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1.
PLoS Pathog ; 17(1): e1009214, 2021 01.
Article in English | MEDLINE | ID: mdl-33465157

ABSTRACT

The precise role of CD4 T cell turnover in maintaining HIV persistence during antiretroviral therapy (ART) has not yet been well characterized. In resting CD4 T cell subpopulations from 24 HIV-infected ART-suppressed and 6 HIV-uninfected individuals, we directly measured cellular turnover by heavy water labeling, HIV reservoir size by integrated HIV-DNA (intDNA) and cell-associated HIV-RNA (caRNA), and HIV reservoir clonality by proviral integration site sequencing. Compared to HIV-negatives, ART-suppressed individuals had similar fractional replacement rates in all subpopulations, but lower absolute proliferation rates of all subpopulations other than effector memory (TEM) cells, and lower plasma IL-7 levels (p = 0.0004). Median CD4 T cell half-lives decreased with cell differentiation from naïve to TEM cells (3 years to 3 months, p<0.001). TEM had the fastest replacement rates, were most highly enriched for intDNA and caRNA, and contained the most clonal proviral expansion. Clonal proviruses detected in less mature subpopulations were more expanded in TEM, suggesting that they were maintained through cell differentiation. Earlier ART initiation was associated with lower levels of intDNA, caRNA and fractional replacement rates. In conclusion, circulating integrated HIV proviruses appear to be maintained both by slow turnover of immature CD4 subpopulations, and by clonal expansion as well as cell differentiation into effector cells with faster replacement rates.


Subject(s)
Anti-Retroviral Agents/therapeutic use , CD4-Positive T-Lymphocytes/pathology , Cell Differentiation , HIV Infections/virology , HIV-1/immunology , Viral Load , Virus Replication , Adult , CD4-Positive T-Lymphocytes/drug effects , CD4-Positive T-Lymphocytes/virology , Case-Control Studies , DNA, Viral/analysis , HIV Infections/drug therapy , HIV Infections/immunology , HIV Infections/pathology , HIV-1/drug effects , HIV-1/genetics , Humans , Male , Middle Aged
2.
J Infect Dis ; 223(9): 1621-1630, 2021 05 20.
Article in English | MEDLINE | ID: mdl-32915986

ABSTRACT

BACKGROUND: Despite early antiretroviral therapy (ART), ART-suppressed people with human immunodeficiency virus (HIV) (PWH) remain at higher risk for infections and infection-related cancers than the general population. The immunologic pathways that remain abnormal in this setting, potentially contributing to these complications, are unclear. METHODS: ART-suppressed PWH and HIV-negative controls, all cytomegalovirus seropositive and enriched for HIV risk factors, were sampled from an influenza vaccine responsiveness study. PWH were stratified by timing of ART initiation (within 6 months of infection [early ART] vs later) and nadir CD4+ T-cell count among later initiators. Between-group differences in kynurenine-tryptophan (KT) ratio, interferon-inducible protein 10, soluble CD14 and CD163, soluble tumor necrosis factor receptor 2, interleukin 6, and soluble urokinase plasminogen activator receptor were assessed after confounder adjustment. RESULTS: Most participants (92%) were male, reflecting the demographics of early-ART initiators in San Francisco. Most biomarkers were higher among later-ART initiators. Participants in the early-ART group achieved near-normal soluble tumor necrosis factor receptor 2, interleukin 6, and soluble urokinase plasminogen activator receptor levels, but substantially higher KT ratio than those without HIV after confounder adjustment (P = .008). Soluble CD14, soluble CD163, and interferon-inducible protein 10 trended similarly. CONCLUSIONS: While early-ART initiators restore near-normal levels of many inflammatory markers, the kynurenine pathway of tryptophan catabolism remains abnormally high. Because this pathway confers adaptive immune defects and predicts tuberculosis and cancer progression, this it may contribute to persistent risks of these complications in this setting.


Subject(s)
Anti-HIV Agents , Biomarkers/blood , HIV Infections , Immune System , Anti-HIV Agents/therapeutic use , Antigens, CD , Antigens, Differentiation, Myelomonocytic , Chemokine CXCL10 , HIV Infections/drug therapy , HIV Infections/immunology , Humans , Interleukin-6 , Kynurenine , Lipopolysaccharide Receptors , Male , Receptors, Cell Surface , Receptors, Tumor Necrosis Factor, Type II , Receptors, Urokinase Plasminogen Activator , Tryptophan
3.
Nat Biotechnol ; 38(10): 1174-1183, 2020 10.
Article in English | MEDLINE | ID: mdl-32855547

ABSTRACT

Appropriate use and interpretation of serological tests for assessments of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) exposure, infection and potential immunity require accurate data on assay performance. We conducted a head-to-head evaluation of ten point-of-care-style lateral flow assays (LFAs) and two laboratory-based enzyme-linked immunosorbent assays to detect anti-SARS-CoV-2 IgM and IgG antibodies in 5-d time intervals from symptom onset and studied the specificity of each assay in pre-coronavirus disease 2019 specimens. The percent of seropositive individuals increased with time, peaking in the latest time interval tested (>20 d after symptom onset). Test specificity ranged from 84.3% to 100.0% and was predominantly affected by variability in IgM results. LFA specificity could be increased by considering weak bands as negative, but this decreased detection of antibodies (sensitivity) in a subset of SARS-CoV-2 real-time PCR-positive cases. Our results underline the importance of seropositivity threshold determination and reader training for reliable LFA deployment. Although there was no standout serological assay, four tests achieved more than 80% positivity at later time points tested and more than 95% specificity.


