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1.
PLoS Pathog ; 20(8): e1012446, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-39116185

RESUMO

HIV persistence in the brain is a barrier to cure, and potentially contributes to HIV-associated neurocognitive disorders. Whether HIV transcription persists in the brain despite viral suppression with antiretroviral therapy (ART) and is subject to the same blocks to transcription seen in other tissues and blood, is unclear. Here, we quantified the level of HIV transcripts in frontal cortex tissue from virally suppressed or non-virally suppressed people with HIV (PWH). HIV transcriptional profiling of frontal cortex brain tissue (and PBMCs where available) from virally suppressed (n = 11) and non-virally suppressed PWH (n = 13) was performed using digital polymerase chain reaction assays (dPCR). CD68+ myeloid cells or CD3+ T cells expressing HIV p24 protein present in frontal cortex tissue was detected using multiplex immunofluorescence imaging. Frontal cortex brain tissue from PWH had HIV TAR (n = 23/24) and Long-LTR (n = 20/24) transcripts. Completion of HIV transcription was evident in brain tissue from 12/13 non-virally suppressed PWH and from 5/11 virally suppressed PWH, with HIV p24+CD68+ cells detected in these individuals. While a block to proximal elongation was present in frontal cortex tissue from both PWH groups, this block was more extensive in virally suppressed PWH. These findings suggest that the brain is a transcriptionally active HIV reservoir in a subset of virally suppressed PWH.


Assuntos
Encéfalo , Infecções por HIV , HIV-1 , Humanos , Infecções por HIV/metabolismo , Infecções por HIV/virologia , Masculino , Encéfalo/metabolismo , Encéfalo/virologia , Adulto , Pessoa de Meia-Idade , Feminino , Transcrição Gênica , Lobo Frontal/metabolismo , Lobo Frontal/virologia
2.
J Immunol ; 212(10): 1553-1563, 2024 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-38558245

RESUMO

HIV is associated with NK cell dysfunction and expansion of adaptive-like NK cells that persist despite antiretroviral therapy (ART). We investigated the timing of NK cell perturbations during acute HIV infection and the impact of early ART initiation. PBMCs and plasma were obtained from people with HIV (PWH; all men who have sex with men; median age, 26.0 y) diagnosed during Fiebig stages I, II, III, or IV/V. Participants initiated ART a median of 3 d after diagnosis, and immunophenotyping was performed at diagnosis and longitudinally after ART. Anti-CMV Abs were assessed by ELISA. Samples from matched HIV-uninfected males were also analyzed. Proportions of adaptive NK cells (A-NKs; defined as Fcε-Receptor-1γ-) were expanded at HIV diagnosis at all Fiebig stages (pooled median 66% versus 25% for controls; p < 0.001) and were not altered by early ART initiation. Abs to CMV immediate early protein were elevated in PWH diagnosed in Fiebig stages III and IV/V (p < 0.03 for both). Proportions of A-NKs defined as either Fcε-Receptor-1γ- or NKG2C+/CD57+ were significantly associated with HIV DNA levels at diagnosis (p = 0.046 and 0.029, respectively) and trended toward an association after 48 wk of ART. Proportions of activated HLA-DR+/CD38+ NK cells remained elevated in PWH despite early ART initiation. NK cell activation and A-NK expansion occur very early after HIV transmission, before T cell activation, and are not altered by ART initiation during acute infection. A-NKs may contribute to HIV control and thus be useful for HIV cure.


Assuntos
Infecções por HIV , Células Matadoras Naturais , Humanos , Infecções por HIV/imunologia , Infecções por HIV/tratamento farmacológico , Células Matadoras Naturais/imunologia , Masculino , Adulto , HIV-1/imunologia , Antirretrovirais/uso terapêutico , Imunidade Adaptativa , Doença Aguda , Adulto Jovem
3.
Ann Neurol ; 94(4): 798-802, 2023 10.
Artigo em Inglês | MEDLINE | ID: mdl-37493435

RESUMO

Here, we provide the first regional analysis of intact and defective HIV reservoirs within the brain. Brain tissue from both viremic and virally suppressed people with HIV (PWH) harbored HIV pol DNA in all regions tested, with lower levels present in basal ganglia and cerebellum relative to frontal white matter. Intact proviruses were primarily found in the frontal white matter but also detected in other brain regions of PWH, demonstrating frontal white matter as a major brain reservoir of intact, potentially replication competent HIV DNA that persists despite antiretroviral therapy. ANN NEUROL 2023;94:798-802.


