RESUMO
Human immunodeficiency virus 1 (HIV-1) infection is associated with heightened inflammation and excess risk of cardiovascular disease, cancer and other complications. These pathologies persist despite antiretroviral therapy. In two independent cohorts, we found that innate lymphoid cells (ILCs) were depleted in the blood and gut of people with HIV-1, even with effective antiretroviral therapy. ILC depletion was associated with neutrophil infiltration of the gut lamina propria, type 1 interferon activation, increased microbial translocation and natural killer (NK) cell skewing towards an inflammatory state, with chromatin structure and phenotype typical of WNT transcription factor TCF7-dependent memory T cells. Cytokines that are elevated during acute HIV-1 infection reproduced the ILC and NK cell abnormalities ex vivo. These results show that inflammatory cytokines associated with HIV-1 infection irreversibly disrupt ILCs. This results in loss of gut epithelial integrity, microbial translocation and memory NK cells with heightened inflammatory potential, and explains the chronic inflammation in people with HIV-1.
Assuntos
Citocinas/sangue , HIV-1/imunologia , HIV-1/patogenicidade , Imunidade Inata , Células Matadoras Naturais/imunologia , Linfócitos/imunologia , Fator 1 de Transcrição de Linfócitos T/imunologia , Regulação da Expressão Gênica , Infecções por HIV/genética , Infecções por HIV/imunologia , Infecções por HIV/virologia , Homeostase/imunologia , Humanos , Memória Imunológica , Técnicas In Vitro , Inflamação/genética , Inflamação/imunologia , Inflamação/virologia , Fator 1 de Transcrição de Linfócitos T/genética , Via de Sinalização Wnt/imunologiaRESUMO
Candida auris is a yeast that is difficult to eradicate and has caused outbreaks in health care facilities. We report a cluster of 5 patients in 1 intensive care unit who were colonized or infected in 2017. The initial 2 patients were recipients of liver transplants who had cultures that grew C auris within 3 days of each other in June 2017 (days 43 and 30 posttransplant). Subsequent screening cultures identified 2 additional patients with C auris colonization. Respiratory and urine cultures from a fifth patient yielded C auris. All isolates were fluconazole resistant but susceptible to echinocandins. Whole genome sequencing showed the strains were clonal, suggesting in-hospital transmission, and related but distinct from New York/New Jersey strains, consistent with a separate introduction. However, no source or contact was found. Two of the 5 patients died. C auris infection likely contributed to 1 patient death by infecting a vascular aneurysm at the graft anastomosis. Strict infection control precautions were initiated to control the outbreak. Our experience reveals that although severe disease from C auris can occur in transplant recipients, outbreaks can be controlled using recommended infection control practices. We have had no further patients infected with C auris to date.
Assuntos
Transplante de Fígado , Antifúngicos/uso terapêutico , Candida , Candidíase Invasiva , Cuidados Críticos , Surtos de Doenças , Humanos , Unidades de Terapia Intensiva , Transplante de Fígado/efeitos adversos , Testes de Sensibilidade MicrobianaRESUMO
Virus-specific CD8(+) T cells expand dramatically during acute EBV infection, and their persistence is important for lifelong control of EBV-related disease. To better define the generation and maintenance of these effective CD8(+) T cell responses, we used microarrays to characterize gene expression in total and EBV-specific CD8(+) T cells isolated from the peripheral blood of 10 individuals followed from acute infectious mononucleosis (AIM) into convalescence (CONV). In total CD8(+) T cells, differential expression of genes in AIM and CONV was most pronounced among those encoding proteins important in T cell activation/differentiation, cell division/metabolism, chemokines/cytokines and receptors, signaling and transcription factors (TF), immune effector functions, and negative regulators. Within these categories, we identified 28 genes that correlated with CD8(+) T cell expansion in response to an acute EBV infection. In EBV-specific CD8(+) T cells, we identified 33 genes that were differentially expressed in AIM and CONV. Two important TF, T-bet and eomesodermin, were upregulated and maintained at similar levels in both AIM and CONV; in contrast, protein expression declined from AIM to CONV. Expression of these TF varied among cells with different epitope specificities. Collectively, gene and protein expression patterns suggest that a large proportion, if not a majority of CD8(+) T cells in AIM are virus specific, activated, dividing, and primed to exert effector activities. High expression of T-bet and eomesodermin may help to maintain effector mechanisms in activated cells and to enable proliferation and transition to earlier differentiation states in CONV.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Herpesvirus Humano 4/imunologia , Mononucleose Infecciosa/imunologia , Transcriptoma , ADP-Ribosil Ciclase 1/genética , Doença Aguda , Adolescente , Adulto , Feminino , Humanos , Masculino , Receptores de Interleucina-7/genética , Fatores de Transcrição/genéticaRESUMO
We report a case of probable Zaire Ebola virus-related ophthalmologic complications in a physician from the United States who contracted Ebola virus disease in Liberia. Uveitis, immune activation, and nonspecific increase in antibody titers developed during convalescence. This case highlights immune phenomena that could complicate management of Ebola virus disease-related uveitis during convalescence.
