inflammation, including asthma, is usually characterized by the predominant recruitment of eosinophils. However, neutrophilia is also prominent during severe exacerbations. Cell surface-expressed glycans play a role in leukocyte trafficking and recruitment during inflammation. Here, the involvement of UDP-N-acetylglucosamine:α-6-D-mannoside ß1,6-N-acetylglucosaminyltransferase V (MGAT5)-modified N-glycans in eosinophil and neutrophil recruitment during allergic airway inflammation was investigated. Allergen-challenged Mgat5-deficient (Mgat5(-/-)) mice exhibited significantly attenuated airway eosinophilia and inflammation (decreased Th2 cytokines, mucus production) compared with WT counterparts, attributable to decreased rolling, adhesion, and survival of Mgat5(-/-) eosinophils. Interestingly, allergen-challenged Mgat5(-/-) mice developed airway neutrophilia and increased airway reactivity with persistent elevated levels of proinflammatory cytokines (IL-17A, TNFα, IFNγ)). This increased neutrophil recruitment was also observed in LPS- and thioglycollate (TG)-induced inflammation in Mgat5(-/-) mice. Furthermore, there was significantly increased recruitment of infused Mgat5(-/-) neutrophils compared with WT neutrophils in the peritoneal cavity of TG-exposed WT mice. Mgat5(-/-) neutrophils demonstrated enhanced adhesion to P-selectin as well as increased migration toward keratinocyte-derived chemokine compared with WT neutrophils in vitro along with increased calcium mobilization upon activation and expression of elevated levels of CXCR2, which may contribute to the increased neutrophil recruitment. These data indicate an important role for MGAT5-modified N-glycans in differential regulation of eosinophil and neutrophil recruitment during allergic airway inflammation.


Asunto(s)
Eosinófilos/metabolismo , Inflamación , Infiltración Neutrófila , Neutrófilos/metabolismo , Polisacáridos/química , Animales , Líquido del Lavado Bronquioalveolar , Carbohidratos/química , Movimiento Celular , Quimiotaxis , Hipersensibilidad/metabolismo , Pulmón/patología , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , N-Acetilglucosaminiltransferasas/metabolismo , Células Th2/citología
7.
Am J Physiol Lung Cell Mol Physiol ; 302(11): L1179-91, 2012 Jun 01.
Artículo en Inglés | MEDLINE | ID: mdl-22427531

RESUMEN

Trafficking and recruitment of eosinophils during allergic airway inflammation is mediated by the phosphatidylinositol 3-kinase (PI3K) family of signaling molecules. The role played by the p110δ subunit of PI3K (PI3K p110δ) in regulating eosinophil trafficking and recruitment was investigated using a selective pharmacological inhibitor (IC87114). Treatment with the PI3K p110δ inhibitor significantly reduced murine bone marrow-derived eosinophil (BM-Eos) adhesion to VCAM-1 as well as ICAM-1 and inhibited activation-induced changes in cell morphology associated with reduced Mac-1 expression and aberrant cell surface localization/distribution of Mac-1 and α4. Infused BM-Eos demonstrated significantly decreased rolling and adhesion in inflamed cremaster muscle microvessels of mice treated with IC87114 compared with vehicle-treated mice. Furthermore, inhibition of PI3K p110δ significantly attenuated eotaxin-1-induced BM-Eos migration and prevented eotaxin-1-induced changes in the cytoskeleton and cell morphology. Knockdown of PI3K p110δ with siRNA in BM-Eos resulted in reduced rolling, adhesion, and migration, as well as inhibition of activation-induced changes in cell morphology, validating its role in regulating trafficking and migration. Finally, in a mouse model of cockroach antigen-induced allergic airway inflammation, oral administration of the PI3K p110δ inhibitor significantly inhibited airway eosinophil recruitment, resulting in attenuation of airway hyperresponsiveness in response to methacholine, reduced mucus secretion, and expression of proinflammatory molecules (found in inflammatory zone-1 and intelectin-1). Overall, these findings indicate the important role played by PI3K p110δ in mediating BM-Eos trafficking and migration by regulating adhesion molecule expression and localization/distribution as well as promoting changes in cell morphology that favor recruitment during inflammation.