Assuntos
Infecções por HIV , HIV-1 , Humanos , Provírus/genética , Linfócitos T CD4-Positivos , HIV-1/genética , Carga Viral , Infecções por HIV/tratamento farmacológico , Encéfalo
4.
Ann Neurol ; 92(4): 532-544, 2022 10.
Artigo em Inglês | MEDLINE | ID: mdl-35867351

RESUMO

OBJECTIVE: Human immunodeficiency virus (HIV) persistence in blood and tissue reservoirs, including the brain, is a major barrier to HIV cure and possible cause of comorbid disease. However, the size and replication competent nature of the central nervous system (CNS) reservoir is unclear. Here, we used the intact proviral DNA assay (IPDA) to provide the first quantitative assessment of the intact and defective HIV reservoir in the brain of people with HIV (PWH). METHODS: Total, intact, and defective HIV proviruses were measured in autopsy frontal lobe tissue from viremic (n = 18) or virologically suppressed (n = 12) PWH. Total or intact/defective proviruses were measured by detection of HIV pol or the IPDA, respectively, through use of droplet digital polymerase chain reaction (ddPCR). HIV-seronegative individuals were included as controls (n = 6). RESULTS: Total HIV DNA was present at similar levels in brain tissues from untreated viremic and antiretroviral (ART)-suppressed individuals (median = 22.3 vs 26.2 HIV pol copies/106 cells), reflecting a stable CNS reservoir of HIV that persists despite therapy. Furthermore, 8 of 10 viremic and 6 of 9 virally suppressed PWH also harbored intact proviruses in the CNS (4.63 vs 12.7 intact copies/106 cells). Viral reservoirs in CNS and matched lymphoid tissue were similar in the composition of intact and/or defective proviruses, albeit at lower levels in the brain. Importantly, CNS resident CD68+ myeloid cells in virally suppressed individuals harbored HIV DNA, directly showing the presence of a CNS resident HIV reservoir. INTERPRETATION: Our results demonstrate the first evidence for an intact, potentially replication competent HIV reservoir in the CNS of virally suppressed PWH. ANN NEUROL 2022;92:532-544.


Assuntos
Infecções por HIV , Provírus , Antirretrovirais/uso terapêutico , Encéfalo , Linfócitos T CD4-Positivos , DNA Viral/genética , DNA Viral/uso terapêutico , Infecções por HIV/tratamento farmacológico , Humanos , Provírus/genética , Carga Viral/métodos
5.
Retrovirology ; 17(1): 24, 2020 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-32762760

RESUMO

BACKGROUND: HIV-1 infects a wide range of CD4+ T cells with different phenotypic properties and differing expression levels of entry coreceptors. We sought to determine the viral tropism of subtype C (C-HIV) Envelope (Env) clones for different CD4+ T cell subsets and whether tropism changes during acute to chronic disease progression. HIV-1 envs were amplified from the plasma of five C-HIV infected women from three untreated time points; less than 2 months, 1-year and 3-years post-infection. Pseudoviruses were generated from Env clones, phenotyped for coreceptor usage and CD4+ T cell subset tropism was measured by flow cytometry. RESULTS: A total of 50 C-HIV envs were cloned and screened for functionality in pseudovirus infection assays. Phylogenetic and variable region characteristic analysis demonstrated evolution in envs between time points. We found 45 pseudoviruses were functional and all used CCR5 to mediate entry into NP2/CD4/CCR5 cells. In vitro infection assays showed transitional memory (TM) and effector memory (EM) CD4+ T cells were more frequently infected (median: 46% and 25% of total infected CD4+ T cells respectively) than naïve, stem cell memory, central memory and terminally differentiated cells. This was not due to these subsets contributing a higher proportion of the CD4+ T cell pool, rather these subsets were more susceptible to infection (median: 5.38% EM and 2.15% TM cells infected), consistent with heightened CCR5 expression on EM and TM cells. No inter- or intra-participant changes in CD4+ T cell subset tropism were observed across the three-time points. CONCLUSIONS: CD4+ T cell subsets that express more CCR5 were more susceptible to infection with C-HIV Envs, suggesting that these may be the major cellular targets during the first 3 years of infection. Moreover, we found that viral tropism for different CD4+ T cell subsets in vitro did not change between Envs cloned from acute to chronic disease stages. Finally, central memory, naïve and stem cell memory CD4+ T cell subsets were susceptible to infection, albeit inefficiently by Envs from all time-points, suggesting that direct infection of these cells may help establish the latent reservoir early in infection.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Infecções por HIV/virologia , HIV-1/fisiologia , Subpopulações de Linfócitos T/imunologia , Tropismo Viral , Produtos do Gene env do Vírus da Imunodeficiência Humana/metabolismo , Adulto , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/virologia , Feminino , Variação Genética , Infecções por HIV/imunologia , HIV-1/classificação , HIV-1/genética , Humanos , Memória Imunológica , Estudos Longitudinais , Filogenia , Receptores de HIV/metabolismo , Subpopulações de Linfócitos T/metabolismo , Subpopulações de Linfócitos T/virologia , Produtos do Gene env do Vírus da Imunodeficiência Humana/genética
6.
J Immunol ; 194(10): 4688-97, 2015 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-25855354