Assuntos
Ebolavirus , Doença pelo Vírus Ebola/complicações , Doença pelo Vírus Ebola/virologia , Uveíte/diagnóstico , Uveíte/etiologia , Ebolavirus/genética , Doença pelo Vírus Ebola/diagnóstico , Humanos , Libéria , Masculino , Tomografia de Coerência Óptica , Uveíte/tratamento farmacológicoRESUMO
UNLABELLED: We report the diversity of latent membrane protein 1 (LMP1) gene founder sequences and the level of Epstein-Barr virus (EBV) genome variability over time and across anatomic compartments by using virus genomes amplified directly from oropharyngeal wash specimens and peripheral blood B cells during acute infection and convalescence. The intrahost nucleotide variability of the founder virus was 0.02% across the region sequences, and diversity increased significantly over time in the oropharyngeal compartment (P = 0.004). The LMP1 region showing the greatest level of variability in both compartments, and over time, was concentrated within the functional carboxyl-terminal activating regions 2 and 3 (CTAR2 and CTAR3). Interestingly, a deletion in a proline-rich repeat region (amino acids 274 to 289) of EBV commonly reported in EBV sequenced from cancer specimens was not observed in acute infectious mononucleosis (AIM) patients. Taken together, these data highlight the diversity in circulating EBV genomes and its potential importance in disease pathogenesis and vaccine design. IMPORTANCE: This study is among the first to leverage an improved high-throughput deep-sequencing methodology to investigate directly from patient samples the degree of diversity in Epstein-Barr virus (EBV) populations and the extent to which viral genome diversity develops over time in the infected host. Significant variability of circulating EBV latent membrane protein 1 (LMP1) gene sequences was observed between cellular and oral wash samples, and this variability increased over time in oral wash samples. The significance of EBV genetic diversity in transmission and disease pathogenesis are discussed.
Assuntos
Linfócitos B/virologia , Infecções por Vírus Epstein-Barr/virologia , Variação Genética , Herpesvirus Humano 4/genética , Orofaringe/virologia , Proteínas da Matriz Viral/genética , Análise por Conglomerados , DNA Viral/química , DNA Viral/genética , Herpesvirus Humano 4/isolamento & purificação , Humanos , Dados de Sequência Molecular , Filogenia , Análise de Sequência de DNA , Fatores de Tempo , Adulto JovemRESUMO
The resistance of human immunodeficiency virus type 1 (HIV-1) to antibody-mediated immunity often prevents the detection of antibodies that neutralize primary isolates of HIV-1. However, conventional assays for antibody functions other than neutralization are suboptimal. Current methods for measuring the killing of virus-infected cells by antibody-dependent cell-mediated cytotoxicity (ADCC) are limited by the number of natural killer (NK) cells obtainable from individual donors, donor-to-donor variation, and the use of nonphysiological targets. We therefore developed an ADCC assay based on NK cell lines that express human or macaque CD16 and a CD4(+) T-cell line that expresses luciferase from a Tat-inducible promoter upon HIV-1 or simian immunodeficiency virus (SIV) infection. NK cells and virus-infected targets are mixed in the presence of serial plasma dilutions, and ADCC is measured as the dose-dependent loss of luciferase activity. Using this approach, ADCC titers were measured in plasma samples from HIV-infected human donors and SIV-infected macaques. For the same plasma samples paired with the same test viruses, this assay was approximately 2 orders of magnitude more sensitive than optimized assays for neutralizing antibodies-frequently allowing the measurement of ADCC in the absence of detectable neutralization. Although ADCC correlated with other measures of Env-specific antibodies, neutralizing and gp120 binding titers did not consistently predict ADCC activity. Hence, this assay affords a sensitive method for measuring antibodies capable of directing ADCC against HIV- or SIV-infected cells expressing native conformations of the viral envelope glycoprotein and reveals incomplete overlap of the antibodies that direct ADCC and those measured in neutralization and binding assays.
