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1.
PLoS One ; 15(8): e0238316, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32866201

RESUMO

BACKGROUND: Perinatally HIV-infected children on anti-retroviral treatment (ART) are reported to have metabolic abnormalities such as dyslipidemia, lipodystrophy, and insulin resistance which potentially increase the risk of diabetes, kidney, liver and cardiovascular disease. OBJECTIVE: To elucidate HIV-mediated metabolic complications that sustain even during ART in perinatally HIV-infected children. METHOD: We have carried out metabolic profiling of the plasma of treatment-naïve and ART-suppressed perinatally HIV-infected children and uninfected controls using 1H nuclear magnetic resonance (NMR) spectroscopy followed by statistical analysis and annotation. RESULT: Validated multivariate analysis showed clear distinction among our study groups. Our results showed elevated levels of lactate, glucose, phosphoenolpyruvic acid, propionic acid, 2-ketobutyric acid and tricarboxylic acid (TCA) cycle metabolites in untreated HIV-infected children compared to uninfected controls. ART normalized the levels of several metabolites, however the level of lactate, phosphoenolpyruvic acid, oxoglutaric acid, oxaloacetic acid, myoinositol and glutamine remained upregulated despite ART in HIV-infected children. Pathway analysis revealed perturbed propanoate metabolism, amino acid metabolism, glycolysis and TCA cycle in untreated and ART-suppressed HIV-infected children. CONCLUSION: Developing therapeutic strategies targeting metabolic abnormalities may be beneficial for preventing diabetes, cardiovascular disease or other associated complications in perinatally HIV-infected children.


Assuntos
Infecções por HIV/metabolismo , Plasma/metabolismo , Antirretrovirais/uso terapêutico , Criança , Estudos Transversais , Feminino , Infecções por HIV/tratamento farmacológico , Humanos , Espectroscopia de Ressonância Magnética/métodos , Masculino , Metaboloma/fisiologia , Metabolômica/métodos , Projetos Piloto , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia
2.
PLoS Pathog ; 16(9): e1008813, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32925973

RESUMO

HIV Nef counteracts cellular host restriction factors SERINC3 and SERINC5, but our understanding of how naturally occurring global Nef sequence diversity impacts these activities is limited. Here, we quantify SERINC3 and SERINC5 internalization function for 339 Nef clones, representing the major pandemic HIV-1 group M subtypes A, B, C and D. We describe distinct subtype-associated hierarchies for Nef-mediated internalization of SERINC5, for which subtype B clones display the highest activities on average, and of SERINC3, for which subtype B clones display the lowest activities on average. We further identify Nef polymorphisms that modulate its ability to counteract SERINC proteins, including substitutions in the N-terminal domain that selectively impair SERINC3 internalization. Our findings demonstrate that the SERINC antagonism activities of HIV Nef differ markedly among major viral subtypes and between individual isolates within a subtype, suggesting that variation in these functions may contribute to global differences in viral pathogenesis.


Assuntos
Infecções por HIV/virologia , HIV-1/fisiologia , Glicoproteínas de Membrana/antagonistas & inibidores , Proteínas de Membrana/antagonistas & inibidores , Polimorfismo Genético , Replicação Viral , Produtos do Gene nef do Vírus da Imunodeficiência Humana/metabolismo , Infecções por HIV/genética , Infecções por HIV/metabolismo , Soropositividade para HIV , Interações Hospedeiro-Patógeno , Humanos , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras/virologia , Células Tumorais Cultivadas , Produtos do Gene nef do Vírus da Imunodeficiência Humana/genética
3.
PLoS Pathog ; 16(8): e1008696, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32760139

RESUMO

HLA-B*35Px is associated with HIV-1 disease rapid progression to AIDS. However, the mechanism(s) underlying this deleterious effect of this HLA allele on HIV-1 infection outcome has not fully understood. CD8+ T cells play a crucial role to control the viral replication but impaired CD8+ T cells represent a major hallmark of HIV-1 infection. Here, we examined the effector functions of CD8+ T cells restricted by HLA-B*35Px (HLA-B*35:03 and HLA-B*35:02), HLA-B*27/B57 and non-HLA-B*27/B57 (e.g. HLA-A*01, A*02, A*03, A*11, A*24, A*26, B*40, B*08, B*38, B*44). CD8+ T cells restricted by HLA-B*35Px exhibited an impaired phenotype compared with those restricted by HLA-B*27/B57 and even non-HLA-B*27/B57. CD8+ T cells restricted by non-HLA-B*27/B57 when encountered their cognate epitopes upregulated TIM-3 and thus became suppressed by regulatory T cells (Tregs) via TIM-3: Galectin-9 (Gal-9). Strikingly, CD8+ T cells restricted by HLA-B*35Px expressed fewer TIM-3 and therefore did not get suppressed by Tregs, which was similar to CD8+ T cells restricted by HLA-B*27/B57. Instead, CD8+ T cells restricted by HLA-B*35Px upon recognition of their cognate epitopes upregulated CTLA-4. The transcriptional and impaired phenotype (e.g. poor effector functions) of HIV-specific CD8+ T cells restricted by HLA-B*35 was related to persistent CTLA-4, elevated Eomes and blimp-1 but poor T-bet expression. As such, anti-CTLA-4 antibody, Ipilimumab, reversed the impaired proliferative capacity of antigen-specific CD8+ T cells restricted by HLA-B*35Px but not others. This study supports the concept that CD8+ T resistance to Tregs-mediated suppression is related to allele restriction rather than the epitope specificity. Our results aid to explain a novel mechanism for the inability of HIV-specific CD8+ T cells restricted by HLA-B*35Px to control viral replication.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Antígeno CTLA-4/metabolismo , Epitopos de Linfócito T/imunologia , Infecções por HIV/imunologia , HIV-1/imunologia , Antígeno HLA-B35/imunologia , Replicação Viral , Antígeno CTLA-4/imunologia , Citocinas/metabolismo , Infecções por HIV/metabolismo , Infecções por HIV/virologia , HIV-1/metabolismo , Humanos , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/metabolismo , Ativação Linfocitária/imunologia , Fenótipo , Linfócitos T Reguladores/imunologia
4.
PLoS Pathog ; 16(8): e1008646, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32776974