RESUMO

We previously demonstrated that NK cells from HIV-infected individuals have elevated expression of activation markers, spontaneously degranulate ex vivo, and decrease expression of a signal-transducing protein for NK-activating receptors, FcRγ. Importantly, these changes were maintained in virologically suppressed (VS) individuals receiving combination antiretroviral therapy (cART). In this study, we show that loss of FcRγ is caused by the expansion of a novel subset of FcRγ(-)CD56(dim) NK cells with an altered activation receptor repertoire and biological properties. In a cross-sectional study, FcRγ(-) NK cells as a proportion of total CD56(dim) NK cells increased in cART-naive viremic HIV-infected individuals (median [interquartile range] = 25.9 [12.6-56.1] compared with 3.80 [1.15-11.5] for HIV(-) controls, p < 0.0001) and in VS HIV-infected individuals (22.7 [13.1-56.2] compared with 3.80 [1.15-11.5], p = 0.0004), with no difference between cART-naive and VS patients (p = 0.93). FcRγ(-) NK cells expressed no NKp30 or NKp46. They showed greater Ab-dependent cellular cytotoxicity activity against rituximab-opsonized Raji cells and in a whole-blood assay measuring NK responses to overlapping HIV peptides, despite having reduced CD16 expression compared with conventional NK cells. Their prevalence correlated with CMV Ab titers in HIV(-) subjects but not in HIV(+) individuals, and with the inflammatory marker CXCL10 in both groups. The expansion of a subset of NK cells that lacks NKp30 and NKp46 to ∼90% of CD56(dim) NK cells in some VS HIV(+) individuals may influence NK-mediated immunosurveillance in patients receiving cART.


Assuntos
Infecções por HIV/imunologia , Células Matadoras Naturais/imunologia , Subpopulações de Linfócitos/imunologia , Receptores de IgG/imunologia , Antirretrovirais/uso terapêutico , Doença Crônica , Estudos Transversais , Ensaio de Imunoadsorção Enzimática , Infecções por HIV/tratamento farmacológico , Humanos
7.
Immunol Rev ; 254(1): 114-42, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23772618

RESUMO

Despite significant reductions in morbidity and mortality secondary to availability of effective combination anti-retroviral therapy (cART), human immunodeficiency virus (HIV) infection still accounts for 1.5 million deaths annually. The majority of deaths occur in sub-Saharan Africa where rates of opportunistic co-infections are disproportionately high. In this review, we discuss the immunopathogenesis of five common infections that cause significant morbidity in HIV-infected patients globally. These include co-infection with Mycobacterium tuberculosis, Cryptococcus neoformans, hepatitis B virus, hepatitis C virus, and Plasmodium falciparum. Specifically, we review the natural history of each co-infection in the setting of HIV, the specific immune defects induced by HIV, the effects of cART on the immune response to the co-infection, the pathogenesis of immune restoration disease (IRD) associated with each infection, and advances in the areas of prevention of each co-infection via vaccination. Finally, we discuss the opportunities and gaps in knowledge for future research.


Assuntos
Infecções Oportunistas Relacionadas com a AIDS/imunologia , Infecções por HIV/imunologia , Infecções Oportunistas Relacionadas com a AIDS/prevenção & controle , África Subsaariana , Animais , Criptococose/imunologia , Criptococose/prevenção & controle , Infecções por HIV/tratamento farmacológico , Hepatite B/imunologia , Hepatite B/prevenção & controle , Hepatite C/imunologia , Hepatite C/prevenção & controle , Humanos , Malária/imunologia , Malária/prevenção & controle , Tuberculose/imunologia , Tuberculose/prevenção & controle
8.
Retrovirology ; 13(1): 49, 2016 07 26.
Artigo em Inglês | MEDLINE | ID: mdl-27459960

RESUMO

BACKGROUND: Eradication of HIV cannot be achieved with combination antiretroviral therapy (cART) because of the persistence of long-lived latently infected resting memory CD4(+) T cells. We previously reported that HIV latency could be established in resting CD4(+) T cells in the presence of the chemokine CCL19. To define how CCL19 facilitated the establishment of latent HIV infection, the role of chemokine receptor signalling was explored. RESULTS: In resting CD4(+) T cells, CCL19 induced phosphorylation of RAC-alpha serine/threonine-protein kinase (Akt), nuclear factor kappa B (NF-κB), extracellular-signal-regulated kinase (ERK) and p38. Inhibition of the phosphoinositol-3-kinase (PI3K) and Ras/Raf/Mitogen-activated protein kinase/ERK kinase (MEK)/ERK signalling pathways inhibited HIV integration, without significant reduction in HIV nuclear entry (measured by Alu-LTR and 2-LTR circle qPCR respectively). Inhibiting activation of MEK1/ERK1/2, c-Jun N-terminal kinase (JNK), activating protein-1 (AP-1) and NF-κB, but not p38, also inhibited HIV integration. We also show that HIV integrases interact with Pin1 in CCL19-treated CD4(+) T cells and inhibition of JNK markedly reduced this interaction, suggesting that CCL19 treatment provided sufficient signals to protect HIV integrase from degradation via the proteasome pathway. Infection of CCL19-treated resting CD4(+) T cells with mutant strains of HIV, lacking NF-κB binding sites in the HIV long terminal repeat (LTR) compared to infection with wild type virus, led to a significant reduction in integration by up to 40-fold (range 1-115.4, p = 0.03). This was in contrast to only a modest reduction of 5-fold (range 1.7-11, p > 0.05) in fully activated CD4(+) T cells infected with the same mutants. Finally, we demonstrated significant differences in integration sites following HIV infection of unactivated, CCL19-treated, and fully activated CD4(+) T cells. CONCLUSIONS: HIV integration in CCL19-treated resting CD4(+) T cells depends on NF-κB signalling and increases the stability of HIV integrase, which allow subsequent integration and establishment of latency. These findings have implications for strategies needed to prevent the establishment, and potentially reverse, latent infection.