Subject(s)
Betacoronavirus , Clinical Laboratory Techniques/methods , Coronavirus Infections/diagnosis , Pneumonia, Viral/diagnosis , Adult , Aged , Aged, 80 and over , Antibodies, Viral/blood , Betacoronavirus/genetics , Betacoronavirus/immunology , Betacoronavirus/isolation & purification , Biotechnology , COVID-19 , COVID-19 Testing , Chromatography, Affinity , Clinical Laboratory Techniques/statistics & numerical data , Coronavirus Infections/epidemiology , Coronavirus Infections/immunology , Enzyme-Linked Immunosorbent Assay , Female , Humans , Immunoglobulin G/blood , Immunoglobulin M/blood , Male , Middle Aged , Pandemics , Pneumonia, Viral/epidemiology , Pneumonia, Viral/immunology , Point-of-Care Testing , Reverse Transcriptase Polymerase Chain Reaction , SARS-CoV-2 , Sensitivity and Specificity , Young Adult
4.
medRxiv ; 2020 May 17.
Article in English | MEDLINE | ID: mdl-32511497

ABSTRACT

BACKGROUND: Serological tests are crucial tools for assessments of SARS-CoV-2 exposure, infection and potential immunity. Their appropriate use and interpretation require accurate assay performance data. METHOD: We conducted an evaluation of 10 lateral flow assays (LFAs) and two ELISAs to detect anti-SARS-CoV-2 antibodies. The specimen set comprised 128 plasma or serum samples from 79 symptomatic SARS-CoV-2 RT-PCR-positive individuals; 108 pre-COVID-19 negative controls; and 52 recent samples from individuals who underwent respiratory viral testing but were not diagnosed with Coronavirus Disease 2019 (COVID-19). Samples were blinded and LFA results were interpreted by two independent readers, using a standardized intensity scoring system. RESULTS: Among specimens from SARS-CoV-2 RT-PCR-positive individuals, the percent seropositive increased with time interval, peaking at 81.8-100.0% in samples taken >20 days after symptom onset. Test specificity ranged from 84.3-100.0% in pre-COVID-19 specimens. Specificity was higher when weak LFA bands were considered negative, but this decreased sensitivity. IgM detection was more variable than IgG, and detection was highest when IgM and IgG results were combined. Agreement between ELISAs and LFAs ranged from 75.7-94.8%. No consistent cross-reactivity was observed. CONCLUSION: Our evaluation showed heterogeneous assay performance. Reader training is key to reliable LFA performance, and can be tailored for survey goals. Informed use of serology will require evaluations covering the full spectrum of SARS-CoV-2 infections, from asymptomatic and mild infection to severe disease, and later convalescence. Well-designed studies to elucidate the mechanisms and serological correlates of protective immunity will be crucial to guide rational clinical and public health policies.

5.
EBioMedicine ; 20: 217-229, 2017 Jun.
Article in English | MEDLINE | ID: mdl-28529033

ABSTRACT

Reactivation of latent viral reservoirs is on the forefront of HIV-1 eradication research. However, it is unknown if latency reversing agents (LRAs) increase the level of viral transcription from cells producing HIV RNA or harboring transcriptionally-inactive (latent) infection. We therefore developed a microfluidic single-cell-in-droplet (scd)PCR assay to directly measure the number of CD4+ T cells that produce unspliced (us)RNA and multiply spliced (ms)RNA following ex vivo latency reversal with either an histone deacetylase inhibitor (romidepsin) or T cell receptor (TCR) stimulation. Detection of HIV-1 transcriptional activity can also be performed on hundreds of thousands of CD4+ T-cells in a single experiment. The scdPCR method was then applied to CD4+ T cells obtained from HIV-1-infected individuals on antiretroviral therapy. Overall, our results suggest that effects of LRAs on HIV-1 reactivation may be heterogeneous-increasing transcription from active cells in some cases and increasing the number of transcriptionally active cells in others. Genomic DNA and human mRNA isolated from HIV-1 reactivated cells could also be detected and quantified from individual cells. As a result, our assay has the potential to provide needed insight into various reservoir eradication strategies.