Asunto(s)
Asma/inmunología , Eosinofilia/inmunología , Eosinófilos/inmunología , Fosfatidilinositol 3-Quinasas/metabolismo , Sistema Respiratorio/inmunología , Adenina/análogos & derivados , Adenina/farmacología , Animales , Asma/metabolismo , Células de la Médula Ósea , Adhesión Celular/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Quimiocina CCL11/metabolismo , Fosfatidilinositol 3-Quinasa Clase I , Eosinofilia/metabolismo , Eosinófilos/metabolismo , Hipersensibilidad/inmunología , Hipersensibilidad/metabolismo , Molécula 1 de Adhesión Intercelular/inmunología , Molécula 1 de Adhesión Intercelular/metabolismo , Antígeno de Macrófago-1/biosíntesis , Cloruro de Metacolina/farmacología , Ratones , Ratones Endogámicos BALB C , Fosfatidilinositol 3-Quinasas/genética , Inhibidores de las Quinasa Fosfoinosítidos-3 , Quinazolinas/farmacología , Interferencia de ARN , ARN Interferente Pequeño , Sistema Respiratorio/metabolismo , Molécula 1 de Adhesión Celular Vascular/inmunología , Molécula 1 de Adhesión Celular Vascular/metabolismo
8.
J Immunol ; 185(2): 1205-14, 2010 Jul 15.
Artículo en Inglés | MEDLINE | ID: mdl-20543100

RESUMEN

The role played by the beta-galactoside-binding lectin galectin-3 (Gal-3) in airway remodeling, a characteristic feature of asthma that leads to airway dysfunction and poor clinical outcome in humans, was investigated in a murine model of chronic allergic airway inflammation. Wild-type (WT) and Gal-3 knockout (KO) mice were subjected to repetitive allergen challenge with OVA up to 12 wk, and bronchoalveolar lavage fluid (BALF) and lung tissue collected after the last challenge were evaluated for cellular features associated with airway remodeling. Compared to WT mice, chronic OVA challenge in Gal-3 KO mice resulted in diminished remodeling of the airways with significantly reduced mucus secretion, subepithelial fibrosis, smooth muscle thickness, and peribronchial angiogenesis. The higher degree of airway remodeling in WT mice was associated with higher Gal-3 expression in the BALF as well as lung tissue. Cell counts in BALF and lung immunohistology demonstrated that eosinophil infiltration in OVA-challenged Gal-3 KO mice was significantly reduced compared with that WT mice. Evaluation of cellular mediators associated with eosinophil recruitment and airway remodeling revealed that levels of eotaxin-1, IL-5, IL-13, found in inflammatory zone 1, and TGF-beta were substantially lower in Gal-3 KO mice. Finally, leukocytes from Gal-3 KO mice demonstrated decreased trafficking (rolling) on vascular endothelial adhesion molecules compared with that of WT cells. Overall, these studies demonstrate that Gal-3 is an important lectin that promotes airway remodeling via airway recruitment of inflammatory cells, specifically eosinophils, and the development of a Th2 phenotype as well as increased expression of eosinophil-specific chemokines and profibrogenic and angiogenic mediators.


Asunto(s)
Remodelación de las Vías Aéreas (Respiratorias)/inmunología , Alérgenos/inmunología , Galectina 3/inmunología , Inflamación/inmunología , Remodelación de las Vías Aéreas (Respiratorias)/genética , Animales , Western Blotting , Líquido del Lavado Bronquioalveolar/química , Líquido del Lavado Bronquioalveolar/inmunología , Quimiocina CCL11/genética , Quimiocina CCL11/metabolismo , Eosinófilos/inmunología , Eosinófilos/metabolismo , Eosinófilos/patología , Femenino , Citometría de Flujo , Galectina 3/deficiencia , Galectina 3/genética , Interleucina-13/metabolismo , Interleucina-5/metabolismo , Rodamiento de Leucocito/inmunología , Pulmón/inmunología , Pulmón/metabolismo , Pulmón/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ovalbúmina/inmunología , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Molécula 1 de Adhesión Celular Vascular/genética , Molécula 1 de Adhesión Celular Vascular/inmunología , Molécula 1 de Adhesión Celular Vascular/metabolismo
9.
J Nanosci Nanotechnol ; 12(7): 5178-84, 2012 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-22966542