Assuntos
Linfócitos T CD4-Positivos/virologia , Quimiocina CCL19/farmacologia , NF-kappa B/metabolismo , Receptores CCR/genética , Integração Viral , Latência Viral , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/fisiologia , Regulação Viral da Expressão Gênica/efeitos dos fármacos , Integrase de HIV/genética , HIV-1/enzimologia , HIV-1/fisiologia , Humanos , NF-kappa B/genética , Receptores CCR/metabolismo , Transdução de Sinais/efeitos dos fármacos , Integração Viral/efeitos dos fármacos , Latência Viral/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos
9.
J Immunol ; 193(11): 5595-603, 2014 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-25367121

RESUMO

Monocyte activation during HIV-1 infection is associated with increased plasma levels of inflammatory markers and increased risk for premature development of age-related diseases. Because activated monocytes primarily use glucose to support cellular metabolism, we hypothesized that chronic monocyte activation during HIV-1 infection induces a hypermetabolic response with increased glucose uptake. To test this hypothesis, we evaluated glucose transporter 1 (Glut1) expression and glucose uptake by monocyte subpopulations in HIV-seropositive (HIV(+)) treatment-naive individuals (n = 17), HIV(+) individuals on combination antiretroviral therapy with viral loads below detection (n = 11), and HIV-seronegative (HIV(-)) individuals (n = 16). Surface expression of Glut1 and cellular uptake of the fluorescent glucose analog 2-(N-(7-nitrobenz-2-oxa-1, 3-diazol-4-yl) amino)-2 deoxyglucose were analyzed by flow cytometry on monocyte subpopulations. Irrespective of treatment status, monocytes from HIV(+) persons had significantly increased surface expression of Glut1 compared with those from HIV(-) controls. Nonclassical (CD14(+)CD16(++)) and intermediate (CD14(++)CD16(+)) monocyte subpopulations showed higher Glut1 expression than did classical (CD14(++)CD16(-)) monocytes. Intermediate monocytes from treatment-naive HIV(+) individuals also showed increased uptake of 2-(N-(7-nitrobenz-2-oxa-1, 3-diazol-4-yl) amino)-2 deoxyglucose compared with those from HIV(-) controls. Our results show that HIV infection is associated with increased glucose metabolism in monocytes and that Glut1 expression by proinflammatory monocytes is a potential marker of inflammation in HIV-infected subjects. However, the possibility exists whereby other Gluts such as Glut3 and Glut4 may also support the influx of glucose into activated and inflammatory monocyte populations.


Assuntos
Transportador de Glucose Tipo 1/metabolismo , Glucose/metabolismo , Infecções por HIV/imunologia , HIV/imunologia , Monócitos/imunologia , Adulto , Idoso , Antirretrovirais/administração & dosagem , Antirretrovirais/efeitos adversos , Austrália , Biomarcadores/metabolismo , Combinação de Medicamentos , Feminino , Glucose/análogos & derivados , Transportador de Glucose Tipo 1/genética , Infecções por HIV/terapia , Soropositividade para HIV , Humanos , Mediadores da Inflamação/metabolismo , Masculino , Pessoa de Meia-Idade , Monócitos/efeitos dos fármacos , Monócitos/virologia , Regulação para Cima
10.
BMC Med ; 13: 154, 2015 Jul 07.
Artigo em Inglês | MEDLINE | ID: mdl-26149666