Assuntos
Anticorpos Antivirais/química , HIV-1/metabolismo , Testes de Neutralização/métodos , Vírus da Imunodeficiência Símia/metabolismo , Animais , Citotoxicidade Celular Dependente de Anticorpos , Linfócitos T CD4-Positivos/virologia , Linhagem Celular , Ensaio de Imunoadsorção Enzimática/métodos , Proteína gp120 do Envelope de HIV/metabolismo , Humanos , Imunoglobulina G/química , Células Matadoras Naturais/virologia , Macaca , Ligação Proteica , Receptores de IgG/biossíntese , Reprodutibilidade dos Testes , Virologia/métodosRESUMO
Viral reservoirs that persist in HIV-1 infected individuals on antiretroviral therapy (ART) are the major obstacle to viral eradication. The identification and definition of viral reservoirs in patients on ART is needed in order to understand viral persistence and achieve the goal of viral eradication. We examined whether analysis of episomal HIV-1 genomes provided the means to characterize virus that persists during ART and whether it could reveal the virus that contributes to treatment failure in patients on ART. For six individuals in which virus replication was highly suppressed for at least 20 months, proviral and episomal genomes present just prior to rebound were phylogenetically compared to RNA genomes of rebounding virus after therapy interruption. Episomal envelope sequences, but not proviral envelope sequences, were highly similar to sequences in rebounding virus. Since episomes are products of recent infections, the phylogenetic relationships support the conclusion that viral rebound originated from a cryptic viral reservoir. To evaluate whether the reservoir revealed by episomal sequence analysis was of clinical relevance, we examined whether episomal sequences define a viral population that contributes to virologic failure in individuals receiving the CCR5 antagonist, Vicriviroc. Episomal envelope sequences at or near baseline predicted treatment failure due to the presence of X4 or D/M (dual/mixed) viral variants. In patients that did not harbor X4 or D/M viruses, the basis for Vicriviroc treatment failure was indeterminate. Although these samples were obtained from viremic patients, the assay would be applicable to a large percentage of aviremic patients, based on previous studies. Summarily, the results support the use of episomal HIV-1 as an additional or alternative approach to traditional assays to characterize virus that is maintained during long-term, suppressive ART.
Assuntos
DNA Complementar/genética , Infecções por HIV/tratamento farmacológico , HIV-1/genética , Viremia/tratamento farmacológico , Adolescente , Terapia Antirretroviral de Alta Atividade , Antagonistas dos Receptores CCR5 , Farmacorresistência Viral/genética , Genoma Viral , Infecções por HIV/genética , Infecções por HIV/imunologia , Humanos , Filogenia , Piperazinas/uso terapêutico , Pirimidinas/uso terapêutico , RNA Viral , Falha de Tratamento , Carga Viral , Viremia/genética , Viremia/imunologia , Replicação ViralRESUMO
SARS coronavirus 1 (SARS-CoV-1) causes a respiratory infection that can lead to acute respiratory distress characterized by inflammation and high levels of cytokines in the lung tissue. In this study we constructed a herpes simplex virus 1 replication-defective mutant vector expressing SARS-CoV-1 spike protein as a potential vaccine vector and to probe the effects of spike protein on host cells. The spike protein expressed from this vector is functional in that it localizes to the surface of infected cells and induces fusion of ACE2-expressing cells. In immunized mice, the recombinant vector induced antibodies that bind to spike protein in an ELISA assay and that show neutralizing activity. The spike protein expressed from this vector can induce the expression of cytokines in an ACE2-independent, MyD88-dependent process. These results argue that the SARS-CoV-1 spike protein intrinsically activates signaling pathways that induce cytokines and contribute directly to the inflammatory process of SARS.