RESUMO

Inositol hexakisphosphate (IP6) potently stimulates HIV-1 particle assembly in vitro and infectious particle production in vivo. However, knockout cells lacking inositol-pentakisphosphate 2-kinase (IPPK-KO), the enzyme that produces IP6 by phosphorylation of inositol pentakisphosphate (IP5), were still able to produce infectious HIV-1 particles at a greatly reduced rate. HIV-1 in vitro assembly can also be stimulated to a lesser extent with IP5, but until recently, it was not known if IP5 could also function in promoting assembly in vivo. Here we addressed whether there is an absolute requirement for IP6 or IP5 in the production of infectious HIV-1 particles. IPPK-KO cells expressed no detectable IP6 but elevated IP5 levels and displayed a 20-100-fold reduction in infectious particle production, correlating with lost virus release. Transient transfection of an IPPK expression vector stimulated infectious particle production and release in IPPK-KO but not wildtype cells. Several attempts to make IP6/IP5 deficient stable cells were not successful, but transient expression of the enzyme multiple inositol polyphosphate phosphatase-1 (MINPP1) into IPPK-KOs resulted in near ablation of IP6 and IP5. Under these conditions, we found that HIV-1 infectious particle production and virus release were essentially abolished (1000-fold reduction) demonstrating an IP6/IP5 requirement. However, other retroviruses including a Gammaretrovirus, a Betaretrovirus, and two non-primate Lentiviruses displayed only a modest (3-fold) reduction in infectious particle production from IPPK-KOs and were not significantly altered by expression of IPPK or MINPP1. The only other retrovirus found to show a clear IP6/IP5 dependence was the primate (macaque) Lentivirus Simian Immunodeficiency Virus, which displayed similar sensitivity as HIV-1. We were not able to determine if producer cell IP6/IP5 is required at additional steps beyond assembly because viral particles devoid of both molecules could not be generated. Finally, we found that loss of IP6/IP5 in viral target cells had no effect on permissivity to HIV-1 infection.


Assuntos
Vetores Genéticos/administração & dosagem , Infecções por HIV/virologia , Fosfatos de Inositol/metabolismo , Lentivirus de Primatas/genética , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Ácido Fítico/metabolismo , Vírion/fisiologia , Animais , Vetores Genéticos/genética , HIV/fisiologia , Infecções por HIV/metabolismo , Infecções por HIV/patologia , Humanos , Fosforilação , Primatas
5.
Proc Natl Acad Sci U S A ; 117(30): 18002-18009, 2020 07 28.
Artigo em Inglês | MEDLINE | ID: mdl-32665438

RESUMO

In combating viral infections, the Fab portion of an antibody could mediate virus neutralization, whereas Fc engagement of Fc-γ receptors (FcγRs) could mediate an array of effector functions. Evidence abounds that effector functions are important in controlling infections by influenza, Ebola, or HIV-1 in animal models. However, the relative contribution of virus neutralization versus effector functions to the overall antiviral activity of an antibody remains unknown. To address this fundamental question in immunology, we utilized our knowledge of HIV-1 dynamics to compare the kinetics of the viral load decline (ΔVL) in infected animals given a wild-type (WT) anti-HIV-1 immunoglobulin G1 (IgG1) versus those given a Fc-Null variant of the same antibody. In three independent experiments in HIV-1-infected humanized mice and one pivotal experiment in simian-human immunodeficiency virus (SHIV)-infected rhesus macaques, an earlier and sharper decline in viral load was consistently detected for the WT antibody. Quantifications of the observed differences indicate that Fc-mediated effector functions accounted for 25-45% of the total antiviral activity in these separate experiments. In this study, Fc-mediated effector functions have been quantified in vivo relative to the contribution of virus neutralization mediated by the Fab.


Assuntos
Anticorpos Anti-HIV/imunologia , Infecções por HIV/imunologia , Infecções por HIV/metabolismo , HIV-1/imunologia , Imunoglobulina G/imunologia , Receptores de IgG/metabolismo , Animais , Anticorpos Neutralizantes/imunologia , Modelos Animais de Doenças , Infecções por HIV/virologia , Interações Hospedeiro-Patógeno/imunologia , Humanos , Camundongos , Camundongos Transgênicos , Testes de Neutralização
6.
Molecules ; 25(12)2020 Jun 26.
Artigo em Inglês | MEDLINE | ID: mdl-32604797

RESUMO

Viruses can be spread from one person to another; therefore, they may cause disorders in many people, sometimes leading to epidemics and even pandemics. New, previously unstudied viruses and some specific mutant or recombinant variants of known viruses constantly appear. An example is a variant of coronaviruses (CoV) causing severe acute respiratory syndrome (SARS), named SARS-CoV-2. Some antiviral drugs, such as remdesivir as well as antiretroviral drugs including darunavir, lopinavir, and ritonavir are suggested to be effective in treating disorders caused by SARS-CoV-2. There are data on the utilization of antiretroviral drugs against SARS-CoV-2. Since there are many studies aimed at the identification of the molecular mechanisms of human immunodeficiency virus type 1 (HIV-1) infection and the development of novel therapeutic approaches against HIV-1, we used HIV-1 for our case study to identify possible molecular pathways shared by SARS-CoV-2 and HIV-1. We applied a text and data mining workflow and identified a list of 46 targets, which can be essential for the development of infections caused by SARS-CoV-2 and HIV-1. We show that SARS-CoV-2 and HIV-1 share some molecular pathways involved in inflammation, immune response, cell cycle regulation.