RESUMEN

Polyethyleneimine (PEI) has been described as a highly efficient gene carrier due to its efficient proton sponge effect within endosomes. However, many studies have demonstrated that PEI is toxic and associated with a lack of cell specificity despite high transfection efficiency. In order to minimize the toxicity of PEI, we prepared chitosan-graft-spermine (CHI-g-SPE) in a previous study. CHI-g-SPE showed low toxicity and high transfection efficiency. However, this compound also had limited target cell specificity. In the present study, we synthesized galactosylated CHI-g-SPE (GCS) because this modified GCS could be delivered specifically into the liver due to hepatocyte-specific galactose receptors. The DNA-binding properties of GCS at various copolymer/DNA weight ratios were evaluated by a gel retardation assay. The GCS copolymer exhibited significant DNA-binding ability and efficiently protected DNA from nuclease attack. Using energy-filtered transmission electron microscopy (EF-TEM), we observed dense spherical, nano-sized GCS/DNA complexes with a homogenous distribution. Most importantly, GCS was associated with remarkably low cytotoxicity compared to PEI in HepG2, HeLa, and A549 cells. Moreover, GCS carriers specifically delivered the gene-of-interest into hepatocytes in vitro as well as in vivo. Our results suggest that the novel GCS described here is a safe and highly efficient carrier for hepatocyte-targeted gene delivery.


Asunto(s)
Quitosano/química , ADN/farmacocinética , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Nanocápsulas/química , Espermina/química , Transfección/métodos , Animales , Línea Celular Tumoral , ADN/administración & dosificación , Portadores de Fármacos/síntesis química , Humanos , Masculino , Ratones , Ratones Transgénicos
10.
J Immunol ; 183(6): 3971-9, 2009 Sep 15.
Artículo en Inglés | MEDLINE | ID: mdl-19710461

RESUMEN

The effect of targeted inactivation of the gene encoding N-deacetylase/N-sulfotransferase-1 (Ndst1), a key enzyme involved in the biosynthesis of heparan sulfate (HS) chains, on the inflammatory response associated with allergic inflammation in a murine model of OVA-induced acute airway inflammation was investigated. OVA-exposed Ndst1(f/f)TekCre(+) (mutant) mice deficient in endothelial and leukocyte Ndst1 demonstrated significantly decreased allergen-induced airway hyperresponsiveness and inflammation characterized by a significant reduction in airway recruitment of inflammatory cells (eosinophils, macrophages, neutrophils, and lymphocytes), diminished IL-5, IL-2, TGF-beta1, and eotaxin levels, as well as decreased expression of TGF-beta1 and the angiogenic protein FIZZ1 (found in inflammatory zone 1) in lung tissue compared with OVA-exposed Ndst1(f/f)TekCre(-) wild-type littermates. Furthermore, murine eosinophils demonstrated significantly decreased rolling on lung endothelial cells (ECs) from mutant mice compared with wild-type ECs under conditions of flow in vitro. Treatment of wild-type ECs, but not eosinophils, with anti-HS Abs significantly inhibited eosinophil rolling, mimicking that observed with Ndst1-deficient ECs. In vivo, trafficking of circulating leukocytes in lung microvessels of allergen-challenged Ndst1-deficient mice was significantly lower than that observed in corresponding WT littermates. Endothelial-expressed HS plays an important role in allergic airway inflammation through the regulation of recruitment of inflammatory cells to the airways by mediating interaction of leukocytes with the vascular endothelium. Furthermore, HS may also participate by sequestering and modulating the activity of allergic asthma-relevant mediators such as IL-5, IL-2, and TGF-beta1.