Assuntos
Anticorpos Neutralizantes/imunologia , Herpesvirus Humano 1/genética , Imunidade Inata , Coronavírus Relacionado à Síndrome Respiratória Aguda Grave/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Animais , Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Fusão Celular , Linhagem Celular , Citocinas/imunologia , Vetores Genéticos , Humanos , Camundongos , Coronavírus Relacionado à Síndrome Respiratória Aguda Grave/genética , Transdução de Sinais , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/metabolismo , Vacinas de Partículas Semelhantes a Vírus/imunologia , Vacinas Virais/imunologiaRESUMO
In most human immunodeficiency virus type 1 (HIV-1)-infected individuals who achieve viral loads of <50 copies/ml during highly active antiretroviral therapy (HAART), low levels of plasma virus remain detectable for years by ultrasensitive methods. The relative contributions of ongoing virus replication and virus production from HIV-1 reservoirs to persistent low-level viremia during HAART remain controversial. HIV-1 vaccination of HAART-treated individuals provides a model for examining low-level viremia, as immunizations may facilitate virus replication and sequence evolution. In a phase 1 trial of modified vaccinia virus Ankara/fowlpox virus-based HIV-1 vaccines in 20 HIV-infected young adults receiving HAART, we assessed the prevalence of low-level viremia and sequence evolution, using ultrasensitive viral load (<6.5 copies/ml) and genotyping (five-copy sensitivity) assays. Viral evolution, consisting of new drug resistance mutations and novel amino acid changes within a relevant HLA-restricted allele (e.g., methionine, isoleucine, glutamine, or arginine for leucine at position 205 of RT), was found in 1 and 3 of 20 subjects, respectively. Sequence evolution was significantly correlated with levels of viremia of between 6.5 and <50 copies/ml (P = 0.03) and was more likely to occur within epitopes presented by relevant HLA alleles (P < 0.001). These findings suggest that ongoing virus replication contributes to low-level viremia in patients on HAART and that this ongoing replication is subject to CD8(+) T-cell selective pressures.
Assuntos
Antirretrovirais/farmacologia , Infecções por HIV/virologia , HIV-1/metabolismo , Poxviridae/genética , Adulto , Terapia Antirretroviral de Alta Atividade , Linfócitos T CD4-Positivos/virologia , Linfócitos T CD8-Positivos/virologia , Estudos de Coortes , Feminino , Genótipo , Infecções por HIV/tratamento farmacológico , Humanos , Imunização , Masculino , Análise de Sequência de DNARESUMO
Spike (S) proteins of coronaviruses, including the coronavirus that causes severe acute respiratory syndrome (SARS), associate with cellular receptors to mediate infection of their target cells. Here we identify a metallopeptidase, angiotensin-converting enzyme 2 (ACE2), isolated from SARS coronavirus (SARS-CoV)-permissive Vero E6 cells, that efficiently binds the S1 domain of the SARS-CoV S protein. We found that a soluble form of ACE2, but not of the related enzyme ACE1, blocked association of the S1 domain with Vero E6 cells. 293T cells transfected with ACE2, but not those transfected with human immunodeficiency virus-1 receptors, formed multinucleated syncytia with cells expressing S protein. Furthermore, SARS-CoV replicated efficiently on ACE2-transfected but not mock-transfected 293T cells. Finally, anti-ACE2 but not anti-ACE1 antibody blocked viral replication on Vero E6 cells. Together our data indicate that ACE2 is a functional receptor for SARS-CoV.