Subject(s)
HIV Infections/virology , HIV-1/genetics , High-Throughput Screening Assays , Polymerase Chain Reaction , RNA, Viral , Single-Cell Analysis , Virus Latency , Adult , CD4-Positive T-Lymphocytes/virology , Cells, Cultured , HIV Infections/drug therapy , Humans , Middle Aged , Sequence Analysis, DNA , Viral Load , Virus Activation/genetics
6.
J Infect Dis ; 213(4): 569-73, 2016 Feb 15.
Article in English | MEDLINE | ID: mdl-26310308

ABSTRACT

We leveraged data from the Preexposure Prophylaxis Initiative (iPrEx), a global trial of preexposure chemoprophylaxis against human immunodeficiency virus type 1 (HIV-1) infection, to compare T-cell activation between those who remained negative for HIV-1 and those who became infected during the trial. The frequency of CD38(+)HLA-DR(+) CD8(+) T cells was greater in those who seroconverted, relative to the frequency in those who remained uninfected (1.30% vs 0.82%, respectively; P = .005). This translated to an odds ratio of 4.26 (95% confidence interval, 1.54-11.78) for the association between CD8(+) T-cell activation and infection with HIV-1. T-cell activation may be a biomarker for elevated HIV-1 infection risk.


Subject(s)
CD8-Positive T-Lymphocytes/immunology , HIV Infections/pathology , Lymphocyte Activation , T-Lymphocyte Subsets/immunology , ADP-ribosyl Cyclase 1/analysis , Adolescent , Adult , CD8-Positive T-Lymphocytes/chemistry , HIV Infections/virology , HIV-1/immunology , HLA-DR Antigens/analysis , Humans , Male , Membrane Glycoproteins/analysis , Young Adult
7.
Proc Natl Acad Sci U S A ; 112(27): 8379-84, 2015 Jul 07.
Article in English | MEDLINE | ID: mdl-26100867

ABSTRACT

HIV-1-specific T-cell responses in exposed seronegative subjects suggest that a viral breach of the exposure site is more common than current transmission rates would suggest and that host immunity can extinguish subsequent infection foci. The Preexposure Prophylaxis Initiative (iPrEx) chemoprophylaxis trial provided an opportunity to rigorously investigate these responses in a case-control immunology study; 84 preinfection peripheral blood mononuclear cell samples from individuals enrolled in the iPrEx trial who later seroconverted were matched with 480 samples from enrolled subjects who remained seronegative from both the placebo and active treatment arms. T-cell responses to HIV-1 Gag, Protease, Integrase, Reverse Transcriptase, Vif, and Nef antigens were quantified for all subjects in an IFN-γ enzyme-linked immunospot (ELISpot) assay. IFN-γ responses varied in magnitude and frequency across subjects. A positive response was more prevalent in those who remained persistently HIV-1-negative for Gag (P = 0.007), Integrase (P < 0.001), Vif (P < 0.001), and Nef (P < 0.001). When correlated with outcomes in the iPrEx trial, Vif- and Integrase-specific T-cell responses were associated with reduced HIV-1 infection risk [hazard ratio (HR) = 0.36, 95% confidence interval (95% CI) = 0.19-0.66 and HR = 0.52, 95% CI = 0.28-0.96, respectively]. Antigen-specific responses were independent of emtricitabine/tenofovir disoproxil fumarate use. IFN-γ secretion in the ELISpot was confirmed using multiparametric flow cytometry and largely attributed to effector memory CD4+ or CD8+ T cells. Our results show that HIV-1-specific T-cell immunity can be detected in exposed but uninfected individuals and that these T-cell responses can differentiate individuals according to infection outcomes.


Subject(s)
HIV Infections/immunology , HIV-1/immunology , Immunity, Cellular/immunology , Leukocytes, Mononuclear/immunology , Adult , CD4-Positive T-Lymphocytes/immunology , CD4-Positive T-Lymphocytes/metabolism , CD8-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/metabolism , Case-Control Studies , Enzyme-Linked Immunosorbent Assay , Female , Flow Cytometry , HIV Infections/blood , HIV Infections/virology , HIV Seropositivity/immunology , HIV-1/metabolism , HIV-1/physiology , Human Immunodeficiency Virus Proteins/immunology , Human Immunodeficiency Virus Proteins/metabolism , Humans , Interferon-gamma/immunology , Interferon-gamma/metabolism , Leukocytes, Mononuclear/metabolism , Logistic Models , Male , Multivariate Analysis , Young Adult
8.
Bio Protoc ; 5(12)2015 Jun 20.
Article in English | MEDLINE | ID: mdl-27453912

ABSTRACT

Natural killer (NK) cells comprise 5-20% of peripheral blood mononuclear cells (PBMC) in humans. In addition to their fundamental roles in the defense against viral infections and tumor surveillance, NK cells help shape adaptive immune responses through their production of cytokines. NK cells are traditionally identified as CD3neg, CD14neg, CD19neg lymphocytes expressing CD56. Using a combination of markers that includes CD56 and CD7 greatly increases the ability to define the phenotype and function of NK cell subsets. Two key markers of NK cell function are the production of IFNγ and the release of cytotoxic granules measured by the expression of CD107a. Here we describe a method to assess IFNγ and CD107a expression in NK cells following stimulation with target cells or cytokines. This method can be used to assess the general functional capacity of NK cells in peripheral blood mononuclear cells from a wide range of study participants.