RESUMO

BACKGROUND: With more than 600,000 deaths from malaria, mainly of children under five years old and caused by infection with Plasmodium falciparum, comes an urgent need for an effective anti-malaria vaccine. Limited details on the mechanisms of protective immunity are a barrier to vaccine development. Antibodies play an important role in immunity to malaria and monocytes are key effectors in antibody-mediated protection by phagocytosing antibody-opsonised infected erythrocytes (IE). Eliciting antibodies that enhance phagocytosis of IE is therefore an important potential component of an effective vaccine, requiring robust assays to determine the ability of elicited antibodies to stimulate this in vivo. The mechanisms by which monocytes ingest IE and the nature of the monocytes which do so are unknown. METHODS: Purified trophozoite-stage P. falciparum IE were stained with ethidium bromide, opsonised with anti-erythrocyte antibodies and incubated with fresh whole blood. Phagocytosis of IE and TNF production by individual monocyte subsets was measured by flow cytometry. Ingestion of IE was confirmed by imaging flow cytometry. RESULTS: CD14(hi)CD16+ monocytes phagocytosed antibody-opsonised IE and produced TNF more efficiently than CD14(hi)CD16- and CD14(lo)CD16+ monocytes. Blocking experiments showed that Fcγ receptor IIIa (CD16) but not Fcγ receptor IIa (CD32a) or Fcγ receptor I (CD64) was necessary for phagocytosis. CD14(hi)CD16+ monocytes ingested antibody-opsonised IE when peripheral blood mononuclear cells were reconstituted with autologous serum but not heat-inactivated autologous serum. Antibody-opsonised IE were rapidly opsonised with complement component C3 in serum (t1/2 = 2-3 minutes) and phagocytosis of antibody-opsonised IE was inhibited in a dose-dependent manner by an inhibitor of C3 activation, compstatin. Compared to other monocyte subsets, CD14(hi)CD16+ monocytes expressed the highest levels of complement receptor 4 (CD11c) and activated complement receptor 3 (CD11b) subunits. CONCLUSIONS: We show a special role for CD14(hi)CD16+ monocytes in phagocytosing opsonised P. falciparum IE and production of TNF. While ingestion was mediated by Fcγ receptor IIIa, this receptor was not sufficient to allow phagocytosis; despite opsonisation with antibody, phagocytosis of IE also required complement opsonisation. Assays which measure the ability of vaccines to elicit a protective antibody response to P. falciparum should consider their ability to promote phagocytosis and fix complement.


Assuntos
Anticorpos Antiprotozoários/imunologia , Receptores de Lipopolissacarídeos/imunologia , Malária Falciparum , Monócitos/imunologia , Plasmodium falciparum/imunologia , Receptores de IgG/imunologia , Células Cultivadas , Eritrócitos/parasitologia , Citometria de Fluxo , Humanos , Leucócitos Mononucleares/imunologia , Malária Falciparum/imunologia , Malária Falciparum/prevenção & controle , Fagocitose/imunologia
11.
BMC Med ; 12: 108, 2014 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-24980799

RESUMO

BACKGROUND: An understanding of the mechanisms mediating protective immunity against malaria in humans is currently lacking, but critically important to advance the development of highly efficacious vaccines. Antibodies play a key role in acquired immunity, but the functional basis for their protective effect remains unclear. Furthermore, there is a strong need for immune correlates of protection against malaria to guide vaccine development. METHODS: Using a validated assay to measure opsonic phagocytosis of Plasmodium falciparum merozoites, we investigated the potential role of this functional activity in human immunity against clinical episodes of malaria in two independent cohorts (n = 109 and n = 287) experiencing differing levels of malaria transmission and evaluated its potential as a correlate of protection. RESULTS: Antibodies promoting opsonic phagocytosis of merozoites were cytophilic immunoglobulins (IgG1 and IgG3), induced monocyte activation and production of pro-inflammatory cytokines, and were directed against major merozoite surface proteins (MSPs). Consistent with protective immunity in humans, opsonizing antibodies were acquired with increasing age and malaria exposure, were boosted on re-infection, and levels were related to malaria transmission intensity. Opsonic phagocytosis was strongly associated with a reduced risk of clinical malaria in longitudinal studies in children with current or recent infections. In contrast, antibodies to the merozoite surface in standard immunoassays, or growth-inhibitory antibodies, were not significantly associated with protection. In multivariate analyses including several antibody responses, opsonic phagocytosis remained significantly associated with protection against malaria, highlighting its potential as a correlate of immunity. Furthermore, we demonstrate that human antibodies against MSP2 and MSP3 that are strongly associated with protection in this population are effective in opsonic phagocytosis of merozoites, providing a functional link between these antigen-specific responses and protection for the first time. CONCLUSIONS: Opsonic phagocytosis of merozoites appears to be an important mechanism contributing to protective immunity in humans. The opsonic phagocytosis assay appears to be a strong correlate of protection against malaria, a valuable biomarker of immunity, and provides a much-needed new tool for assessing responses to blood-stage malaria vaccines and measuring immunity in populations.


Assuntos
Vacinas Antimaláricas/imunologia , Malária Falciparum/parasitologia , Merozoítos/imunologia , Plasmodium falciparum/imunologia , Animais , Antígenos de Protozoários/imunologia , Criança , Pré-Escolar , Estudos de Coortes , Feminino , Humanos , Lactente , Estudos Longitudinais , Masculino , Fagocitose
12.
Guang Pu Xue Yu Guang Pu Fen Xi ; 34(3): 704-8, 2014 Mar.
Artigo em Zh | MEDLINE | ID: mdl-25208396