Asunto(s)
Células Endoteliales/enzimología , Heparitina Sulfato/deficiencia , Leucocitos/enzimología , Hipersensibilidad Respiratoria/patología , Sulfotransferasas/deficiencia , Animales , Quimiotaxis , Inflamación/etiología , Inflamación/patología , Interleucina-2/análisis , Interleucina-5/análisis , Ratones , Hipersensibilidad Respiratoria/etiología , Factor de Crecimiento Transformador beta1/análisis
11.
Exp Lung Res ; 37(5): 279-90, 2011 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-21309736

RESUMEN

Allergic inflammation is associated with increased generation and trafficking of inflammatory cells, especially eosinophils, to sites of inflammation. The effect of acute versus chronic airway allergen challenge on hematopoietic activity in the bone marrow (BM) and lungs was investigated using murine models of allergic airway inflammation. Acute allergen challenge induced proliferation of BM cells and significantly increased generation of eosinophil, but not multipotent, granulocyte-macrophage (GM), or B-lymphocyte progenitor cells. However, no hematopoietic activity was observed in the lungs. With chronic challenge, BM cells failed to proliferate, but exhibited increased capacity to generate multipotent as well as eosinophil, GM, and B-lymphocyte progenitors. In addition, increased generation of eosinophil- and GM-specific progenitors was observed in the lungs. Although no differences were observed in their ability to roll on BM endothelium in vitro or in vivo, CD34-enriched hematopoietic/stem progenitor cells (HSPCs) from chronic-, but not acute-, challenged mice demonstrated reduced migration across BM endothelial cells associated with decreased CXCR4 expression. Overall, these studies demonstrate that chronic allergen exposure can alter BM homing due to decreased transendothelial migration enabling noninteracting HSPCs to egress out of the BM and recruit to sites of inflammation such as the airways, resulting in extramedullary hematopoiesis.


Asunto(s)
Alérgenos/inmunología , Hematopoyesis Extramedular/inmunología , Inflamación/inmunología , Pulmón/inmunología , Alérgenos/administración & dosificación , Animales , Antígenos CD34/inmunología , Línea Celular , Movimiento Celular/inmunología , Eosinofilia/inmunología , Eosinofilia/metabolismo , Eosinófilos/inmunología , Eosinófilos/metabolismo , Inflamación/metabolismo , Pulmón/metabolismo , Ratones , Células Madre Multipotentes/inmunología , Células Madre Multipotentes/metabolismo , Células Progenitoras Mieloides/inmunología , Células Progenitoras Mieloides/metabolismo , Células Precursoras de Linfocitos B/inmunología , Células Precursoras de Linfocitos B/metabolismo , Receptores CXCR4/metabolismo
12.
Artículo en Inglés | MEDLINE | ID: mdl-30651985

RESUMEN

The gut microbiome has long been known to play fundamentally important roles in the animal health and the well-being of its host. As such, the establishment and maintenance of a beneficial gut microbiota early in life is crucial in pigs, since early gut colonizers are pivotal in the establishment of permanent microbial community structures affecting the health and growth performance of pigs later in life. Emphasizing this importance of early gut colonizers, it is critical to understand the factors impacting the establishment of the piglet gut microbiome at weaning. Factors include, among others, diet, in-feed antibiotics, probiotics and prebiotic administration. The impact of these factors on establishment of the gut microbiome of piglets at weaning includes effects on piglet gut microbial diversity, structure, and succession. In this review, we thoroughly reviewed the most recent findings on the piglet gut microbiome shifts as influenced by weaning, and how these microbiome changes brought about by various factors that have been shown to affect the development of microbiota in piglets. This review will provide a general overview of recent studies that can help to facilitate the design of new strategies to modulate the gut microbiome in order to enhance gastrointestinal health, growth performance and well-being of piglets.