Assuntos
Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/metabolismo , Mineração de Dados/métodos , Infecções por HIV/epidemiologia , Infecções por HIV/metabolismo , Interações Hospedeiro-Patógeno/imunologia , Pandemias , Pneumonia Viral/epidemiologia , Pneumonia Viral/metabolismo , Anti-Inflamatórios/uso terapêutico , Antígenos de Diferenciação/genética , Antígenos de Diferenciação/imunologia , Antivirais/uso terapêutico , Betacoronavirus/efeitos dos fármacos , Betacoronavirus/imunologia , Betacoronavirus/patogenicidade , Proteínas do Sistema Complemento/genética , Proteínas do Sistema Complemento/imunologia , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/imunologia , Bases de Dados Genéticas , Regulação da Expressão Gênica , Infecções por HIV/tratamento farmacológico , Infecções por HIV/imunologia , HIV-1/efeitos dos fármacos , HIV-1/imunologia , HIV-1/patogenicidade , Interações Hospedeiro-Patógeno/efeitos dos fármacos , Interações Hospedeiro-Patógeno/genética , Humanos , Imunidade Inata/efeitos dos fármacos , Fatores Imunológicos/uso terapêutico , Inflamação , Interferons/genética , Interferons/imunologia , Interleucinas/genética , Interleucinas/imunologia , Redes e Vias Metabólicas/efeitos dos fármacos , Redes e Vias Metabólicas/genética , Redes e Vias Metabólicas/imunologia , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/imunologia , Proteínas Repressoras/genética , Proteínas Repressoras/imunologia , Transdução de Sinais , Receptores Toll-Like/genética , Receptores Toll-Like/imunologia , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/imunologia
7.
PLoS One ; 15(7): e0233877, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32645021

RESUMO

BACKGROUND: The impact of nutritional supplements on weight gain in HIV-infected children on antiretroviral treatment (ART) remains uncertain. Starting supplements depends upon current weight-for-age or other acute malnutrition indicators, producing time-dependent confounding. However, weight-for-age at ART initiation may affect subsequent weight gain, independent of supplement use. Implications for marginal structural models (MSMs) with inverse probability of treatment weights (IPTW) are unclear. METHODS: In the ARROW trial, non-randomised supplement use and weight-for-age were recorded monthly from ART initiation. The effect of supplements on weight-for-age over the first year was estimated using generalised estimating equation MSMs with IPTW, both with and without interaction terms between baseline weight-for-age and time. Separately, data were simulated assuming no supplement effect, with use depending on current weight-for-age, and weight-for-age trajectory depending on baseline weight-for-age to investigate potential bias associated with different MSM specifications. RESULTS: In simulations, despite correctly specifying IPTW, omitting an interaction in the MSM between baseline weight-for-age and time produced increasingly biased estimates as associations between baseline weight-for-age and subsequent weight trajectory increased. Estimates were unbiased when the interaction between baseline weight-for-age and time was included, even if the data were simulated with no such interaction. In ARROW, without an interaction the estimated effect was +0.09 (95%CI +0.02,+0.16) greater weight-for-age gain per month's supplement use; this reduced to +0.03 (-0.04,+0.10) including the interaction. DISCUSSION: This study highlights a specific situation in which MSM model misspecification can occur and impact the resulting estimate. Since an interaction in the MSM (outcome) model does not bias the estimate of effect if the interaction does not exist, it may be advisable to include such a term when fitting MSMs for repeated measures.


Assuntos
Infecções por HIV/dietoterapia , Apoio Nutricional/métodos , Ganho de Peso/efeitos dos fármacos , Antirretrovirais/uso terapêutico , Peso Corporal/efeitos dos fármacos , Peso Corporal/fisiologia , Pré-Escolar , Suplementos Nutricionais/análise , Feminino , HIV/patogenicidade , Infecções por HIV/metabolismo , Humanos , Lactente , Masculino , Modelos Estatísticos , Modelagem Computacional Específica para o Paciente , Projetos de Pesquisa
8.
DNA Cell Biol ; 39(9): 1478-1485, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32584609

RESUMO

Combined antiretroviral therapy has improved quality and life expectancy of people living with human immunodeficiency virus (HIV). However, this therapy increases oxidative stress (OS), which in turn causes alterations in lipid and carbon metabolism, kidney disease, liver cirrhosis, and increased risk of cardiovascular disease. The Klotho gene has been implicated in cardiovascular risk increase. Klotho protein expression at X level decreases the risk of heart disease. HIV-positive people usually present low plasma levels of Klotho; thus, contributing to some extent to an increase in cardiovascular risk for these types of patients, mostly by favoring atherosclerosis. Therefore, our aim is to provide an overview of the effect of OS on Klotho protein and its consequent cardiometabolic alterations in HIV-positive patients on antiretroviral therapy.


Assuntos
Doenças Cardiovasculares/metabolismo , Glucuronidase/metabolismo , Infecções por HIV/metabolismo , Estresse Oxidativo , Animais , Terapia Antirretroviral de Alta Atividade/efeitos adversos , Doenças Cardiovasculares/etiologia , Doenças Cardiovasculares/genética , Glucuronidase/genética , Infecções por HIV/tratamento farmacológico , Humanos
9.
PLoS Pathog ; 16(6): e1008378, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32492044