9.
Mol Med ; 20: 179-90, 2014 Apr 22.
Article in English | MEDLINE | ID: mdl-24687160

ABSTRACT

Chronic stress has deleterious effects on immune function, which can lead to adverse health outcomes. However, studies investigating the impact of stress reduction interventions on immunity in clinical research have yielded divergent results, potentially stemming from differences in study design and genetic heterogeneity, among other clinical research challenges. To test the hypothesis that reducing glucocorticoid levels enhances certain immune functions, we administered influenza vaccine once (prime) or twice (boost) to mice housed in either standard control caging or environmental enrichment (EE) caging. We have shown that this approach reduces mouse corticosterone production. Compared with controls, EE mice had significantly lower levels of fecal corticosterone metabolites (FCMs) and increased splenic B and T lymphocyte numbers. Corticosterone levels were negatively associated with the numbers of CD19(+) (r(2) = 0.43, p = 0.0017), CD4(+) (r(2) = 0.28, p = 0.0154) and CD8(+) cells (r(2) = 0.20, p = 0.0503). Vaccinated mice showed nonsignificant differences in immunoglobulin G (IgG) titer between caging groups, although EE mice tended to exhibit larger increases in titer from prime to boost than controls; the interaction between the caging group (control versus EE) and vaccine group (prime versus boost) showed a strong statistical trend (cage-group*vaccine-group, F = 4.27, p = 0.0555), suggesting that there may be distinct effects of EE caging on primary versus secondary IgG vaccine responses. Vaccine-stimulated splenocytes from boosted EE mice had a significantly greater frequency of interleukin 5 (IL-5)-secreting cells than boosted controls (mean difference 7.7, IL-5 spot-forming units/10(6) splenocytes, 95% confidence interval 0.24-135.1, p = 0.0493) and showed a greater increase in the frequency of IL-5-secreting cells from prime to boost. Our results suggest that corticosterone reduction via EE caging was associated with enhanced secondary vaccine responses, but had little effect on primary responses in mice. These findings help identify differences in primary and secondary vaccine responses in relationship to stress mediators that may be relevant in clinical studies.


Subject(s)
B-Lymphocytes/immunology , Corticosterone/metabolism , Cytokines/metabolism , Influenza Vaccines/immunology , Spleen/immunology , T-Lymphocytes/immunology , Animals , Environment, Controlled , Immunization, Secondary , Male , Mice , Mice, Inbred BALB C
10.
Retrovirology ; 10: 158, 2013 Dec 18.
Article in English | MEDLINE | ID: mdl-24351015

ABSTRACT

BACKGROUND: A subset of CD3(neg)CD56(neg)CD16⁺ Natural Killer (NK) cells is highly expanded during chronic HIV-1 infection. The role of this subset in HIV-1 pathogenesis remains unclear. The lack of NK cell lineage-specific markers has complicated the study of minor NK cell subpopulations. RESULTS: Using CD7 as an additional NK cell marker, we found that CD3(neg)CD56(neg)CD16⁺ cells are a heterogeneous population comprised of CD7⁺ NK cells and CD7(neg) non-classical myeloid cells. CD7⁺CD56(neg)CD16⁺ NK cells are significantly expanded in HIV-1 infection. CD7⁺CD56(neg)CD16⁺ NK cells are mature and express KIRs, the C-type lectin-like receptors NKG2A and NKG2C, and natural cytotoxicity receptors similar to CD7⁺CD56⁺CD16⁺ NK cells. CD7⁺CD56(neg) NK cells in healthy donors produced minimal IFNγ following K562 target cell or IL-12 plus IL-18 stimulation; however, they degranulated in response to K562 stimulation similar to CD7⁺CD56⁺ NK cells. HIV-1 infection resulted in reduced IFNγ secretion following K562 or cytokine stimulation by both NK cell subsets compared to healthy donors. Decreased granzyme B and perforin expression and increased expression of CD107a in the absence of stimulation, particularly in HIV-1-infected subjects, suggest that CD7⁺CD56(neg)CD16⁺ NK cells may have recently engaged target cells. Furthermore, CD7⁺CD56(neg)CD16⁺ NK cells have significantly increased expression of CD95, a marker of NK cell activation. CONCLUSIONS: Taken together, CD7⁺CD56(neg)CD16⁺ NK cells are activated, mature NK cells that may have recently engaged target cells.