13.
J Anim Sci Technol ; 61(2): 55-60, 2019 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-31333862

RESUMEN

Colibacillosis is one of the major health problems in young piglets resulting in poor health and death caused by Escherichia coli producing F18 pili and Shiga toxin 2e. It is pivotal to reduce colibacillosis in weaned piglets to enhance production performance. In this study, we evaluated synbiotics as the gut health improvement agents in the mouse model challenged with Shiga toxin-producing E. coli (STEC) isolated from piglets. Prebiotic lactulose was formulated with each 5.0 × 106 CFU/mL of Pediococcus acidilactici GB-U15, Lactobacillus plantarum GB-U17, and Lactobacillus plantarum GB 1-3 to produce 3 combinations of synbiotics. A total of 40 three weeks old BALB/c mice were randomly assigned to 4 groups (n = 10): a control group and 3 synbiotics treated groups. Each treatment groups were daily administrated with 5.0 × 106 CFU/mL of one synbiotics for the first week, and every 3 days during the second week. All the mice were challenged with 8.0 × 108 CFU/mL of STEC 5 days after animals began to receive synbiotics. Mice treated with synbiotics based on Pediococcus acidilactici GB-U15 and Lactobacillus plantarum GB-U17 significantly improved daily weight gain compared to mice in other groups. While mice treated with GB-U15 showed better fecal index, no significant differences were observed among groups. Gross lesion and histopathological evaluations showed that mice treated with GB-U15 moderately improved recovery from STEC infection. In conclusion, our results suggest that the synbiotics formulated with lactulose and Pediococcus acidilactici GB-U15 have potential benefits to prevent and improve colibacillosis in weaned piglets.

14.
Respir Res ; 9: 26, 2008 Mar 14.
Artículo en Inglés | MEDLINE | ID: mdl-18341691

RESUMEN

BACKGROUND: CD38 is expressed in human airway smooth muscle (HASM) cells, regulates intracellular calcium, and its expression is augmented by tumor necrosis factor alpha (TNF-alpha). CD38 has a role in airway hyperresponsiveness, a hallmark of asthma, since deficient mice develop attenuated airway hyperresponsiveness compared to wild-type mice following intranasal challenges with cytokines such as IL-13 and TNF-alpha. Regulation of CD38 expression in HASM cells involves the transcription factor NF-kappaB, and glucocorticoids inhibit this expression through NF-kappaB-dependent and -independent mechanisms. In this study, we determined whether the transcriptional regulation of CD38 expression in HASM cells involves response elements within the promoter region of this gene. METHODS: We cloned a putative 3 kb promoter fragment of the human cd38 gene into pGL3 basic vector in front of a luciferase reporter gene. Sequence analysis of the putative cd38 promoter region revealed one NF-kappaB and several AP-1 and glucocorticoid response element (GRE) motifs. HASM cells were transfected with the 3 kb promoter, a 1.8 kb truncated promoter that lacks the NF-kappaB and some of the AP-1 sites, or the promoter with mutations of the NF-kappaB and/or AP-1 sites. Using the electrophoretic mobility shift assays, we determined the binding of nuclear proteins to oligonucleotides encoding the putative cd38 NF-kappaB, AP-1, and GRE sites, and the specificity of this binding was confirmed by gel supershift analysis with appropriate antibodies. RESULTS: TNF-alpha induced a two-fold activation of the 3 kb promoter following its transfection into HASM cells. In cells transfected with the 1.8 kb promoter or promoter constructs lacking NF-kappaB and/or AP-1 sites or in the presence of dexamethasone, there was no induction in the presence of TNF-alpha. The binding of nuclear proteins to oligonucleotides encoding the putative cd38 NF-kappaB site and some of the six AP-1 sites was increased by TNF-alpha, and to some of the putative cd38 GREs by dexamethasone. CONCLUSION: The EMSA results and the cd38 promoter-reporter assays confirm the functional role of NF-kappaB, AP-1 and GREs in the cd38 promoter in the transcriptional regulation of CD38.