RESUMO

The HIV-1 reservoir consists of latently infected cells that persist despite antiretroviral therapy (ART). Elucidating the proviral genetic composition of the reservoir, particularly in the context of pre-therapy viral diversity, is therefore important to understanding reservoir formation and the persistence of latently infected cells. Here we investigate reservoir proviral variants from 13 Zambian acutely-infected individuals with additional pre-therapy sampling for a unique comparison to the ART-naïve quasispecies. We identified complete transmitted/founder (TF) viruses from seroconversion plasma samples, and additionally amplified and sequenced HIV-1 from plasma obtained one year post-infection and just prior to ART initiation. While the majority of proviral variants in the reservoir were most closely related to viral variants from the latest pre-therapy time point, we also identified reservoir proviral variants dating to or near the time of infection, and to intermediate time points between infection and treatment initiation. Reservoir proviral variants differing by five or fewer nucleotide changes from the TF virus persisted during treatment in five individuals, including proviral variants that exactly matched the TF in two individuals, one of whom had remained ART-naïve for more than six years. Proviral variants during treatment were significantly less divergent from the TF virus than plasma variants present at the last ART-naïve time point. These findings indicate that reservoir proviral variants are archived throughout infection, recapitulating much of the viral diversity that arises throughout untreated HIV-1 infection, and strategies to target and reduce the reservoir must therefore permit for the clearance of proviruses encompassing this extensive diversity.


Assuntos
Variação Genética , Infecções por HIV/genética , HIV-1/genética , Filogenia , Doença Aguda , Adulto , Antirretrovirais , Feminino , Infecções por HIV/tratamento farmacológico , Infecções por HIV/metabolismo , HIV-1/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Zâmbia
10.
PLoS Pathog ; 16(6): e1008381, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32525948

RESUMO

HIV invades the brain during acute infection. Yet, it is unknown whether long-lived infected brain cells release productive virus that can egress from the brain to re-seed peripheral organs. This understanding has significant implication for the brain as a reservoir for HIV and most importantly HIV interplay between the brain and peripheral organs. Given the sheer number of astrocytes in the human brain and their controversial role in HIV infection, we evaluated their infection in vivo and whether HIV infected astrocytes can support HIV egress to peripheral organs. We developed two novel models of chimeric human astrocyte/human peripheral blood mononuclear cells: NOD/scid-IL-2Rgc null (NSG) mice (huAstro/HuPBMCs) whereby we transplanted HIV (non-pseudotyped or VSVg-pseudotyped) infected or uninfected primary human fetal astrocytes (NHAs) or an astrocytoma cell line (U138MG) into the brain of neonate or adult NSG mice and reconstituted the animals with human peripheral blood mononuclear cells (PBMCs). We also transplanted uninfected astrocytes into the brain of NSG mice and reconstituted with infected PBMCs to mimic a biological infection course. As expected, the xenotransplanted astrocytes did not escape/migrate out of the brain and the blood brain barrier (BBB) was intact in this model. We demonstrate that astrocytes support HIV infection in vivo and egress to peripheral organs, at least in part, through trafficking of infected CD4+ T cells out of the brain. Astrocyte-derived HIV egress persists, albeit at low levels, under combination antiretroviral therapy (cART). Egressed HIV evolved with a pattern and rate typical of acute peripheral infection. Lastly, analysis of human cortical or hippocampal brain regions of donors under cART revealed that astrocytes harbor between 0.4-5.2% integrated HIV gag DNA and 2-7% are HIV gag mRNA positive. These studies establish a paradigm shift in the dynamic interaction between the brain and peripheral organs which can inform eradication of HIV reservoirs.


Assuntos
Astrócitos , Barreira Hematoencefálica , Infecções por HIV , HIV-1/metabolismo , Hipocampo , Liberação de Vírus , Animais , Antirretrovirais/farmacologia , Astrócitos/metabolismo , Astrócitos/patologia , Astrócitos/virologia , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/patologia , Barreira Hematoencefálica/virologia , Linhagem Celular Tumoral , Infecções por HIV/genética , Infecções por HIV/metabolismo , Infecções por HIV/patologia , HIV-1/genética , Hipocampo/metabolismo , Hipocampo/patologia , Hipocampo/virologia , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID
11.
PLoS One ; 15(5): e0223464, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32379830

RESUMO

Resistance associated mutations (RAMs) threaten the long-term success of combination antiretroviral therapy (cART) outcomes for HIV-1 treatment. HIV-1 Integrase (IN) strand transfer inhibitors (INSTIs) have proven to be a viable option for highly specific HIV-1 therapy. The INSTI, Dolutegravir is recommended by the World Health Organization for use as first-line cART. This study aims to understand how RAMs affect the stability of IN, as well as the binding of the drug Dolutegravir to the catalytic pocket of the protein. A homology model of HIV-1 subtype C IN was successfully constructed and validated. The site directed mutator webserver was used to predict destabilizing and/or stabilizing effects of known RAMs while FoldX confirmed any changes in protein energy upon introduction of mutation. Also, interaction analysis was performed between neighbouring residues. Three mutations known to be associated with Raltegravir, Elvitegravir and Dolutegravir resistance were selected; E92Q, G140S and Y143R, for molecular dynamics simulations. The structural quality assessment indicated high reliability of the HIV-1C IN tetrameric structure, with more than 90% confidence in modelled regions. Change in free energy for the three mutants indicated different effects, while simulation analysis showed G140S to have the largest affect on protein stability and flexibility. This was further supported by weaker non-bonded pairwise interaction energy and binding free energy values between the drug DTG and E92Q, Y143R and G140S mutants suggesting reduced binding affinity, as indicated by interaction analysis in comparison to the WT. Our findings suggest the G140S mutant has the strongest effect on the HIV-1C IN protein structure and Dolutegravir binding. To the best of our knowledge, this is the first study that uses the consensus wild type HIV-1C IN sequence to build an accurate 3D model to understand the effect of three known mutations on DTG drug binding in a South Africa context.