Subject(s)
CD56 Antigen/analysis , HIV Infections/immunology , HIV-1/immunology , Killer Cells, Natural/chemistry , Killer Cells, Natural/immunology , Receptors, IgG/analysis , Antigens, CD7/analysis , GPI-Linked Proteins/analysis , Granzymes/biosynthesis , Healthy Volunteers , Humans , Interferon-gamma/metabolism , Lysosomal-Associated Membrane Protein 1/analysis , Perforin/biosynthesis
11.
PLoS One ; 8(2): e52144, 2013.
Article in English | MEDLINE | ID: mdl-23468834

ABSTRACT

BACKGROUND: The immunopathogenic mechanisms leading to psoriasis remain unresolved. CD57 is a marker of replicative inability and immunosenescence on CD8+ T cells and the proportion of CD57 expressing CD8+ T cells is increased in a number of inflammatory conditions. METHODOLOGY: We examined the expression of CD57 on T cells in the skin of patients affected with psoriasis, comparing lesional and unaffected skin. We also assessed functionality of the T cells by evaluating the secretion of several inflammatory cytokines (IL-17A, IFN-gamma, IL-2, IL-33, TNF-alpha, IL-21, IL-22, and IL-27), from cell-sorted purified CD4+ and CD8+ T cells isolated from lesional and unaffected skin biopsies of psoriasis patients. PRINCIPAL FINDINGS: We observed that the frequency of CD57+CD4+ and CD57+CD8+ T cells was significantly higher in unaffected skin of psoriasis patients compared to lesional skin. Sorted CD4+ T cells from psoriatic lesional skin produced higher levels of IL-17A, IL-22, and IFN-gamma compared to unaffected skin, while sorted CD8+ T cells from lesional skin produced higher levels of IL-17, IL-22, IFN-gamma, TNF-alpha, and IL-2 compared to unaffected skin. CONCLUSIONS/SIGNIFICANCE: These findings suggest that T cells in unaffected skin from psoriasis patients exhibit a phenotype compatible with replicative inability. As they have a lower replicative capacity, CD57+ T cells are less frequent in lesional tissue due to the high cellular turnover.


Subject(s)
CD57 Antigens/metabolism , Cytokines/biosynthesis , Psoriasis/metabolism , Skin/metabolism , Skin/pathology , T-Lymphocytes/metabolism , Adult , CD4-Positive T-Lymphocytes/immunology , CD4-Positive T-Lymphocytes/metabolism , CD8-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/metabolism , Female , Humans , Male , Middle Aged , Psoriasis/immunology , Skin/immunology , T-Lymphocytes/immunology
12.
J Virol ; 87(11): 6073-80, 2013 Jun.
Article in English | MEDLINE | ID: mdl-23536679

ABSTRACT

APOBEC3 proteins mediate potent antiretroviral activity by hypermutating the retroviral genome during reverse transcription. To counteract APOBEC3 and gain a replicative advantage, lentiviruses such as human immunodeficiency virus type 1 (HIV-1) and simian immunodeficiency virus (SIV) have evolved the Vif protein, which targets APOBEC3 proteins for proteasomal degradation. However, the proteasome plays a critical role in the generation of T cell peptide epitopes. Whether Vif-mediated destruction of APOBEC3 proteins leads to the generation and presentation of APOBEC3-derived T cell epitopes on the surfaces of lentivirus-infected cells remains unknown. Here, using peptides derived from multiple Vif-sensitive APOBEC3 proteins, we identified APOBEC3-specific T cell responses in both HIV-1-infected patients and SIV-infected rhesus macaques. These results raise the possibility that these T cell responses may be part of the larger antiretroviral immune response.


Subject(s)
CD8-Positive T-Lymphocytes/virology , Cytidine Deaminase/immunology , Cytosine Deaminase/immunology , HIV Infections/enzymology , HIV-1/physiology , Simian Acquired Immunodeficiency Syndrome/enzymology , Simian Immunodeficiency Virus/physiology , APOBEC-3G Deaminase , Adult , Animals , CD8-Positive T-Lymphocytes/immunology , Cytidine Deaminase/genetics , Cytosine Deaminase/genetics , Female , Gene Products, vif/genetics , Gene Products, vif/immunology , HIV Infections/immunology , HIV Infections/virology , HIV-1/genetics , HIV-1/immunology , Humans , Macaca mulatta , Male , Simian Acquired Immunodeficiency Syndrome/immunology , Simian Acquired Immunodeficiency Syndrome/virology , Simian Immunodeficiency Virus/genetics , Simian Immunodeficiency Virus/immunology
13.
Exp Dermatol ; 22(1): 64-6, 2013 Jan.
Article in English | MEDLINE | ID: mdl-23278897

ABSTRACT

Psoriasis is a hyper-proliferative disease of the skin in which immunological mechanisms play a direct pathogenetic role. There have been limited studies of natural killer (NK) cells in psoriasis. The aim of this study was to examine the phenotype of NK cells in skin biopsies and peripheral blood mononuclear cells from patients with psoriasis and healthy controls. CD56(+) CD16(-) and CD56(+) CD16(+) NK cells were isolated from lesional skin, unaffected skin and PBMC of psoriasis patients, and normal skin and PBMC from healthy controls. The expression of CD57, NKG2A and NKG2C was assessed by flow cytometry. NK cells in psoriasis skin lesions were skewed in their expression of CD57, a marker of NK cell maturity, with CD57 expression significantly reduced and NKG2A expression increased on NK cells in lesional and unaffected skin compared to controls. These data suggest that in this patient cohort, NK cells could be isolated from psoriasis lesions and exhibit an immature phenotype.