Asunto(s)
ADP-Ribosil Ciclasa 1/genética , Dexametasona/farmacología , Glucocorticoides/farmacología , Glicoproteínas de Membrana/genética , Miocitos del Músculo Liso/fisiología , Tráquea/citología , Factor de Necrosis Tumoral alfa/farmacología , Hiperreactividad Bronquial/fisiopatología , Células Cultivadas , Dexametasona/metabolismo , Ensayo de Cambio de Movilidad Electroforética , Regulación de la Expresión Génica/efectos de los fármacos , Glucocorticoides/metabolismo , Humanos , Luciferasas/genética , Miocitos del Músculo Liso/citología , Miocitos del Músculo Liso/efectos de los fármacos , FN-kappa B/metabolismo , Regiones Promotoras Genéticas/fisiología , Receptores de Glucocorticoides/metabolismo , Factor de Transcripción AP-1/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo
15.
Artículo en Inglés | MEDLINE | ID: mdl-30069307

RESUMEN

BACKGROUND: Understanding the composition of the microbial community and its functional capacity during weaning is important for pig production as bacteria play important roles in the pig's health and growth performance. However, limited information is available regarding the composition and function of the gut microbiome of piglets in early-life. Therefore, we performed 16S rRNA gene and whole metagenome shotgun sequencing of DNA from fecal samples from healthy piglets during weaning to measure microbiome shifts, and to identify the potential contribution of the early-life microbiota in shaping piglet health with a focus on microbial stress responses, carbohydrate and amino acid metabolism. RESULTS: The analysis of 16S rRNA genes and whole metagenome shotgun sequencing revealed significant compositional and functional differences between the fecal microbiome in nursing and weaned piglets. The fecal microbiome of the nursing piglets showed higher relative abundance of bacteria in the genus Bacteroides with abundant gene families related to the utilization of lactose and galactose. Prevotella and Lactobacillus were enriched in weaned piglets with an enrichment for the gene families associated with carbohydrate and amino acid metabolism. In addition, an analysis of the functional capacity of the fecal microbiome showed higher abundances of genes associated with heat shock and oxidative stress in the metagenome of weaned piglets compared to nursing piglets. CONCLUSIONS: Overall, our data show that microbial shifts and changes in functional capacities of the piglet fecal microbiome resulted in potential reductions in the effects of stress, including dietary changes that occur during weaning. These results provide us with new insights into the piglet gut microbiome that contributes to the growth of the animal.

16.
FASEB J ; 20(7): 1000-2, 2006 May.
Artículo en Inglés | MEDLINE | ID: mdl-16571778

RESUMEN

The transmembrane glycoprotein CD38 catalyzes the synthesis of the calcium mobilizing molecule cyclic ADP-ribose from NAD. In human airway smooth muscle (HASM) cells, the expression and function of CD38 are augmented by the inflammatory cytokine tumor necrosis factor-alpha (TNF-alpha), leading to increased intracellular calcium response to agonists. A glucocorticoid response element in the CD38 gene has been computationally described, providing evidence for transcriptional regulation of its expression. In the present study, we investigated the effects of dexamethasone, a glucocorticoid, on CD38 expression and ADP-ribosyl cyclase activity in HASM cells stimulated with TNF-alpha. In HASM cells, TNF-alpha augmented CD38 expression and ADP-ribosyl cyclase activity, which were attenuated by dexamethasone. TNF-alpha increased NF-kappaB expression and its activation, and dexamethasone partially reversed these effects. TNF-alpha increased the expression of IkappaBalpha, and dexamethasone increased it further. An inhibitor of NF-kappaB activation or transfection of cells with IkappaB mutants decreased TNF-alpha-induced CD38 expression. The results indicate that TNF-alpha-induced CD38 expression involves NF-kappaB expression and its activation and dexamethasone inhibits CD38 expression through NF-kappaB-dependent and -independent mechanisms.