RESUMO

Petrochemical wastewater is of huge quantity released during the production and complicated contaminants of petrochemical wastewater will have immense negative impact on ecology environment. Three-dimensional excitation-emission matrix fluorescence(3D-EEM) was used to investigate the characteristic fluorescence of influent and effluent from each processing unit of Hydrolysis-acidification +A/O+ Contact-oxidation Process in a typical petrochemical wastewater treatment plant . The results showed that there were 4 fluorescence peaks named Peak A, Peak B, Peak D, Peak E in the spectrum chart of influent, they are around lambda(ex/lambda(em) = 220/300, 225/340, 270/300, 275/340 nm, the primary source of fluorescence organic matter(FOM) is industrial wastewater. The fluorescence intensity of each fluorescence peak was decreased, while location was unchanged in the effluent of Hydrolysis-acidification. Peak C appeared from the effluent of anaerobic tank at lambda(ex)/lambda(em) = 250/425 nm, then the fluorescence intensity of Peak C was enhanced in the effluent of aerobic tank. Peak A disappeared from the effluent of secondary sedimentation tank. The spectrum chart of the wastewater had no obvious variation after secondary sedimentation tank. The removal rate of FOM was expressed with the degradation percentage of the fluorescence intensity, the total FOM was reduced by 92.0% after processing, and the removal rate of the FOM fluoresce around Peak A, Peak B, Peak D, Peak E were 100.0%, 91.2%, 80.3%, 92.0% respectively. A volatile I(Peak B)/I(Peak E) value of influent but a relatively stable value of effluent demonstrated that the wastewater treatment plant operated steadily and the process has higher capacity in resistance to shock loading.


Assuntos
Compostos Orgânicos , Eliminação de Resíduos Líquidos , Indústrias Extrativas e de Processamento , Petróleo , Espectrometria de Fluorescência , Águas Residuárias
13.
Malar J ; 11: 343, 2012 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-23046548

RESUMO

BACKGROUND: Antibody opsonization of Plasmodium falciparum-infected erythrocytes (IE) plays a crucial role in anti-malarial immunity by promoting clearance of blood-stage infection by monocytes and macrophages. The effects of phagocytosis of opsonized IE on macrophage pro-inflammatory cytokine responses are poorly understood. METHODS: Phagocytic clearance, cytokine response and intracellular signalling were measured using IFN-γ-primed human monocyte-derived macrophages (MDM) incubated with opsonized and unopsonized trophozoite-stage CS2 IE, a chondroitin sulphate-binding malaria strain. Cytokine secretion was measured by bead array or ELISA, mRNA using quantitative PCR, and activation of NF-κB by Western blot and electrophoretic mobility shift assay. Data were analysed using the Mann-Whitney U test or the Wilcoxon signed rank test as appropriate. RESULTS: Unopsonized CS2 IE were not phagocytosed whereas IE opsonized with pooled patient immune serum (PPS) were (Phagocytic index (PI)=18.4, [SE 0.38] n=3). Unopsonized and opsonized IE induced expression of TNF, IL-1ß and IL-6 mRNA by MDM and activated NF-κB to a similar extent. Unopsonized IE induced secretion of IL-6 (median= 622 pg/ml [IQR=1,250-240], n=9) but no IL-1ß or TNF, whereas PPS-opsonized IE induced secretion of IL-1ß (18.6 pg/mL [34.2-14.4]) and TNF (113 pg/ml [421-17.0]) and increased IL-6 secretion (2,195 pg/ml [4,658-1,095]). Opsonized, but not unopsonized, CS2 IE activated caspase-1 cleavage and enzymatic activity in MDM showing that Fc receptor-mediated phagocytosis activates the inflammasome. MDM attached to IgG-coated surfaces however secreted IL-1ß in response to unopsonized IE, suggesting that internalization of IE is not absolutely required to activate the inflammasome and stimulate IL-1ß secretion. CONCLUSIONS: It is concluded that IL-6 secretion from MDM in response to CS2 IE does not require phagocytosis, whereas secretion of TNF and IL-1ß is dependent on Fcγ receptor-mediated phagocytosis; for IL-1ß, this occurs by activation of the inflammasome. The data presented in this paper show that generating antibody responses to blood-stage malaria parasites is potentially beneficial both in reducing parasitaemia via Fcγ receptor-dependent macrophage phagocytosis and in generating a robust pro-inflammatory response.


Assuntos
Citocinas/metabolismo , Eritrócitos/parasitologia , Inflamassomos/metabolismo , Macrófagos/imunologia , Malária Falciparum/imunologia , Proteínas Opsonizantes/imunologia , Plasmodium falciparum/imunologia , Western Blotting , Ensaio de Imunoadsorção Enzimática , Humanos , Malária Falciparum/parasitologia , Reação em Cadeia da Polimerase em Tempo Real
14.
Materials (Basel) ; 16(1)2022 Dec 29.
Artigo em Inglês | MEDLINE | ID: mdl-36614677

RESUMO

The effect of Y2O3 addition on the microstructure and properties of the laser cladded Al-Si alloy coating on the surface of AZ91D magnesium alloy was investigated in this study. The experimental results showed that the Al-Si + Y2O3 cladding layers contained α-Mg, Mg2Si, Al4MgY and a small amount of Al12Mg17 phases. The coarse dendrites, reticulated eutectic structures and massive phases in the coatings tended to be refined and gradually uniformly distributed with the increased amount of Y2O3. The introduction of Y2O3 into the cladding layer favored the improvement of microhardness and wear resistance due to the grain refinement strengthening and dispersion strengthening. The addition of Y2O3 also promoted the reduction of localized corrosion sites and made the corrosion surface smoother, implying that the corrosion resistance of the Y2O3-modified coatings was better than that of the unmodified cladding layer.