Assuntos
Farmacorresistência Viral/genética , Infecções por HIV/metabolismo , Inibidores de Integrase de HIV/metabolismo , Integrase de HIV/genética , Integrase de HIV/metabolismo , HIV-1/enzimologia , Compostos Heterocíclicos com 3 Anéis/metabolismo , Mutação , Sequência de Aminoácidos , Domínio Catalítico/genética , Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , Integrase de HIV/química , Inibidores de Integrase de HIV/uso terapêutico , Compostos Heterocíclicos com 3 Anéis/uso terapêutico , Humanos , Simulação de Dinâmica Molecular , Ligação Proteica/genética , Estabilidade Proteica , Quinolonas/metabolismo , Quinolonas/uso terapêutico , Raltegravir Potássico/metabolismo , Raltegravir Potássico/uso terapêutico , África do Sul , Replicação Viral/efeitos dos fármacos , Replicação Viral/genética
12.
PLoS Biol ; 18(5): e3000660, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32453744

RESUMO

Increased life expectancy of patients diagnosed with HIV in the current era of antiretroviral therapy is unfortunately accompanied with the prevalence of HIV-associated neurocognitive disorders (HANDs) and risk of comorbidities such as Alzheimer-like pathology. HIV-1 transactivator of transcription (Tat) protein has been shown to induce the production of toxic neuronal amyloid protein and also enhance neurotoxicity. The contribution of astrocytes in Tat-mediated amyloidosis remains an enigma. We report here, in simian immunodeficiency virus (SIV)+ rhesus macaques and patients diagnosed with HIV, brain region-specific up-regulation of amyloid precursor protein (APP) and Aß (40 and 42) in astrocytes. In addition, we find increased expression of ß-site cleaving enzyme (BACE1), APP, and Aß in human primary astrocytes (HPAs) exposed to Tat. Mechanisms involved up-regulation of hypoxia-inducible factor (HIF-1α), its translocation and binding to the long noncoding RNA (lncRNA) BACE1-antisense transcript (BACE1-AS), resulting, in turn, in the formation of the BACE1-AS/BACE1 RNA complex, subsequently leading to increased BACE1 protein, and activity and generation of Aß-42. Gene silencing approaches confirmed the regulatory role of HIF-1α in BACE1-AS/BACE1 in Tat-mediated amyloidosis. This is the first report implicating the role of the HIF-1α/lncRNABACE1-AS/BACE1 axis in Tat-mediated induction of astrocytic amyloidosis, which could be targeted as adjunctive therapies for HAND-associated Alzheimer-like comorbidity.


Assuntos
Amiloidose/virologia , Astrócitos/metabolismo , Infecções por HIV/complicações , Transtornos Neurocognitivos/virologia , Produtos do Gene tat do Vírus da Imunodeficiência Humana/metabolismo , Peptídeos beta-Amiloides/metabolismo , Amiloidose/metabolismo , Animais , Encéfalo/metabolismo , Células Cultivadas , Infecções por HIV/metabolismo , HIV-1 , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Macaca mulatta , Pessoa de Meia-Idade , Transtornos Neurocognitivos/metabolismo , Fragmentos de Peptídeos/metabolismo , RNA Longo não Codificante/metabolismo , Regulação para Cima
13.
J Virol ; 94(11)2020 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-32238582

RESUMO

Retroviral replication proceeds through obligate integration of the viral DNA into the host genome. In particular, for the HIV-1 genome to enter the nucleus, it must be led through the nuclear pore complex (NPC). During the HIV-1 cytoplasmic journey, the viral core acts as a shell to protect the viral genetic material from antiviral sensors and ensure an adequate environment for reverse transcription. However, the relatively narrow size of the nuclear pore channel requires that the HIV-1 core is reshaped into a structure that fits the pore. On the other hand, the organization of the viral CA proteins that remain associated with the preintegration complex (PIC) during and after nuclear translocation is still enigmatic. In this study, we analyzed the progressive organizational changes of viral CA proteins within the cytoplasm and the nucleus by immunogold labeling. Furthermore, we set up a novel technology, HIV-1 ANCHOR, which enables the specific detection of the retrotranscribed DNA by fluorescence microscopy, thereby offering the opportunity to uncover the architecture of the potential HIV-1 PIC. Thus, we combined the immunoelectron microscopy and ANCHOR technologies to reveal the presence of DNA- and CA-positive complexes by correlated light and electron microscopy (CLEM). During and after nuclear translocation, HIV-1 appears as a complex of viral DNA decorated by multiple viral CA proteins remodeled in a pearl necklace-like shape. Thus, we could describe how CA proteins are reshaped around the viral DNA to permit the entrance of the HIV-1 in the nucleus. This particular CA protein complex composed of the integrase and the retrotranscribed DNA leads the HIV-1 genome inside the host nucleus. Our findings contribute to the understanding of the early steps of HIV-1 infection and provide new insights into the organization of HIV-1 CA proteins during and after viral nuclear entry. Of note, we are now able to visualize the viral DNA in viral complexes, opening up new perspectives for future studies on virus's fate in the cell nucleus.IMPORTANCE How the reverse-transcribed genome reaches the host nucleus remains a main open question related to the infectious cycle of HIV-1. The HIV-1 core has a size of ∼100 nm, largely exceeding that of the NPC channel (∼39 nm). Thus, a rearrangement of the viral CA protein organization is required to achieve an effective nuclear translocation. The mechanism of this process remains undefined due to the lack of a technology capable of visualizing potential CA subcomplexes in association with the viral DNA in the nucleus of HIV-1-infected cells. By the means of state-of-the-art technologies (HIV-1 ANCHOR system combined with CLEM), our study shows that remodeled viral complexes retain multiple CA proteins but not an intact core or only a single CA monomer. These viral CA complexes associated with the retrotranscribed DNA can be observed inside the nucleus, and they represent a potential PIC. Thus, our study shed light on critical early steps characterizing HIV-1 infection, thereby revealing novel, therapeutically exploitable points of intervention. Furthermore, we developed and provided a powerful tool enabling direct, specific, and high-resolution visualization of intracellular and intranuclear HIV-1 subviral structures.