Assuntos
Coronavírus Relacionado à Síndrome Respiratória Aguda Grave/metabolismo , Enzima de Conversão de Angiotensina 2 , Animais , Anticorpos/imunologia , Anticorpos/farmacologia , Carboxipeptidases/antagonistas & inibidores , Carboxipeptidases/genética , Carboxipeptidases/imunologia , Carboxipeptidases/metabolismo , Linhagem Celular , Chlorocebus aethiops , Células Gigantes/citologia , Células Gigantes/metabolismo , Humanos , Glicoproteínas de Membrana/química , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Peso Molecular , Peptidil Dipeptidase A/imunologia , Peptidil Dipeptidase A/metabolismo , Ligação Proteica , Estrutura Terciária de Proteína , Receptores Virais/antagonistas & inibidores , Receptores Virais/genética , Receptores Virais/imunologia , Receptores Virais/metabolismo , Coronavírus Relacionado à Síndrome Respiratória Aguda Grave/genética , Coronavírus Relacionado à Síndrome Respiratória Aguda Grave/crescimento & desenvolvimento , Solubilidade , Glicoproteína da Espícula de Coronavírus , Transfecção , Células Vero , Proteínas do Envelope Viral/química , Proteínas do Envelope Viral/genética , Proteínas do Envelope Viral/metabolismo , Replicação Viral/efeitos dos fármacosRESUMO
Plasmacytoid dendritic cells (PDC) are major producers of type I interferons (IFN) in response to human immunodeficiency virus type 1 (HIV-1) infection. To better define the underlying mechanisms, we studied the magnitude of alpha IFN (IFN-alpha) induction by recombinant viruses containing changes in the Env protein that impair or disrupt CD4 binding or expressing primary env alleles with differential coreceptor tropism. We found that the CD4 binding affinity but not the viral coreceptor usage is critical for the attachment of autofluorescing HIV-1 to PDC and for subsequent IFN-alpha induction. Our results illustrate the importance of the gp120-CD4 interaction in determining HIV-1-induced immune stimulation via IFN-alpha production.
Assuntos
Antígenos CD4/imunologia , Células Dendríticas/imunologia , HIV-1/imunologia , Interferon-alfa/biossíntese , Adolescente , Adulto , Antígenos CD4/metabolismo , Linhagem Celular , Células Cultivadas , Criança , HIV-1/isolamento & purificação , HIV-1/fisiologia , Humanos , Rim/citologia , Pessoa de Meia-Idade , Transfecção , Replicação ViralRESUMO
OBJECTIVE: Timely initiation of combination antiretroviral therapy (ART) limits latent HIV reservoir size and should also limit reservoir genetic complexity. However, the relationship between these two factors remains unclear, particularly among HIV-infected youth. DESIGN: Retrospective analysis of replication-competent latent HIV clones serially isolated by limiting-dilution culture from resting CD4 T-cell reservoirs from ART-suppressed, young adult participants of a historic phase I therapeutic vaccine trial (PACTG/IMPAACT-P1059). METHODS: Replication-competent latent HIV clones isolated from resting CD4 T cells of four perinatally and 10 nonperinatally infected young adults (average 22 versus 6 years uncontrolled infection, respectively) were sequenced in Pol and Nef. Within-host HIV sequence datasets were characterized with respect to their genetic diversity and inferred immune escape mutation burden. RESULTS: Although participants were comparable in terms of sociodemographic and HIV sampling characteristics (e.g. on average, a mean 17 Pol sequences were recovered at five timepoints over up to 70 weeks) and the length of ART suppression at study entry (average 3 years), replication-competent HIV reservoir size, genetic diversity, immune escape mutation burden and variant complexity were significantly higher among the perinatally infected participants who experienced longer durations of uncontrolled viremia. Nevertheless, viral sequences inferred to retain susceptibility to host cellular immune responses were detected in all participants, irrespective of uncontrolled viremia duration. CONCLUSION: HIV elimination in late-suppressed youth may be doubly challenged by larger and more genetically complex reservoirs. Strategies that integrate host and viral genetic complexity to achieve HIV remission or cure may merit consideration in such cases.