15.
Biochem Biophys Res Commun ; 391(1): 570-4, 2010 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-19932088

RESUMO

The influenza virus RNA polymerase (RdRp) was purified from insect cells (around 0.2mg/l). The RdRp catalyzed all the biochemical reactions of influenza virus transcription and replication in vitro; dinucleotide ApG and globin mRNA-primed transcription, de novo initiation (replication), and polyadenylation. The optimal Mg concentration, pH and temperature were 8mM, 8.0 and 25 degrees C, respectively, which were slightly different from those measured for RdRp of virions. This system is a single-round transcription system. K(m) (microM) were 10.74+/-0.26 (GTP), 33.22+/-3.37 (ATP), 28.93+/-0.48 (CTP) and 22.01+/-1.48 (UTP), and V(max) (fmol nucleotide/pmol RdRp/min) were 2.40+/-0.032 (GTP), 1.95+/-0.17 (ATP), 2.07+/-0.17 (CTP), and 1.52+/-0.38 (UTP), which agreed with high mutation of influenza viruses.


Assuntos
Vírus da Influenza A Subtipo H1N1/enzimologia , Virus da Influenza A Subtipo H5N1/enzimologia , RNA Polimerase Dependente de RNA/química , Proteínas Virais/química , Animais , Células Cultivadas , Humanos , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H1N1/isolamento & purificação , Virus da Influenza A Subtipo H5N1/genética , Virus da Influenza A Subtipo H5N1/isolamento & purificação , Insetos/citologia , Cinética , RNA Polimerase Dependente de RNA/biossíntese , RNA Polimerase Dependente de RNA/isolamento & purificação , Proteínas Virais/biossíntese , Proteínas Virais/isolamento & purificação
17.
Arch Virol ; 154(5): 765-73, 2009.
Artigo em Inglês | MEDLINE | ID: mdl-19340394

RESUMO

Hepatitis C virus (HCV) JFH1 efficiently replicates and produces infectious virus particles in cultured cells. We compared polymerase activity between JFH1 and 1b strains in vitro. The RNA polymerase activity of 1b was 6.4% of that of JFH1. In order to study the mechanism and identify domains responsible for the high polymerase activity of JFH1, we converted the amino acids of 1b RdRp to those of JFH1, and compared their Km, Vmax and template binding activity. Four amino acid mutations in the thumb domain of 1b RdRp, S377R, A450S, E455N and Y561F increased 1b polymerase activity, and their activity was 23.1, 45.8, 28.9, and 36.1% of JFH1, respectively. Vmax and RNA binding activity of JFH1, 1bwt and 1bA450S was JFH1 > 1bA450S > 1b, which indicated both high processivity and slightly higher template binding activity contributed to the high polymerase activity of JFH1.


Assuntos
Hepacivirus/enzimologia , Hepacivirus/genética , RNA Polimerase Dependente de RNA/metabolismo , Proteínas Virais/metabolismo , Substituição de Aminoácidos , Hepacivirus/fisiologia , Concentração de Íons de Hidrogênio , Cinética , Mutação , RNA Viral/genética , RNA Polimerase Dependente de RNA/genética , Temperatura , Proteínas Virais/genética , Replicação Viral
18.
Front Immunol ; 8: 731, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28713370

RESUMO

Innate immune dysfunction persists in HIV+ individuals despite effective combination antiretroviral therapy (cART). We recently demonstrated that an adaptive-like CD56dim NK cell population lacking the signal transducing protein FcRγ is expanded in HIV+ individuals. Here, we analyzed a cohort of HIV+ men who have sex with men (MSM, n = 20) at baseline and following 6, 12, and 24 months of cART and compared them with uninfected MSM (n = 15) to investigate the impact of cART on NK cell dysfunction. Proportions of NK cells expressing markers of early (CD69+) and late (HLA-DR+/CD38+) activation were elevated in cART-naïve HIV+ MSM (p = 0.004 and 0.015, respectively), as were FcRγ- NK cells (p = 0.003). Using latent growth curve modeling, we show that cART did not reduce levels of FcRγ- NK cells (p = 0.115) or activated HLA-DR+/CD38+ NK cells (p = 0.129) but did reduce T cell and monocyte activation (p < 0.001 for all). Proportions of FcRγ- NK cells were not associated with NK cell, T cell, or monocyte activation, suggesting different factors drive CD56dim FcRγ- NK cell expansion and immune activation in HIV+ individuals. While proportions of activated CD69+ NK cells declined significantly on cART (p = 0.003), the rate was significantly slower than the decline of T cell and monocyte activation, indicating a reduced potency of cART against NK cell activation. Our findings indicate that 2 years of suppressive cART have no impact on CD56dim FcRγ- NK cell expansion and that NK cell activation persists after normalization of other immune parameters. This may have implications for the development of malignancies and co-morbidities in HIV+ individuals on cART.