Assuntos
Linfócitos T CD4-Positivos/metabolismo , Infecções por HIV/metabolismo , Integrase de HIV/metabolismo , HIV-1/metabolismo , Complexos Multiproteicos/metabolismo , Integração Viral , Transporte Ativo do Núcleo Celular , Linfócitos T CD4-Positivos/virologia , Núcleo Celular/genética , Núcleo Celular/metabolismo , Núcleo Celular/virologia , Células HEK293 , Infecções por HIV/genética , Integrase de HIV/genética , HIV-1/genética , Células HeLa , Humanos , Complexos Multiproteicos/genética
14.
PLoS Pathog ; 16(4): e1008507, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32282853

RESUMO

The HIV-1 capsid protein makes up the core of the virion and plays a critical role in early steps of HIV replication. Due to its exposure in the cytoplasm after entry, HIV capsid is a target for host cell factors that act directly to block infection such as TRIM5α and MxB. Several host proteins also play a role in facilitating infection, including in the protection of HIV-1 capsid from recognition by host cell restriction factors. Through an unbiased screening approach, called HIV-CRISPR, we show that the CPSF6-binding deficient, N74D HIV-1 capsid mutant is sensitive to restriction mediated by human TRIM34, a close paralog of the well-characterized HIV restriction factor TRIM5α. This restriction occurs at the step of reverse transcription, is independent of interferon stimulation, and limits HIV-1 infection in key target cells of HIV infection including CD4+ T cells and monocyte-derived dendritic cells. TRIM34 can also restrict some SIV capsids. TRIM34 restriction requires TRIM5α as knockout or knockdown of TRIM5α results in a loss of antiviral activity. Through immunofluorescence studies, we show that TRIM34 and TRIM5α colocalize to cytoplasmic bodies and are more frequently observed to be associated with infecting N74D capsids than with WT HIV-1 capsids. Our results identify TRIM34 as an HIV-1 CA-targeting restriction factor and highlight the potential role for heteromultimeric TRIM interactions in contributing to restriction of HIV-1 infection in human cells.


Assuntos
Proteínas do Capsídeo/metabolismo , Proteínas de Transporte/metabolismo , Infecções por HIV/metabolismo , HIV-1/fisiologia , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Capsídeo/metabolismo , Proteínas do Capsídeo/genética , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Células HEK293 , Infecções por HIV/virologia , Soropositividade para HIV , HIV-1/metabolismo , Células HeLa , Humanos , Transcrição Reversa , Integração Viral/fisiologia
15.
Proc Natl Acad Sci U S A ; 117(19): 10286-10293, 2020 05 12.
Artigo em Inglês | MEDLINE | ID: mdl-32341150

RESUMO

HIV-1 maturation involves conversion of the immature Gag polyprotein lattice, which lines the inner surface of the viral membrane, to the mature capsid protein (CA) lattice, which encloses the viral RNA. Maturation inhibitors such as bevirimat (BVM) bind within six-helix bundles, formed by a segment that spans the junction between the CA and spacer peptide 1 (SP1) subunits of Gag, and interfere with cleavage between CA and SP1 catalyzed by the HIV-1 protease (PR). We report solid-state NMR (ssNMR) measurements on spherical virus-like particles (VLPs), facilitated by segmental isotopic labeling, that provide information about effects of BVM on the structure and dynamics of CA-SP1 junction helices in the immature lattice. Although BVM strongly blocks PR-catalyzed CA-SP1 cleavage in VLPs and blocks conversion of VLPs to tubular CA assemblies, 15N and 13C ssNMR chemical shifts of segmentally labeled VLPs with and without BVM are very similar, indicating that interaction with BVM does not alter the six-helix bundle structure appreciably. Only the 15N chemical shift of A280 (the first residue of SP1) changes significantly, consistent with BVM binding to an internal ring of hydrophobic side chains of L279 residues. Measurements of transverse 15N spin relaxation rates reveal a reduction in the amplitudes and/or timescales of backbone N-H bond motions, corresponding to a rigidification of the six-helix bundles. Overall, our data show that inhibition of HIV-1 maturation by BVM involves changes in structure and dynamics that are surprisingly subtle, but still sufficient to produce a large effect on CA-SP1 cleavage.


Assuntos
Proteínas do Capsídeo/química , HIV-1/efeitos dos fármacos , Fragmentos de Peptídeos/química , Succinatos/farmacologia , Triterpenos/farmacologia , Vírion/efeitos dos fármacos , Replicação Viral , Produtos do Gene gag do Vírus da Imunodeficiência Humana/metabolismo , Fármacos Anti-HIV/farmacologia , Proteínas do Capsídeo/genética , Proteínas do Capsídeo/metabolismo , Farmacorresistência Viral , Infecções por HIV/tratamento farmacológico , Infecções por HIV/metabolismo , Infecções por HIV/virologia , HIV-1/genética , HIV-1/metabolismo , Humanos , Modelos Moleculares , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/metabolismo , Conformação Proteica , Vírion/genética , Vírion/metabolismo , Montagem de Vírus , Produtos do Gene gag do Vírus da Imunodeficiência Humana/genética
16.
Nat Commun ; 11(1): 1767, 2020 04 14.
Artigo em Inglês | MEDLINE | ID: mdl-32286302

RESUMO

Female children and adults typically generate more efficacious immune responses to vaccines and infections than age-matched males, but also suffer greater immunopathology and autoimmune disease. We here describe, in a cohort of > 170 in utero HIV-infected infants from KwaZulu-Natal, South Africa, fetal immune sex differences resulting in a 1.5-2-fold increased female susceptibility to intrauterine HIV infection. Viruses transmitted to females have lower replicative capacity (p = 0.0005) and are more type I interferon-resistant (p = 0.007) than those transmitted to males. Cord blood cells from females of HIV-uninfected sex-discordant twins are more activated (p = 0.01) and more susceptible to HIV infection in vitro (p = 0.03). Sex differences in outcome include superior maintenance of aviraemia among males (p = 0.007) that is not explained by differential antiretroviral therapy adherence. These data demonstrate sex-specific innate immune selection of HIV associated with increased female susceptibility to in utero infection and enhanced functional cure potential among infected males.