Asunto(s)
ADP-Ribosil Ciclasa 1/metabolismo , Dexametasona/farmacología , Glicoproteínas de Membrana/metabolismo , Miocitos del Músculo Liso/efectos de los fármacos , Miocitos del Músculo Liso/metabolismo , FN-kappa B/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , ADP-Ribosil Ciclasa 1/genética , Antiinflamatorios/farmacología , Células Cultivadas , Humanos , Glicoproteínas de Membrana/genética , Tráquea , Transcripción Genética
17.
Neuropsychiatr Dis Treat ; 12: 1019-27, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-27175079

RESUMEN

PURPOSE: Decision-making is the process of forming preferences for possible options, selecting and executing actions, and evaluating the outcome. This study used the Iowa Gambling Task (IGT) and the Prospect Valence Learning (PVL) model to investigate deficits in risk-reward related decision-making in patients with chronic schizophrenia, and to identify decision-making processes that contribute to poor IGT performance in these patients. MATERIALS AND METHODS: Thirty-nine patients with schizophrenia and 31 healthy controls participated. Decision-making was measured by total net score, block net scores, and the total number of cards selected from each deck of the IGT. PVL parameters were estimated with the Markov chain Monte Carlo sampling scheme in OpenBugs and BRugs, its interface to R, and the estimated parameters were analyzed with the Mann-Whitney U-test. RESULTS: The schizophrenia group received significantly lower total net scores compared to the control group. In terms of block net scores, an interaction effect of group × block was observed. The block net scores of the schizophrenia group did not differ across the five blocks, whereas those of the control group increased as the blocks progressed. The schizophrenia group obtained significantly lower block net scores in the fourth and fifth blocks of the IGT and selected cards from deck D (advantageous) less frequently than the control group. Additionally, the schizophrenia group had significantly lower values on the utility-shape, loss-aversion, recency, and consistency parameters of the PVL model. CONCLUSION: These results indicate that patients with schizophrenia experience deficits in decision-making, possibly due to failure in learning the expected value of each deck, and incorporating outcome experiences of previous trials into expectancies about options in the present trial.

18.
J Vet Sci ; 3(4): 247-54, 2002 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-12819374

RESUMEN

The detrimental effects of environmental pollutants on the health of the individual are generally accepted, although the mechanisms of these effects remain to be incompletely understood. In the present study, we examined the effects of B[a]P, 2-BP, phenol and TCDD on proinflammatory cytokine gene expression in mice spleen cells which were stimulated with anti-CD3. 10(-9)M TCDD increased IFNgamma and TNFalpha gene expression, but suppressed IL-1 gene expression. 10(-6)M phenol inhibited IL-1, IL-6 and TNFalpha gene expression, and 10(-6)M of 2-BP downregulated TNFalpha gene expression. However, 10(-6)M of B[a]P did not influence on IL-1, IL-6, IFNgamma and TNFalpha gene expression. These findings suggest that TCDD may impair the immune functions of mice by enhancing proinflammatory cytokines production, whereas phenol and 2-BP may impair the functions by inhibiting the production of these cytokines.


Asunto(s)
Benzo(a)pireno/toxicidad , Citocinas/biosíntesis , Contaminantes Ambientales/toxicidad , Hidrocarburos Bromados/toxicidad , Fenol/toxicidad , Dibenzodioxinas Policloradas/toxicidad , Bazo/efectos de los fármacos , Animales , Apoptosis/efectos de los fármacos , Complejo CD3/inmunología , Células Cultivadas , Citocinas/genética , Expresión Génica/efectos de los fármacos , Masculino , Ratones , Ratones Endogámicos C3H , ARN/química , ARN/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Bazo/metabolismo
19.
Nat Commun ; 4: 2479, 2013.
Artículo en Inglés | MEDLINE | ID: mdl-24056518

RESUMEN

ORM (yeast)-like protein isoform 3 (ORMDL3) has recently been identified as a candidate gene for susceptibility to asthma; however, the mechanisms by which it contributes to asthma pathogenesis are not well understood. Here we demonstrate a functional role for ORMDL3 in eosinophils in the context of allergic inflammation. Eosinophils recruited to the airways of allergen-challenged mice express ORMDL3. ORMDL3 expression in bone marrow eosinophils is localized in the endoplasmic reticulum and is induced by interleukin-3 and eotaxin-1. Overexpression of ORMDL3 in eosinophils causes increased rolling, distinct cytoskeletal rearrangement, extracellular signal-regulated kinase (1/2) phosphorylation and nuclear translocation of nuclear factor kappa B. Knockdown of ORMDL3 significantly inhibits activation-induced cell shape changes, adhesion and recruitment to sites of inflammation in vivo, combined with reduced expression of CD49d and CD18. In addition, ORMDL3 regulates interleukin-3-induced expression of CD48 and CD48-mediated eosinophil degranulation. These studies show that ORMDL3 regulates eosinophil trafficking, recruitment and degranulation, further elucidating a role for this molecule in allergic asthma and potentially other eosinophilic disorders.