Assuntos
Antirretrovirais/uso terapêutico , Variação Genética , Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , HIV/classificação , HIV/genética , Latência Viral , Adolescente , Linfócitos T CD4-Positivos/virologia , DNA Viral/química , DNA Viral/genética , Feminino , Humanos , Masculino , Estudos Retrospectivos , Análise de Sequência de DNA , Adulto Jovem , Produtos do Gene nef do Vírus da Imunodeficiência Humana/genética , Produtos do Gene pol do Vírus da Imunodeficiência Humana/genéticaRESUMO
Recent reports have determined that HIV-1 Vif counteracts an innate antiviral cellular factor, Apobec3G. However, the function of Vif during HIV-1 pathogenesis remains poorly understood. To gain a better understanding of Vif function, the viral isolate from an HIV-1-infected long-term nonprogressor (LTNP) that displayed a Vif-mutant replication phenotype was studied. This LTNP has been infected since before 1983 and has no HIV-related disease in the absence of antiretroviral therapy. From separate samples, obtained on more than one study visit, virus grew in cocultures of LTNP cells with Vif-complementing T cell lines, but not the parental T cell lines. An unusual amino acid motif (KKRK) was found in the Vif sequence at positions 90 to 93. Since this motif commonly functions as a nuclear localization sequence, experiments were performed to determine the ability of this KKRK motif to mediate nuclear localization of Vif. Wild-type Vif displayed a predominantly cytoplasmic distribution. In contrast, the KKRK Vif showed a predominantly nuclear localization. The effect of the KKRK mutation on virus production and infectivity was also studied. The KKRK motif that mislocalizes Vif to the nucleus also reduces viral replication and infectivity in nonpermissive cells. Our data highlight the importance of Vif in HIV-1 pathogenesis and also provide a unique tool to investigate the interaction of Vif and Apobec3G.
Assuntos
Núcleo Celular/metabolismo , HIV-1/patogenicidade , Motivos de Aminoácidos , Sequência de Aminoácidos , Linhagem Celular , Produtos do Gene vif/química , Produtos do Gene vif/genética , Produtos do Gene vif/metabolismo , Sobreviventes de Longo Prazo ao HIV , HIV-1/genética , HIV-1/metabolismo , Humanos , Dados de Sequência Molecular , Mutação , Análise de Sequência de DNA , Produtos do Gene vif do Vírus da Imunodeficiência HumanaRESUMO
Monoclonal antibodies (Mabs) against the Urbani strain of the SARS-associated coronavirus (SARS-CoV) were developed and characterized for reactivity to SARS-CoV and SARS-CoV S, N, M, and E proteins using enzyme-linked immunoabsorbent (ELISA), radioimmunoprecipitation, immunofluorescence, Western Blot and microneutralization assays. Twenty-six mAbs were reactive to SARS-CoV by ELISA, and nine were chosen for detailed characterization. Five mAbs reacted against the S protein, two against the M protein, and one each against the N and E proteins. Two of five S protein mAbs neutralized SARS-CoV infection of Vero E6 cells and reacted to an epitope within amino acids 490-510 in the S protein. While two of the three non-neutralizing antibodies recognized at second epitope within amino acids 270-350. The mAbs characterized should prove useful for developing SARS-CoV diagnostic assays and for studying the biology of infection and pathogenesis of disease.
Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Antivirais/imunologia , Especificidade de Anticorpos , Coronavírus Relacionado à Síndrome Respiratória Aguda Grave/imunologia , Proteínas Estruturais Virais/imunologia , Animais , Anticorpos Monoclonais/biossíntese , Anticorpos Antivirais/biossíntese , Linhagem Celular , Chlorocebus aethiops , Proteínas M de Coronavírus , Proteínas do Nucleocapsídeo de Coronavírus , Mapeamento de Epitopos , Humanos , Glicoproteínas de Membrana/imunologia , Testes de Neutralização , Proteínas do Nucleocapsídeo/imunologia , Síndrome Respiratória Aguda Grave/virologia , Glicoproteína da Espícula de Coronavírus , Células Vero , Proteínas do Envelope Viral/imunologia , Proteínas da Matriz Viral/imunologia , Proteínas ViroporinasRESUMO
The detection of replication-competent human immunodeficiency virus type 1 (HIV-1) in infected individuals is integral for studies of viral latency and reservoirs of continuing replication during treatment. We describe a modified coculture method to detect/isolate virus from HIV-1-infected individuals with low or undetectable plasma viral loads. We observed a wide range in CD4 and chemokine receptor concentrations on CD4(+) T cells of HIV-1-uninfected donors. We selected cells from donors who expressed high levels of CCR5 after mitogen stimulation to combine in culture with peripheral blood mononuclear cells of HIV-1-infected individuals. Using this donor-enhanced culture method, viruses were isolated from asymptomatic adults with longterm nonprogressive infection, and children receiving effective highly active antiretroviral therapy, whereas parallel cultures with cells expressing lower levels of CCR5 yielded none. Virus was isolated from an individual for whom all previous attempts using reported methods were unsuccessful. Virus growth from cryopreserved cells from this same individual was detected at multiple sampling time points, including a time point at which a sensitive assay to measure 2-LTR (long terminal repeat) circular forms of the viral genome was negative. This will be a useful technique by which to study viral latency and HIV-1 pathogenesis in adult and pediatric populations.
