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1.
Am J Physiol Lung Cell Mol Physiol ; 305(11): L866-77, 2013 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-24077949

RESUMO

We sought to investigate the effects of cockroach allergen (CRA) exposure on the lung macrophage population to determine how different macrophage phenotypes influence exacerbation of disease. CRA exposure caused significantly reduced expression of CD86 on lung macrophages. These effects were not systemic, as peritoneal macrophage CD86 expression was not altered. To investigate whether naïve macrophages could reduce asthma-like pulmonary inflammation, autologous peritoneal macrophages were instilled into the airways 24 h before the final CRA challenge. Pulmonary inflammation was assessed by measurement of airway hyperresponsiveness, mucin production, inflammatory cell recruitment, and cytokine production. Cell transfer did not have significant effects in control mice, nor did it affect pulmonary mucin production or airway hyperresponsiveness in control or CRA-exposed mice. However, there was significant reduction in the number of eosinophils recovered in the bronchoalveolar lavage (BAL) (5.8 × 105 vs. 0.88 × 105), and total cell recruitment to the airways of CRA-exposed mice was markedly reduced (1.1 × 106 vs. 0.57 × 106). The reduced eosinophil recruitment was reflected by substantially lower levels of eosinophil peroxidase in the lung and significantly lower concentrations of eotaxins in BAL (eotaxin 1: 3 pg/ml vs. undetectable; eotaxin 2: 2,383 vs. 131 pg/ml) and lung homogenate (eotaxin 1: 1,043 vs. 218 pg/ml; eotaxin 2: 10 vs. 1.5 ng/ml). We conclude that CRA decreases lung macrophage CD86 expression. Furthermore, supplementation of the lung cell population with peritoneal macrophages inhibits eosinophil recruitment, achieved through reduction of eotaxin production. These data demonstrate that transfer of naïve macrophages will reduce some aspects of asthma-like pulmonary inflammation in response to CRA.


Assuntos
Asma/imunologia , Quimiocina CCL11/biossíntese , Quimiocina CCL24/biossíntese , Baratas/imunologia , Eosinófilos/imunologia , Macrófagos Peritoneais/imunologia , Alérgenos/imunologia , Animais , Animais não Endogâmicos , Líquido da Lavagem Broncoalveolar , Células Cultivadas , Técnicas de Cocultura , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Macrófagos Alveolares/imunologia , Macrófagos Peritoneais/metabolismo , Macrófagos Peritoneais/transplante , Camundongos , Camundongos Endogâmicos ICR , Mucinas/metabolismo , Neutrófilos/imunologia
2.
Am J Pathol ; 181(3): 845-57, 2012 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-22796441

RESUMO

Asthma may be triggered by multiple mediators, including allergen-IgE cross-linking and non-IgE mechanisms. Several clinical studies have shown acute ethanol consumption exacerbates asthma, yet no animal model exists to study this process. We developed a model of ethanol-triggered asthma in allergen-sensitized mice to evaluate the mechanisms of ethanol inducing asthma-like responses. Outbred mice were exposed to cockroach allergens on Days 0 and 14; and on Day 21, mice received ethanol by oral gavage. Tracer studies confirmed alcohol aspiration did not occur. Within 30 minutes, alcohol induced degranulation of over 74% of mast cells, and multiple parameters of asthma-like pulmonary inflammation were triggered. Ethanol-gavaged mice had a fivefold increased production of eotaxin-2 (534 pg/mL) and a sevenfold increase in bronchoalveolar eosinophils (70,080 cells). Ethanol induced a 10-fold increase in IL-13, from 84 pg/mL in sensitized mice to 845 pg/mL in ethanol-gavaged sensitized mice. In cockroach allergen-sensitized mice, ethanol triggered asthma-like changes in respiratory physiology and a significant fivefold increase in airway mucin production. Importantly, none of these asthmatic exacerbations were observed in normal mice gavaged with ethanol. Cromolyn sodium effectively stabilized mast cells, yet increased mucin production and bronchoalveolar eosinophil recruitment. Together, these data show a single oral alcohol exposure will trigger asthma-like pulmonary inflammation in allergen-sensitized mice, providing a novel asthma model.