19.
FEBS Lett ; 591(20): 3319-3332, 2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28892135

RESUMO

High glucose transporter 1 (Glut1) surface expression is associated with increased glycolytic activity in activated CD4+ T cells. Phosphatidylinositide 3-kinases (PI3K) activation measured by p-Akt and OX40 is elevated in CD4+Glut1+ T cells from HIV+ subjects. TCR engagement of CD4+Glut1+ T cells from HIV+ subjects demonstrates hyperresponsive PI3K-mammalian target of rapamycin signaling. High basal Glut1 and OX40 on CD4+ T cells from combination antiretroviral therapy (cART)-treated HIV+ patients represent a sufficiently metabolically active state permissive for HIV infection in vitro without external stimuli. The majority of CD4+OX40+ T cells express Glut1, thus OX40 rather than Glut1 itself may facilitate HIV infection. Furthermore, infection of CD4+ T cells is limited by p110γ PI3K inhibition. Modulating glucose metabolism may limit cellular activation and prevent residual HIV replication in 'virologically suppressed' cART-treated HIV+ persons.


Assuntos
Fármacos Anti-HIV/uso terapêutico , Linfócitos T CD4-Positivos/metabolismo , Transportador de Glucose Tipo 1/imunologia , Infecções por HIV/metabolismo , Receptores OX40/imunologia , Adulto , Terapia Antirretroviral de Alta Atividade , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/virologia , Proliferação de Células , Classe Ib de Fosfatidilinositol 3-Quinase/genética , Classe Ib de Fosfatidilinositol 3-Quinase/imunologia , Regulação da Expressão Gênica , Transportador de Glucose Tipo 1/genética , Infecções por HIV/tratamento farmacológico , Infecções por HIV/imunologia , Infecções por HIV/virologia , HIV-1/efeitos dos fármacos , HIV-1/crescimento & desenvolvimento , Humanos , Ativação Linfocitária , Masculino , Inibidores de Fosfoinositídeo-3 Quinase , Cultura Primária de Células , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/imunologia , Receptores OX40/genética , Transdução de Sinais , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/imunologia , Replicação Viral/efeitos dos fármacos
20.
AIDS Res Hum Retroviruses ; 33(12): 1220-1235, 2017 12.
Artigo em Inglês | MEDLINE | ID: mdl-28797170

RESUMO

Maraviroc (MVC) is an allosteric inhibitor of human immunodeficiency virus type 1 (HIV-1) entry, and is the only CCR5 antagonist licensed for use as an anti-HIV-1 therapeutic. It acts by altering the conformation of the CCR5 extracellular loops, rendering CCR5 unrecognizable by the HIV-1 envelope (Env) glycoproteins. This study aimed to understand the mechanisms underlying the development of MVC resistance in HIV-1-infected patients. To do this, we obtained longitudinal plasma samples from eight subjects who experienced treatment failure with phenotypically verified, CCR5-tropic MVC resistance. We then cloned and characterized HIV-1 Envs (n = 77) from plasma of pretreatment (n = 36) and treatment failure (n = 41) samples. Our results showed variation in the magnitude of MVC resistance as measured by reductions in maximal percent inhibition of Env-pseudotyped viruses, which was more pronounced in 293-Affinofile cells compared to other cells with similar levels of CCR5 expression. Amino acid determinants of MVC resistance localized to the V3 Env region and were strain specific. We also observed minimal cross-resistance to other CCR5 antagonists by MVC-resistant strains. We conclude that 293-Affinofile cells are highly sensitive for detecting and measuring MVC resistance through a mechanism that is CCR5-dependent yet independent of CCR5 expression levels. The strain-specific nature of resistance mutations suggests that sequence-based diagnostics and prognostics will need to be more sophisticated than simple position scoring to be useful for managing resistance in subjects taking MVC. Finally, the minimal levels of cross-resistance suggests that recognition of the MVC-modified form of CCR5 does not necessarily lead to recognition of other antagonist-modified forms of CCR5.


Assuntos
Fármacos Anti-HIV/uso terapêutico , Antagonistas dos Receptores CCR5/uso terapêutico , Cicloexanos/uso terapêutico , Farmacorresistência Viral/genética , Proteína gp120 do Envelope de HIV/genética , Infecções por HIV/tratamento farmacológico , Receptores CCR5/efeitos dos fármacos , Triazóis/uso terapêutico , Adulto , Contagem de Linfócito CD4 , Linhagem Celular , Feminino , Células HEK293 , HIV-1/efeitos dos fármacos , HIV-1/genética , Humanos , Masculino , Maraviroc , Pessoa de Meia-Idade , Falha de Tratamento , Internalização do Vírus/efeitos dos fármacos
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