Assuntos
Infecções por HIV/virologia , HIV-1/isolamento & purificação , Monócitos/virologia , Replicação Viral , Adulto , Criança , Técnicas de Cocultura , Infecções por HIV/sangue , HIV-1/fisiologia , HumanosRESUMO
OBJECTIVES: Therapeutic HIV vaccinations may alter the size of the resting memory CD4 T-cell latent HIV reservoir as HIV establishes latency when memory responses are formed, including those toward HIV. Alternatively, latently infected CD4 T cells maybe killed, while exiting the reservoir upon activation. METHODS: The effect of therapeutic immunization with modified vaccinia Ankara and Fowlpox-based HIV vaccines on the latent reservoir was examined in 19 young adults who were receiving effective antiretroviral therapy. Correlations between size of the reservoir [measured in infectious units per million (IUPM)] resting CD4 T cells and HIV-specific immune responses, including immune activation were examined. Decay of the reservoir was assessed using random-effects model. RESULTS: A modest transient decrease in the size of the reservoir was observed at week 40 [mean -0.31âlog(10) IUPM (95% confidence interval: -0.60 to -0.03; Pâ=â0.03] following HIV vaccinations. The estimated half-life (T1/2) of the reservoir during the 40 weeks following vaccination was 9.8 months and statistically different from zero (Pâ=â0.02), but 35.3 months and not different from zero (Pâ=â0.21) over 72 weeks of study. Latent reservoir size at baseline was not correlated with HIV-specific CD4, CD8 responses or immune activation, but became correlated with CD4 IFNγ (râ=â0.54, Pâ=â0.02) and IL-2 responses at 6 weeks after immunization (râ=â0.48, Pâ=â0.04). CONCLUSION: Therapeutic HIV vaccinations led to a transient increase in decay of latently infected CD4 T cells. Further studies of therapeutic HIV vaccines may provide important insights into facilitating decay of the latent reservoir.
Assuntos
Vacinas contra a AIDS/uso terapêutico , Linfócitos T CD4-Positivos/imunologia , Infecções por HIV/tratamento farmacológico , Vacinas contra a AIDS/imunologia , Infecções por HIV/virologia , Humanos , Resultado do Tratamento , Vacinação , Vacinas Sintéticas/imunologia , Vacinas Sintéticas/uso terapêutico , Adulto JovemRESUMO
Serotype-cross-reactive memory T cells responding to secondary dengue virus (DENV) infection are thought to contribute to disease. However, epitope-specific T cell responses have not been thoroughly compared between subjects with primary versus secondary DENV infection. We studied CD8(+) T cells specific for the HLA-A*1101-restricted NS3(133) epitope in a cohort of A11(+) DENV-infected patients throughout acute illness and convalescence. We compared the expansion, serotype-cross-reactivity, and activation of these cells in PBMC from patients experiencing primary or secondary infection and mild or severe disease by flow cytometry. Our results show expansion and activation of DENV-specific CD8(+) T cells during acute infection, which are predominantly serotype-cross-reactive regardless of DENV infection history. These data confirm marked T cell activation and serotype-cross-reactivity during the febrile phase of dengue; however, A11-NS3(133)-specific responses did not correlate with prior antigenic exposure or current disease severity.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Coinfecção/imunologia , Dengue/imunologia , Dengue/patologia , Criança , Pré-Escolar , Reações Cruzadas/imunologia , Humanos , Lactente , Recém-NascidoRESUMO
BACKGROUND: Programmed Death-1 (PD-1) is an inhibitory member of the CD28 family of molecules expressed on CD8+ T cells in response to antigenic stimulation. To better understand the role of PD-1 in antiviral immunity we examined the expression of PD-1 on Epstein-Barr virus (EBV) epitope-specific CD8+ T cells during acute infectious mononucleosis (AIM) and convalescence. METHODOLOGY/PRINCIPAL FINDINGS: Using flow cytometry, we observed higher frequencies of EBV-specific CD8+ T cells and higher intensity of PD-1 expression on EBV-specific CD8+ T cells during AIM than during convalescence. PD-1 expression during AIM directly correlated with viral load and with the subsequent degree of CD8+ T cell contraction in convalescence. Consistent differences in PD-1 expression were observed between CD8+ T cells with specificity for two different EBV lytic antigen epitopes. Similar differences were observed in the degree to which PD-1 was upregulated on these epitope-specific CD8+ T cells following peptide stimulation in vitro. EBV epitope-specific CD8+ T cell proliferative responses to peptide stimulation were diminished during AIM regardless of PD-1 expression and were unaffected by blocking PD-1 interactions with PD-L1. Significant variability in PD-1 expression was observed on EBV epitope-specific CD8+ T cell subsets defined by V-beta usage. CONCLUSIONS/SIGNIFICANCE: These observations suggest that PD-1 expression is not only dependent on the degree of antigen presentation, but also on undefined characteristics of the responding cell that segregate with epitope specificity and V-beta usage.