Assuntos
Alérgenos/imunologia , Asma/imunologia , Etanol/administração & dosagem , Imunização , Administração Oral , Consumo de Bebidas Alcoólicas/patologia , Intoxicação Alcoólica/complicações , Intoxicação Alcoólica/imunologia , Intoxicação Alcoólica/patologia , Intoxicação Alcoólica/fisiopatologia , Animais , Asma/complicações , Asma/patologia , Asma/fisiopatologia , Degranulação Celular , Citocinas/biossíntese , Modelos Animais de Doenças , Eosinofilia/complicações , Eosinofilia/imunologia , Eosinofilia/patologia , Eosinofilia/fisiopatologia , Eosinófilos/patologia , Feminino , Pulmão/imunologia , Pulmão/patologia , Pulmão/fisiopatologia , Mastócitos/patologia , Mastócitos/fisiologia , Camundongos , Camundongos Endogâmicos ICR , Mucinas/biossíntese , Pneumonia/complicações , Pneumonia/imunologia , Pneumonia/patologia , Pneumonia/fisiopatologia , Testes de Função Respiratória , Células Th2/imunologia
3.
J Immunol ; 182(12): 7763-75, 2009 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-19494300

RESUMO

HLA-B27- and -B57-positive HIV-infected humans have long been associated with control of HIV replication, implying that CD8(+) T cell responses contribute to control of viral replication. In a similar fashion, 50% of Mamu-B*08-positive Indian rhesus macaques control SIVmac239 replication and become elite controllers with chronic-phase viremia <1000 viral RNA copies/ml. Interestingly, Mamu-B*08-restricted SIV-derived epitopes appeared to match the peptide binding profile for HLA-B*2705 in humans. We therefore defined a detailed peptide-binding motif for Mamu-B*08 and investigated binding similarities between the macaque and human MHC class I molecules. Analysis of a panel of approximately 900 peptides revealed that despite substantial sequence differences between Mamu-B*08 and HLA-B*2705, the peptide-binding repertoires of these two MHC class I molecules share a remarkable degree of overlap. Detailed knowledge of the Mamu-B*08 peptide-binding motif enabled us to identify six additional novel Mamu-B*08-restricted SIV-specific CD8(+) T cell immune responses directed against epitopes in Gag, Vpr, and Env. All 13 Mamu-B*08-restricted epitopes contain an R at the position 2 primary anchor and 10 also possess either R or K at the N terminus. Such dibasic peptides are less prone to cellular degradation. This work highlights the relevance of the Mamu-B*08-positive SIV-infected Indian rhesus macaque as a model to examine elite control of immunodeficiency virus replication. The remarkable similarity of the peptide-binding motifs and repertoires for Mamu-B*08 and HLA-B*2705 suggests that the nature of the peptide bound by the MHC class I molecule may play an important role in control of immunodeficiency virus replication.


Assuntos
Antígenos de Histocompatibilidade Classe I/imunologia , Peptídeos/imunologia , Peptídeos/metabolismo , Vírus da Imunodeficiência Símia/imunologia , Replicação Viral , Sequência de Aminoácidos , Animais , DNA Polimerase Dirigida por DNA , Epitopos de Linfócito T/química , Epitopos de Linfócito T/imunologia , Antígenos de Histocompatibilidade Classe I/química , Humanos , Ligantes , Macaca mulatta , Dados de Sequência Molecular , Biblioteca de Peptídeos , Ligação Proteica , Homologia de Sequência de Aminoácidos
4.
J Virol ; 83(22): 11514-27, 2009 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-19726517