Subject(s)
CD57 Antigens/metabolism , Killer Cells, Natural/metabolism , Psoriasis/immunology , Adult , Aged , Aged, 80 and over , CD56 Antigen/metabolism , Cell Differentiation , Humans , Male , Middle Aged , NK Cell Lectin-Like Receptor Subfamily C/metabolism , Phenotype , Psoriasis/blood , Receptors, IgG/metabolism , Skin/cytology , Skin/immunology , Young Adult
14.
PLoS One ; 7(9): e45733, 2012.
Article in English | MEDLINE | ID: mdl-23029209

ABSTRACT

As perinatally HIV-1-infected children grow into adolescents and young adults, they are increasingly burdened with the long-term consequences of chronic HIV-1 infection, with long-term morbidity due to inadequate immunity. In progressive HIV-1 infection in horizontally infected adults, inflammation, T cell activation, and perturbed T cell differentiation lead to an "immune exhaustion", with decline in T cell effector functions. T effector cells develop an increased expression of CD57 and loss of CD28, with an increase in co-inhibitory receptors such as PD-1 and Tim-3. Very little is known about HIV-1 induced T cell dysfunction in vertical infection. In two perinatally antiretroviral drug treated HIV-1-infected groups with median ages of 11.2 yr and 18.5 yr, matched for viral load, we found no difference in the proportion of senescent CD28(-)CD57(+)CD8(+) T cells between the groups. However, the frequency of Tim-3(+)CD8(+) and Tim-3(+)CD4(+) exhausted T cells, but not PD-1(+) T cells, was significantly increased in the adolescents with longer duration of infection compared to the children with shorter duration of HIV-1 infection. PD-1(+)CD8(+) T cells were directly associated with T cell immune activation in children. The frequency of Tim-3(+)CD8(+) T cells positively correlated with HIV-1 plasma viral load in the adolescents but not in the children. These data suggest that Tim-3 upregulation was driven by both HIV-1 viral replication and increased age, whereas PD-1 expression is associated with immune activation. These findings also suggest that the Tim-3 immune exhaustion phenotype rather than PD-1 or senescent cells plays an important role in age-related T cell dysfunction in perinatal HIV-1 infection. Targeting Tim-3 may serve as a novel therapeutic approach to improve immune control of virus replication and mitigate age related T cell exhaustion.


Subject(s)
Aging/metabolism , HIV Infections/metabolism , Infectious Disease Transmission, Vertical , Membrane Proteins/metabolism , T-Lymphocytes/metabolism , Child , HIV Infections/immunology , HIV Infections/transmission , HIV-1 , Hepatitis A Virus Cellular Receptor 2 , Humans , Viral Load
15.
J Virol ; 85(21): 11526-31, 2011 Nov.
Article in English | MEDLINE | ID: mdl-21880743

ABSTRACT

Human endogenous retrovirus (HERV)-specific T cell responses in HIV-1-infected adults have been reported. Whether HERV-specific immunity exists in vertically HIV-1-infected children is unknown. We performed a cross-sectional analysis of HERV-specific T cell responses in 42 vertically HIV-1-infected children. HERV (-H, -K, and -L family)-specific T cell responses were identified in 26 of 42 subjects, with the greatest magnitude observed for the responses to HERV-L. These HERV-specific T cell responses were inversely correlated with the HIV-1 plasma viral load and positively correlated with CD4(+) T cell counts. These data indicate that HERV-specific T cells may participate in controlling HIV-1 replication and that certain highly conserved HERV-derived proteins may serve as promising therapeutic vaccine targets in HIV-1-infected children.


Subject(s)
Endogenous Retroviruses/immunology , HIV Infections/complications , HIV-1/isolation & purification , Infectious Disease Transmission, Vertical , T-Lymphocytes/immunology , CD4 Lymphocyte Count , Cross-Sectional Studies , HIV Infections/transmission , HIV Infections/virology , Humans , Plasma/virology , Viral Load
16.
Proc Natl Acad Sci U S A ; 108(36): 14725-32, 2011 Sep 06.
Article in English | MEDLINE | ID: mdl-21825173

ABSTRACT

During human CMV infection, there is a preferential expansion of natural killer (NK) cells expressing the activating CD94-NKG2C receptor complex, implicating this receptor in the recognition of CMV-infected cells. We hypothesized that NK cells expanded in response to pathogens will be marked by expression of CD57, a carbohydrate antigen expressed on highly mature cells within the CD56(dim)CD16(+) NK cell compartment. Here we demonstrate the preferential expansion of a unique subset of NK cells coexpressing the activating CD94-NKG2C receptor and CD57 in CMV(+) donors. These CD57(+)NKG2C(hi) NK cells degranulated in response to stimulation through their NKG2C receptor. Furthermore, CD57(+)NKG2C(hi) NK cells preferentially lack expression of the inhibitory NKG2A receptor and the inhibitory KIR3DL1 receptor in individuals expressing its HLA-Bw4 ligand. Moreover, in solid-organ transplant recipients with active CMV infection, the percentage of CD57(+)NKG2C(hi) NK cells in the total NK cell population preferentially increased. During acute CMV infection, the NKG2C(+) NK cells proliferated, became NKG2C(hi), and finally acquired CD57. Thus, we propose that CD57 might provide a marker of "memory" NK cells that have been expanded in response to infection.