Asunto(s)
Antígenos CD/genética , Asma/genética , Antígenos CD18/genética , Eosinófilos/metabolismo , Integrina alfa4/genética , Proteínas de la Membrana/genética , Citoesqueleto de Actina/efectos de los fármacos , Animales , Antígenos CD/metabolismo , Asma/metabolismo , Asma/patología , Antígenos CD18/metabolismo , Antígeno CD48 , Adhesión Celular/efectos de los fármacos , Degranulación de la Célula/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Forma de la Célula , Quimiocina CCL11/farmacología , Eosinófilos/efectos de los fármacos , Eosinófilos/patología , Regulación de la Expresión Génica , Integrina alfa4/metabolismo , Interleucina-3/farmacología , Masculino , Proteínas de la Membrana/antagonistas & inhibidores , Proteínas de la Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Proteína Quinasa 1 Activada por Mitógenos/genética , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/genética , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , FN-kappa B/genética , FN-kappa B/metabolismo , Fosforilación , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Transducción de Señal
20.
PLoS One ; 8(1): e54840, 2013.
Artículo en Inglés | MEDLINE | ID: mdl-23372779

RESUMEN

Association of the neurotransmitter serotonin (5-HT) with the pathogenesis of allergic asthma is well recognized and its role as a chemoattractant for eosinophils (Eos) in vitro and in vivo has been previously demonstrated. Here we have examined the regulation of 5-HT-induced human and murine Eos trafficking and migration at a cellular and molecular level. Eos from allergic donors and bone marrow-derived murine Eos (BM-Eos) were found to predominantly express the 5-HT2A receptor. Exposure to 5-HT or 2,5-dimethoxy-4-iodoamphetamine (DOI), a 5-HT2A/C selective agonist, induced rolling of human Eos and AML14.3D10 human Eos-like cells on vascular cell adhesion molecule (VCAM)-1 under conditions of flow in vitro coupled with distinct cytoskeletal and cell shape changes as well as phosphorylation of MAPK. Blockade of 5-HT2A or of ROCK MAPK, PI3K, PKC and calmodulin, but not G(αi)-proteins, with specific inhibitors inhibited DOI-induced rolling, actin polymerization and changes in morphology of VCAM-1-adherent AML14.3D10 cells. More extensive studies with murine BM-Eos demonstrated the role of 5-HT in promoting rolling in vivo within inflamed post-capillary venules of the mouse cremaster microcirculation and confirmed that down-stream signaling of 5-HT2A activation involves ROCK, MAPK, PI3K, PKC and calmodulin similar to AML14.3D10 cells. DOI-induced migration of BM-Eos is also dependent on these signaling molecules and requires Ca(2+). Further, activation of 5-HT2A with DOI led to an increase in intracellular Ca(2+) levels in murine BM-Eos. Overall, these data demonstrate that 5-HT (or DOI)/5-HT2A interaction regulates Eos trafficking and migration by promoting actin polymerization associated with changes in cell shape/morphology that favor cellular trafficking and recruitment via activation of specific intracellular signaling molecules (ROCK, MAPK, PI3K and the PKC-calmodulin pathway).


Asunto(s)
Quimiotaxis de Leucocito/efectos de los fármacos , Eosinófilos/efectos de los fármacos , Eosinófilos/inmunología , Serotonina/farmacología , Animales , Calmodulina/metabolismo , Células Cultivadas , Eosinófilos/metabolismo , Humanos , Rodamiento de Leucocito/efectos de los fármacos , Masculino , Ratones , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Proteína Quinasa C/metabolismo , Transducción de Señal , Quinasas Asociadas a rho/metabolismo
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