RESUMO

An understanding of the mechanism(s) by which some individuals spontaneously control human immunodeficiency virus (HIV)/simian immunodeficiency virus replication may aid vaccine design. Approximately 50% of Indian rhesus macaques that express the major histocompatibility complex (MHC) class I allele Mamu-B*08 become elite controllers after infection with simian immunodeficiency virus SIVmac239. Mamu-B*08 has a binding motif that is very similar to that of HLA-B27, a human MHC class I allele associated with the elite control of HIV, suggesting that SIVmac239-infected Mamu-B*08-positive (Mamu-B*08+) animals may be a good model for the elite control of HIV. The association with MHC class I alleles implicates CD8+ T cells and/or natural killer cells in the control of viral replication. We therefore introduced point mutations into eight Mamu-B*08-restricted CD8+ T-cell epitopes to investigate the contribution of epitope-specific CD8+ T-cell responses to the development of the control of viral replication. Ten Mamu-B*08+ macaques were infected with this mutant virus, 8X-SIVmac239. We compared immune responses and viral loads of these animals to those of wild-type SIVmac239-infected Mamu-B*08+ macaques. The five most immunodominant Mamu-B*08-restricted CD8+ T-cell responses were barely detectable in 8X-SIVmac239-infected animals. By 48 weeks postinfection, 2 of 10 8X-SIVmac239-infected Mamu-B*08+ animals controlled viral replication to <20,000 viral RNA (vRNA) copy equivalents (eq)/ml plasma, while 10 of 15 wild-type-infected Mamu-B*08+ animals had viral loads of <20,000 vRNA copy eq/ml (P = 0.04). Our results suggest that these epitope-specific CD8+ T-cell responses may play a role in establishing the control of viral replication in Mamu-B*08+ macaques.


Assuntos
Antígenos de Histocompatibilidade Classe I/imunologia , Síndrome de Imunodeficiência Adquirida dos Símios/virologia , Vírus da Imunodeficiência Símia/patogenicidade , Replicação Viral/genética , Animais , Linhagem Celular , Primers do DNA , Epitopos de Linfócito T/imunologia , Genes MHC Classe I/imunologia , Variação Genética/imunologia , Macaca mulatta/imunologia , Macaca mulatta/virologia , Mutagênese Sítio-Dirigida , Vírus da Imunodeficiência Símia/genética , Carga Viral
5.
Clin Cancer Res ; 26(8): 1953-1964, 2020 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-31964784

RESUMO

PURPOSE: To assess the potential for CUE-101, a novel therapeutic fusion protein, to selectively activate and expand HPV16 E711-20-specific CD8+ T cells as an off-the shelf therapy for the treatment of HPV16-driven tumors, including head and neck squamous cell carcinoma (HNSCC), cervical, and anal cancers. EXPERIMENTAL DESIGN: CUE-101 is an Fc fusion protein composed of a human leukocyte antigen (HLA) complex, an HPV16 E7 peptide epitope, reduced affinity human IL2 molecules, and an effector attenuated human IgG1 Fc domain. Human E7-specific T cells and human peripheral blood mononuclear cells (PBMC) were tested to demonstrate cellular activity and specificity of CUE-101, whereas in vivo activity of CUE-101 was assessed in HLA-A2 transgenic mice. Antitumor efficacy with a murine surrogate (mCUE-101) was tested in the TC-1 syngeneic tumor model. RESULTS: CUE-101 demonstrates selective binding, activation, and expansion of HPV16 E711-20-specific CD8+ T cells from PBMCs relative to nontarget cells. Intravenous administration of CUE-101 induced selective expansion of HPV16 E711-20-specific CD8+ T cells in HLA-A2 (AAD) transgenic mice, and anticancer efficacy and immunologic memory was demonstrated in TC-1 tumor-bearing mice treated with mCUE-101. Combination therapy with anti-PD-1 checkpoint blockade further enhanced the observed efficacy. CONCLUSIONS: Consistent with its design, CUE-101 demonstrates selective expansion of an HPV16 E711-20-specific population of cytotoxic CD8+ T cells, a favorable safety profile, and in vitro and in vivo evidence supporting its potential for clinical efficacy in an ongoing phase I trial (NCT03978689).