Subject(s)
CD57 Antigens/immunology , Cell Proliferation , Cytomegalovirus Infections/immunology , Cytomegalovirus/immunology , Killer Cells, Natural/immunology , NK Cell Lectin-Like Receptor Subfamily C/immunology , Acute Disease , Biomarkers/metabolism , CD56 Antigen/immunology , CD56 Antigen/metabolism , CD57 Antigens/biosynthesis , Cell Degranulation/immunology , Cytomegalovirus/metabolism , Cytomegalovirus Infections/metabolism , GPI-Linked Proteins/immunology , GPI-Linked Proteins/metabolism , Gene Expression Regulation/immunology , HLA-B Antigens/immunology , HLA-B Antigens/metabolism , Humans , Immunologic Memory , Killer Cells, Natural/metabolism , NK Cell Lectin-Like Receptor Subfamily C/biosynthesis , Organ Transplantation , Receptors, IgG/immunology , Receptors, IgG/metabolism , Receptors, KIR3DL1/immunology , Receptors, KIR3DL1/metabolism , Tissue Donors , Transplantation, Homologous
17.
PLoS Negl Trop Dis ; 5(4): e1030, 2011 Apr 26.
Article in English | MEDLINE | ID: mdl-21541358

ABSTRACT

The T cell immunoglobulin mucin 3 (Tim-3) receptor is highly expressed on HIV-1-specific T cells, rendering them partially "exhausted" and unable to contribute to the effective immune mediated control of viral replication. To elucidate novel mechanisms contributing to the HTLV-1 neurological complex and its classic neurological presentation called HAM/TSP (HTLV-1 associated myelopathy/tropical spastic paraparesis), we investigated the expression of the Tim-3 receptor on CD8(+) T cells from a cohort of HTLV-1 seropositive asymptomatic and symptomatic patients. Patients diagnosed with HAM/TSP down-regulated Tim-3 expression on both CD8(+) and CD4(+) T cells compared to asymptomatic patients and HTLV-1 seronegative controls. HTLV-1 Tax-specific, HLA-A*02 restricted CD8(+) T cells among HAM/TSP individuals expressed markedly lower levels of Tim-3. We observed Tax expressing cells in both Tim-3(+) and Tim-3(-) fractions. Taken together, these data indicate that there is a systematic downregulation of Tim-3 levels on T cells in HTLV-1 infection, sustaining a profoundly highly active population of potentially pathogenic T cells that may allow for the development of HTLV-1 complications.


Subject(s)
CD8-Positive T-Lymphocytes/immunology , Gene Expression Profiling , Gene Products, tax/immunology , Human T-lymphotropic virus 1/immunology , Membrane Proteins/biosynthesis , Paraparesis, Tropical Spastic/immunology , Paraparesis, Tropical Spastic/pathology , Adult , Aged , Female , Hepatitis A Virus Cellular Receptor 2 , Humans , Male , Middle Aged
18.
J Virol ; 85(14): 6977-85, 2011 Jul.
Article in English | MEDLINE | ID: mdl-21525339

ABSTRACT

Eight percent of the human genome is composed of human endogenous retroviruses (HERVs), which are thought to be inactive remnants of ancient infections. Previously, we showed that individuals with early HIV-1 infection have stronger anti-HERV T cell responses than uninfected controls. In this study, we investigated whether these responses persist in chronic HIV-1 infection and whether they have a role in the control of HIV-1. Peripheral blood mononuclear cells (PBMCs) from 88 subjects diagnosed with HIV-1 infection for at least 1 year (median duration of diagnosis, 13 years) were tested for responses against HERV peptides in gamma interferon (IFN-γ) enzyme immunospot (ELISPOT) assays. Individuals who control HIV-1 viremia without highly active antiretroviral therapy (HAART) had stronger and broader HERV-specific T cell responses than HAART-suppressed patients, virologic noncontrollers, immunologic progressors, and uninfected controls (P < 0.05 for each pairwise comparison). In addition, the magnitude of the anti-HERV T cell response was inversely correlated with HIV-1 viral load (r(2) = 0.197, P = 0.0002) and associated with higher CD4(+) T cell counts (r(2) = 0.072, P = 0.027) in untreated patients. Flow cytometric analyses of an HLA-B51-restricted CD8(+) HERV response in one HIV-1-infected individual revealed a less activated and more differentiated phenotype than that stimulated by a homologous HIV-1 peptide. HLA-B51 tetramer dual staining within this individual confirmed two different T cell populations corresponding to these HERV and HIV-1 epitopes, ruling out cross-reactivity. These findings suggest a possible role for anti-HERV immunity in the control of chronic HIV-1 infection and provide support for a larger effort to design an HIV-1 vaccine that targets conserved antigens such as HERV.