Assuntos
Infecções por HIV/imunologia , HIV-1/imunologia , HIV-1/patogenicidade , Imunidade Inata/fisiologia , Antirretrovirais/uso terapêutico , Estudos de Coortes , Feminino , Infecções por HIV/tratamento farmacológico , Infecções por HIV/metabolismo , HIV-1/efeitos dos fármacos , Humanos , Imunidade Inata/genética , Transmissão Vertical de Doença Infecciosa , Interferons/metabolismo , Estimativa de Kaplan-Meier , Masculino , Filogenia , Fatores Sexuais , Pesquisa Médica Translacional
17.
Sex Transm Infect ; 96(5): 337-341, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32245779

RESUMO

OBJECTIVE: To provide insight on viral kinetics and genetic diversity of HIV in seminal plasma at baseline and 1 month after initiating antiretroviral therapy (ART). PATIENTS AND METHODS: Blood and seminal samples from patients with newly diagnosed HIV were obtained before ART initiation (T0) and 1 month after ART initiation (T1). HIV env genetic diversity was studied using deep sequencing Nextera and V3 chemistry in a MiSeq Illumina platform. The number of viral quasispecies (5% cut-off) and Shannon Index were used to analyse diversity. RESULTS: Forty-seven ART-naive patients were recruited between September 2016 and November 2018. At enrolment, the number of quasispecies in blood (median 4 (IQR 2-5)) was lower than in the seminal compartment (median 6, (IQR 4-8)) (p<0.01); the Shannon Index was also higher (p<0.001) in the seminal compartment than in blood (1.77 vs 0.64). At T1, for the 13 patients with detectable HIV in both blood/seminal plasma, viral diversity remained higher (p=0.139) in seminal plasma (median 2 (IQR 1-4.5)) than in blood (median 1 (IQR 1-1.5)) Integrase inhibitors (INI)-based regimens achieved higher levels of undetectability and led more frequently to lower variability (p<0.001) than protease inhibitors (PI) or non-nucleoside reverse transcriptase inhibitors (NNRTI). CONCLUSION: We provide here further evidence of a larger genetic diversity in seminal plasma, both at diagnosis and short term after ART initiation. Our results strengthen previous findings on HIV diversity in seminal plasma. In addition, INIs decrease variability more rapidly than PI and NNRTI in both blood and seminal plasma.


Assuntos
Antirretrovirais/uso terapêutico , Sangue/virologia , Variação Genética , Infecções por HIV/tratamento farmacológico , HIV/genética , Sêmen/virologia , Produtos do Gene env do Vírus da Imunodeficiência Humana/genética , Adulto , Infecções por HIV/sangue , Infecções por HIV/metabolismo , Inibidores de Integrase de HIV/uso terapêutico , Inibidores da Protease de HIV/uso terapêutico , Humanos , Masculino , Inibidores da Transcriptase Reversa/uso terapêutico
18.
PLoS One ; 15(3): e0230707, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32226033

RESUMO

People living with HIV who are on antiretroviral treatment are at increased risk of developing premature cardiovascular disease. Children with perinatal HIV infection (PHIV) have survived through their adolescence and are entering adulthood. We determined the prevalence of metabolic syndrome, abnormal biochemical markers, and characterized body composition parameters in youth living with perinatal HIV infection. This cross-sectional study was conducted at the Research Institute for Health Sciences, Chiang Mai University, Chiang Mai, Thailand from December 2017 to February 2018. PHIV-youths between 15 <25 years of age who were receiving ART were enrolled. Data collection included ART-related history, blood pressure, and anthropometric measurements. Body composition including android, gynoid fat mass, and total body fat were measured by dual-energy X-ray absorptiometry. Fasting blood was drawn to test for lipid profile, glucose, and high sensitivity c-reactive protein (hsCRP). One hundred and twenty PHIV-youths (48% female) were enrolled. Their mean age and the median duration on ART were 20.3 (SD2.6) and 14.1 (IQR 10.4-14.9) years, respectively; 76 (63%) were on first-line non-nucleoside reverse transcriptase inhibitors-based regimens. Thirty-three (28%), 74 (62%), and 13 (11%) of PHIV-youths were underweight (BMI < 18.5 kg/m2), normal (BMI 18.5-24.9 kg/m2), and overweight (BMI ≥ 25.0 kg/m2), respectively. The prevalence of metabolic syndrome was 10.6% (95%CI 5.0-16.0). Seventy-six of 113 (67.3%) of PHIV-youths had lipid alteration; the most prevalent types being low HDL (46.9%) and increased triglycerides (27.4%). Overall 43 (35.9%) had increased hsCRP (16.7% with immediate and 19.2% with high risk for CVD). Females had significantly higher percentage of android and gynoid fat, but lower Android to gynoid ratio (AGR) compared to males. There were 77%, 31%, and 21% of PHIV-youths in the overweight, normal weight, and underweight group with AGR in tertile 3, respectively. In conclusion, we documented presence of metabolic syndrome in 10.6% of PHIV-youths on ART. Increase AGR representing abdominal obesity was detected even in youths with normal BMI or underweight.