Assuntos
Linfócitos T CD8-Positivos/imunologia , Antígeno HLA-A2/imunologia , Fragmentos Fc das Imunoglobulinas/imunologia , Interleucina-2/imunologia , Neoplasias/terapia , Proteínas E7 de Papillomavirus/imunologia , Membro 7 da Superfamília de Receptores de Fatores de Necrose Tumoral/imunologia , Animais , Células Cultivadas , Modelos Animais de Doenças , Feminino , Voluntários Saudáveis , Humanos , Leucócitos Mononucleares , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neoplasias/imunologia , Neoplasias/virologia
6.
J Virol ; 82(4): 1723-38, 2008 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-18057253

RESUMO

Certain major histocompatibility complex (MHC) class I alleles are strongly associated with control of human immunodeficiency virus and simian immunodeficiency virus (SIV). CD8(+) T cells specific for epitopes restricted by these molecules may be particularly effective. Understanding how CD8(+) T cells contribute to control of viral replication should yield important insights for vaccine design. We have recently identified an Indian rhesus macaque MHC class I allele, Mamu-B*08, associated with elite control and low plasma viremia after infection with the pathogenic isolate SIVmac239. Here, we infected four Mamu-B*08-positive macaques with SIVmac239 to investigate why some of these macaques control viral replication. Three of the four macaques controlled SIVmac239 replication with plasma virus concentrations below 20,000 viral RNA copies/ml at 20 weeks postinfection; two of four macaques were elite controllers (ECs). Interestingly, two of the four macaques preserved their CD4(+) memory T lymphocytes during peak viremia, and all four recovered their CD4(+) memory T lymphocytes in the chronic phase of infection. Mamu-B*08-restricted CD8(+) T-cell responses dominated the acute phase and accounted for 23.3% to 59.6% of the total SIV-specific immune responses. Additionally, the ECs mounted strong and broad CD8(+) T-cell responses against several epitopes in Vif and Nef. Mamu-B*08-specific CD8(+) T cells accounted for the majority of mutations in the virus at 18 weeks postinfection. Interestingly, patterns of viral variation in Nef differed between the ECs and the other two macaques. Natural containment of AIDS virus replication in Mamu-B*08-positive macaques may, therefore, be related to a combination of immunodominance and viral escape from CD8(+) T-cell responses.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Macaca mulatta/imunologia , Síndrome de Imunodeficiência Adquirida dos Símios/imunologia , Vírus da Imunodeficiência Símia/fisiologia , Replicação Viral , Alelos , Sequência de Aminoácidos , Animais , Epitopos de Linfócito T/genética , Epitopos de Linfócito T/imunologia , Antígenos de Histocompatibilidade Classe I/genética , Antígenos de Histocompatibilidade Classe I/imunologia , Epitopos Imunodominantes/genética , Epitopos Imunodominantes/imunologia , Macaca mulatta/virologia , Dados de Sequência Molecular , Vírus da Imunodeficiência Símia/genética , Vírus da Imunodeficiência Símia/imunologia
7.
Hum Exp Toxicol ; 22(11): 593-605, 2003 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-14686482