Subject(s)
CD4-Positive T-Lymphocytes/immunology , Endogenous Retroviruses/immunology , HIV Infections/immunology , HIV-1/physiology , Antiretroviral Therapy, Highly Active , Chronic Disease , Enzyme-Linked Immunosorbent Assay , Female , Flow Cytometry , Gene Products, env/immunology , Gene Products, gag/immunology , HIV Infections/drug therapy , HIV Infections/virology , Humans , Immunophenotyping , Male
19.
Blood ; 116(19): 3865-74, 2010 Nov 11.
Article in English | MEDLINE | ID: mdl-20733159

ABSTRACT

Natural killer (NK) cells are innate immune lymphocytes that express a heterogeneous repertoire of germline-encoded receptors and undergo a distinct pattern of maturation. CD57 is a marker of terminal differentiation on human CD8(+) T cells. Very few newborn or fetal NK cells express CD57; however, the frequency of CD57-bearing NK cells increases with age. We assessed the transcriptional, phenotypic, and functional differences between CD57(+) and CD57(-) NK cells within the CD56(dim) mature NK subset. CD57(+) NK cells express a repertoire of NK-cell receptors, suggestive of a more mature phenotype, and proliferate less when stimulated with target cells and/or cytokines. By contrast, a higher frequency of CD57(+) NK cells produced interferon-γ and demonstrated more potent lytic activity when these cells were stimulated through the activating receptor CD16; however, they are less responsive to stimulation by interleukin-12 and interleukin-18. Finally, CD57 expression is induced on CD57(-)CD56(dim) NK cells after activation by interleukin-2. A combination of a mature phenotype, a higher cytotoxic capacity, a higher sensitivity to stimulation via CD16, with a decreased responsiveness to cytokines, and a decreased capacity to proliferate suggest that CD57(+) NK cells are highly mature and might be terminally differentiated.


Subject(s)
CD56 Antigen/metabolism , CD57 Antigens/metabolism , Killer Cells, Natural/classification , Killer Cells, Natural/immunology , Receptors, IgG/metabolism , Adult , Cell Death , Cell Differentiation , Cell Proliferation , Cytotoxicity, Immunologic , GPI-Linked Proteins/metabolism , Gene Expression Profiling , Humans , In Vitro Techniques , Interferon-gamma/biosynthesis , Interleukin-12/pharmacology , Interleukin-18/pharmacology , Killer Cells, Natural/cytology , Lymphocyte Activation
20.
Blood ; 114(23): 4823-31, 2009 Nov 26.
Article in English | MEDLINE | ID: mdl-19805616

ABSTRACT

The lack of natural killer (NK) cell-specific markers, as well as the overlap among several common surface antigens and functional properties, has obscured the delineation between NK cells and dendritic cells. Here, novel subsets of peripheral blood CD3/14/19(neg) NK cells and monocyte/dendritic cell (DC)-like cells were identified on the basis of CD7 and CD4 expression. Coexpression of CD7 and CD56 differentiates NK cells from CD56+ monocyte/DC-like cells, which lack CD7. In contrast to CD7+CD56+ NK cells, CD7(neg)CD56+ cells lack expression of NK cell-associated markers, but share commonalities in their expression of various monocyte/DC-associated markers. Using CD7, we observed approximately 60% of CD4+CD56+ cells were CD7(neg) cells, indicating the actual frequency of activated CD4+ NK cells is much lower in the blood than previously recognized. Functionally, only CD7+ NK cells secrete gamma interferon (IFNgamma) and degranulate after interleukin-12 (IL-12) plus IL-18 or K562 target cell stimulation. Furthermore, using CD7 to separate CD56+ NK cells and CD56+ myeloid cells, we demonstrate that unlike resting CD7+CD56+ NK cells, the CD7(neg)CD56+ myeloid cells stimulate a potent allogeneic response. Our data indicate that CD7 and CD56 coexpression discriminates NK cells from CD7(neg)CD56+ monocyte/DC-like cells, thereby improving our ability to study the intricacies of NK-cell subset phenotypes and functions in vivo.


Subject(s)
Antigens, CD7/analysis , CD4 Antigens/analysis , CD56 Antigen/analysis , Dendritic Cells/classification , Killer Cells, Natural/classification , Monocytes/classification , Adult , Antigen Presentation , Cell Separation , Cytoplasmic Granules/metabolism , Cytotoxicity, Immunologic , Dendritic Cells/chemistry , Dendritic Cells/immunology , Flow Cytometry , Humans , Immunophenotyping , Interferon-gamma/metabolism , Interleukin-12/pharmacology , Interleukin-18/pharmacology , K562 Cells/immunology , Killer Cells, Natural/chemistry , Killer Cells, Natural/drug effects , Killer Cells, Natural/immunology , Killer Cells, Natural/metabolism , Lymphocyte Culture Test, Mixed , Monocytes/chemistry , Monocytes/immunology , Receptors, Chemokine/analysis , Receptors, KIR/analysis
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