Assuntos
Fármacos Anti-HIV/uso terapêutico , Composição Corporal , Infecções por HIV/tratamento farmacológico , Infecções por HIV/metabolismo , Síndrome Metabólica/complicações , Adolescente , Adulto , Biomarcadores/metabolismo , Índice de Massa Corporal , Estudos Transversais , Feminino , Infecções por HIV/complicações , Humanos , Masculino , Adulto Jovem
19.
J Virol ; 94(11)2020 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-32188727

RESUMO

Brain-resident microglia and myeloid cells (perivascular macrophages) are important HIV reservoirs in vivo, especially in the central nervous system (CNS). Despite antiretroviral therapy (ART), low-level persistent HIV replication in these reservoirs remains detectable, which contributes to neuroinflammation and neurological disorders in HIV-infected patients. New approaches complementary to ART to repress residual HIV replication in CNS reservoirs are needed. Our group has recently identified a BRD4-selective small molecule modulator (ZL0580) that induces the epigenetic suppression of HIV. Here, we examined the effects of this compound on HIV in human myeloid cells. We found that ZL0580 induces potent and durable suppression of both induced and basal HIV transcription in microglial cells (HC69) and monocytic cell lines (U1 and OM10.1). Pretreatment of microglia with ZL0580 renders them more refractory to latent HIV reactivation, indicating an epigenetic reprogramming effect of ZL0580 on HIV long terminal repeat (LTR) in microglia. We also demonstrate that ZL0580 induces repressive effect on HIV in human primary monocyte-derived macrophages (MDMs) by promoting HIV suppression during ART treatment. Mechanistically, ZL0580 inhibits Tat transactivation in microglia by disrupting binding of Tat to CDK9, a process key to HIV transcription elongation. High-resolution micrococcal nuclease mapping showed that ZL0580 induces a repressive chromatin structure at the HIV LTR. Taken together, our data suggest that ZL0580 represents a potential approach that could be used in combination with ART to suppress residual HIV replication and/or latent HIV reactivation in CNS reservoirs, thereby reducing HIV-associated neuroinflammation.IMPORTANCE Brain-resident microglia and perivascular macrophages are important HIV reservoirs in the CNS. Persistent viral replication and latent HIV reactivation in the CNS, even under ART, are believed to occur, causing neuroinflammation and neurological disorders in HIV-infected patients. It is critical to identify new approaches that can control residual HIV replication and/or latent HIV reactivation in these reservoirs. We here report that the BRD4-selective small molecule modulator, ZL0580, induces potent and durable suppression of HIV in human microglial and monocytic cell lines. Using an in vitro HIV-infected, ART-treated MDM model, we show that ZL0580 also induces suppressive effect on HIV in human primary macrophages. The significance of our research is that it suggests a potential new approach that has utility in combination with ART to suppress residual HIV replication and/or HIV reactivation in CNS reservoirs, thereby reducing neuroinflammation and neurological disorders in HIV-infected individuals.


Assuntos
Antirreumáticos/farmacologia , Proteínas de Ciclo Celular/antagonistas & inibidores , Epigênese Genética/efeitos dos fármacos , Regulação Viral da Expressão Gênica/efeitos dos fármacos , Infecções por HIV/metabolismo , HIV-1/fisiologia , Microglia , Monócitos , Fatores de Transcrição/antagonistas & inibidores , Replicação Viral/efeitos dos fármacos , Antirreumáticos/química , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular , Infecções por HIV/tratamento farmacológico , Infecções por HIV/patologia , Humanos , Microglia/metabolismo , Microglia/patologia , Microglia/virologia , Monócitos/metabolismo , Monócitos/patologia , Monócitos/virologia , Fatores de Transcrição/metabolismo
20.
Pregnancy Hypertens ; 20: 69-74, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32193148

RESUMO

OBJECTIVES: To immuno-localize histone H2A expression as a marker of neutrophil extracellular traps (NETs) in the placenta; and to quantify and compare the percentage H2A immune-expression as a marker of NETs in the placental intervillous space according to: pregnancy type, HIV status and across the study population. STUDY DESIGN: The participants to the study were a pregnant South African population group of African ancestry (n = 60) stratified as normotensive (N) (n = 30) or pre-eclamptic (PE) (n = 30) and further subdivided as HIV infected (HIV+) (n = 15) or HIV naïve (HIV-) (n = 15). Following informed consent placental tissue samples were obtained at the time of delivery. Immunohistochemistry using the anti-histone 2A (H2A) antibody as a biomarker of NETs, and morphometric image analysis was used to immuno-localize and quantify placental H2A immuno-expression respectively in the placental inter-villous space. Statistical analysis was performed using Graph Pad Prism software (Version 5). MAIN OUTCOME MEASURES: To determine if HIV neutralizes the elevated NETs in PE. RESULTS: NETs were localized within the inter-villous space surrounding the exchange villi and conducting villi of placental tissue. Based on HIV status, a significant elevation in H2A immuno-expression was observed in the HIV+ compared to the HIV- group (p = 0.0008) and in the pre-eclampsia HIV- compared to the normotensive HIV- group (p = 0.0008). However, a significant decline in H2A immuno-expression was observed in the PEHIV+ group compared to the NHIV+ group (p = 0.0072). CONCLUSIONS: Both PE and HIV elevate placental NETs; however, they synergistically downregulate NETs expression. Further investigations are required to interrogate the signaling pathways involved to establish potential NET-targeted therapeutic actions.


Assuntos
Armadilhas Extracelulares/química , Infecções por HIV/metabolismo , Placenta/química , Pré-Eclâmpsia/metabolismo , Complicações Infecciosas na Gravidez/metabolismo , Adolescente , Adulto , Biomarcadores/análise , Vilosidades Coriônicas/química , Feminino , Infecções por HIV/diagnóstico , Infecções por HIV/fisiopatologia , Histonas/análise , Humanos , Pré-Eclâmpsia/diagnóstico , Pré-Eclâmpsia/fisiopatologia , Gravidez , Complicações Infecciosas na Gravidez/diagnóstico , Complicações Infecciosas na Gravidez/fisiopatologia , África do Sul , Adulto Jovem
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