RESUMO

The epithelial Madin Darby canine kidney (MDCK) cell line, Caucasian renal leiomyoblastoma (G-402) cells, human small airways epithelial (HSAE) cells, human bronchial epithelial (HBE) cells and human renal proximal tubule (HRPT) epithelial cells were examined for sensitivity to Clostridium perfringens biotype D epsilon-toxin. MDCK and G-402 cells were confirmed as being the only established cell lines that are sensitive to the toxin. HSAE, HBE and HRPT epithelial cells were only found to be sensitive to the toxin at concentrations of > 1 mg/ mL. Cultures of MDCK and G-402 cells, with increased resistance (tolerance) to the cytotoxic effects of epsilon-toxin, were developed by exposing these cultures to progressively higher concentrations of toxin. The greatest relative increase in tolerance to epsilon-toxin was developed in MDCK cells, in which the LC50 in control cultures was 2 microg/mL as determined by the MTS/PMS assay system; after selection for tolerance, this was raised to 100 microg/mL. This represents a 50-fold increase in tolerance as measured by this index. Using G-402 cells, it was possible to increase the LC50 by twofold from 290 to 590 microg/mL. Subsequent 2-D electrophoresis of membrane preparations from tolerant and control MDCK cells revealed that the expression of a discrete group of proteins found in control cells with a range of molecular weights from 32-36 kDa, all with acidic isoelectric points (IEPs), were either not expressed in epsilon-toxin tolerant cells or had undergone a shift in IEP to a more alkaline pH in tolerant cells. This suggests that epsilon-toxin lethality in MDCK cells may be mediated by membrane-located proteins. Their absence or alteration in toxin-resistant cells would, at least partly, explain the failure of most cell lines to demonstrate sensitivity to this toxin, despite being derived from tissues that are damaged by epsilon-toxin. This approach may have utility in the study of other toxin-cell interactions and could be used in the development of novel medical countermeasures by identifying cellular targets which mediate toxin lethality.


Assuntos
Toxinas Bacterianas/farmacologia , Toxinas Bacterianas/toxicidade , Proteínas de Membrana/fisiologia , Animais , Técnicas de Cultura de Células , Cães , Resistência a Medicamentos , Eletroforese em Gel Bidimensional , Humanos , Rim/citologia , Neoplasias Renais/patologia , Leiomioma Epitelioide/patologia , Células Tumorais Cultivadas
8.
Shock ; 37(1): 56-62, 2012 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-21921828

RESUMO

Sepsis is one of the leading causes of death in hospitals worldwide. Even with optimal therapy, severe sepsis results in 50% mortality, indicating variability in the response of individuals towards treatment. We hypothesize that the presence of preexisting antibodies present in the blood before the onset of sepsis induced by cecal ligation and puncture (CLP) in mice accounts for the differences in their survival. A plasma-enhanced killing (PEK) assay was performed to calculate the PEK capacity of plasma, that is, the ability of plasma to augment polymorphonuclear neutrophil killing of bacteria. Plasma-enhanced killing was calculated as PEK = [1 / log (N)] × 100, where N = number of surviving bacteria; a higher PEK indicated better bacterial killing. A range of PEK in plasma collected from mice before CLP was observed, documenting individual differences in bacterial killing capacity. Mortality was predicted based on plasma IL-6 levels at 24 h after CLP. Mice predicted to die (Die-P) had a lower PEK (<14) and higher peritoneal bacterial counts at 24 h after sepsis compared with those predicted to live (Live-P) with a PEK of greater than 16. Mice with PEK of less than 14 were 3.1 times more likely to die compared with the group with PEK of greater than 16. To understand the mechanism of defense conferred by the preexisting antibodies, binding of IgM or IgG to enteric bacteria was documented by flow cytometry. To determine the relative contribution of IgM or IgG, the immunoglobulins were specifically immunodepleted from the naive plasma samples and the PEK of the depleted plasma measured. Compared with naive plasma, depletion of IgM had no effect on the PEK. However, depletion of IgG increased PEK, suggesting that an inhibitory IgG binds to antigenic sites on bacteria preventing optimal opsonization of the bacteria. These data demonstrate that, before CLP, circulating inhibitory IgG antibodies exist that prevent bacterial killing by polymorphonuclear neutrophils in a CLP model of sepsis.