Assuntos
Antígenos CD/genética , Proteínas Reguladoras de Apoptose/genética , Linfócitos T CD8-Positivos/imunologia , Herpesvirus Humano 4/fisiologia , Mononucleose Infecciosa/genética , Mononucleose Infecciosa/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Antígenos CD/imunologia , Proteínas Reguladoras de Apoptose/imunologia , Linfócitos T CD8-Positivos/virologia , Células Cultivadas , Expressão Gênica , Herpesvirus Humano 4/imunologia , Humanos , Mononucleose Infecciosa/virologia , Receptor de Morte Celular Programada 1 , Receptores de Antígenos de Linfócitos T/genética , Regulação para CimaRESUMO
Memory B cells latently infected with Epstein-Barr virus (mB(Lats)) in the blood disappear rapidly on presentation with acute symptomatic primary infection (acute infectious mononucleosis [AIM]). They undergo a simple exponential decay (average half-life: 7.5 +/- 3.7 days) similar to that of normal memory B cells. The cytotoxic T lymphocyte (CTL) response to immediate early (IE) lytic antigens (CTL(IEs)) also decays over this time period, but no such correlation was observed for the CTL response to lytic or latent antigens or to the levels of virions shed into saliva. We have estimated the average half-life of CTL(IEs) to be 73 (+/- 23) days. We propose that cycles of infection and reactivation occur in the initial stages of infection that produce high levels of mB(Lats) in the circulation. Eventually the immune response arises and minimizes these cycles leaving the high levels of mB(Lats) in the blood to decay through simple memory B-cell homeostasis mechanisms. This triggers the cells to reactivate the virus whereupon most are killed by CTL(IEs) before they can release virus and infect new cells. The release of antigens caused by this large-scale destruction of infected cells may trigger the symptoms of AIM and be a cofactor in other AIM-associated diseases.
Assuntos
Herpesvirus Humano 4/imunologia , Herpesvirus Humano 4/patogenicidade , Mononucleose Infecciosa/imunologia , Mononucleose Infecciosa/virologia , Doença Aguda , Proliferação de Células , Citotoxicidade Imunológica/imunologia , Memória Imunológica/imunologia , Mononucleose Infecciosa/diagnóstico , Fatores de TempoRESUMO
B lymphocyte hyperactivation and elevated immunoglobulin levels (hypergammaglobulinemia) are pathogenic manifestations of HIV-1 infection. Here we provide evidence that these hallmarks are caused by a soluble factor whose production by infected macrophages is induced by the HIV-1 Nef protein. In vitro, HIV-1-infected macrophages or macrophages expressing Nef promoted B cell activation and differentiation to immunoglobulin-secreting cells. Nef-mediated activation of NF-kappaB in macrophages induced secretion of the acute-phase protein ferritin, and ferritin was necessary and sufficient for the observed Nef-dependent B cell changes. The extent of hypergammaglobulinemia in HIV-1-infected individuals correlated directly with plasma ferritin levels and with viral load. Furthermore, the induction of ferritin production and hypergammaglobulinemia was recapitulated when Nef was specifically expressed in macrophages and T cells of transgenic mice. Collectively, these results indicate that the HIV-1 Nef protein carries a pathogenic determinant that governs B cell defects in HIV-1 infection.