Assuntos
Anticorpos Antibacterianos/sangue , Anticorpos Antibacterianos/imunologia , Enterobacteriaceae/imunologia , Sepse/sangue , Sepse/imunologia , Animais , Modelos Animais de Doenças , Feminino , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Imunoglobulina M/sangue , Imunoglobulina M/imunologia , Camundongos , Camundongos Endogâmicos ICR , Neutrófilos/imunologia , Neutrófilos/metabolismo
9.
J Virol ; 81(16): 8827-32, 2007 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-17537848

RESUMO

Certain major histocompatibility complex (MHC) class I alleles are associated with the control of human immunodeficiency virus and simian immunodeficiency virus (SIV) replication. We have designed sequence-specific primers for detection of the rhesus macaque MHC class I allele Mamu-B*08 by PCR and screened a cohort of SIV-infected macaques for this allele. Analysis of 196 SIV(mac)239-infected Indian rhesus macaques revealed that Mamu-B*08 was significantly overrepresented in elite controllers; 38% of elite controllers were Mamu-B*08 positive compared to 3% of progressors (P = 0.00001). Mamu-B*08 was also associated with a 7.34-fold decrease in chronic phase viremia (P = 0.002). Mamu-B*08-positive macaques may, therefore, provide a good model to understand the correlates of MHC class I allele-associated immune protection and viral containment in human elite controllers.


Assuntos
Antígenos de Histocompatibilidade Classe I/genética , Síndrome de Imunodeficiência Adquirida dos Símios/genética , Vírus da Imunodeficiência Símia/fisiologia , Replicação Viral , Alelos , Animais , Sequência de Bases , Testes Genéticos , Antígenos de Histocompatibilidade Classe I/metabolismo , Macaca mulatta , Dados de Sequência Molecular , Síndrome de Imunodeficiência Adquirida dos Símios/imunologia , Viremia/genética , Viremia/imunologia
10.
PLoS One ; 2(11): e1152, 2007 Nov 14.
Artigo em Inglês | MEDLINE | ID: mdl-18000532

RESUMO

BACKGROUND: It is generally accepted that CD8+ T cell responses play an important role in control of immunodeficiency virus replication. The association of HLA-B27 and -B57 with control of viremia supports this conclusion. However, specific correlates of viral control in individuals expressing these alleles have been difficult to define. We recently reported that transient in vivo CD8+ cell depletion in simian immunodeficiency virus (SIV)-infected elite controller (EC) macaques resulted in a brief period of viral recrudescence. SIV replication was rapidly controlled with the reappearance of CD8+ cells, implicating that these cells actively suppress viral replication in ECs. METHODS AND FINDINGS: Here we show that three ECs in that study made at least seven robust CD8+ T cell responses directed against novel epitopes in Vif, Rev, and Nef restricted by the MHC class I molecule Mamu-B*08. Two of these Mamu-B*08-positive animals subsequently lost control of SIV replication. Their breakthrough virus harbored substitutions in multiple Mamu-B*08-restricted epitopes. Indeed, we found evidence for selection pressure mediated by Mamu-B*08-restricted CD8+ T cells in all of the newly identified epitopes in a cohort of chronically infected macaques. CONCLUSIONS: Together, our data suggest that Mamu-B*08-restricted CD8+ T cell responses effectively control replication of pathogenic SIV(mac)239. All seven regions encoding Mamu-B*08-restricted CD8+ T cell epitopes also exhibit amino acid replacements typically seen only in the presence of Mamu-B*08, suggesting that the variation we observe is indeed selected by CD8+ T cell responses. SIV(mac)239 infection of Indian rhesus macaques expressing Mamu-B*08 may therefore provide an animal model for understanding CD8+ T cell-mediated control of HIV replication in humans.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Epitopos/imunologia , Variação Genética , Vírus da Imunodeficiência Símia/imunologia , Animais , Sequência de Bases , Linfócitos T CD8-Positivos/virologia , Primers do DNA , Ensaio de Imunoadsorção Enzimática , Macaca mulatta , Reação em Cadeia da Polimerase , Polimorfismo Conformacional de Fita Simples , RNA Viral/genética , Vírus da Imunodeficiência Símia/genética , Vírus da Imunodeficiência Símia/fisiologia , Replicação Viral
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