JoVE Visualize What is visualize?
Stop Reading. Start Watching.
Advanced Search
Stop Reading. Start Watching.
Regular Search
Find video protocols related to scientific articles indexed in Pubmed.
Preserving HIV-specific T cell responses: does timing of antiretroviral therapy help?
Curr Opin HIV AIDS
PUBLISHED: 11-13-2014
Show Abstract
Hide Abstract
HIV-specific T cell responses are likely to have an important role in HIV cure strategies that aim for long-lasting viral control without antiretroviral therapy (ART). An important issue in enhancing virus-specific T cell responses is whether timing of ART can influence their magnitude and breadth.
Related JoVE Video
Human herpesvirus 8 induces polyfunctional B lymphocytes that drive Kaposi's sarcoma.
MBio
PUBLISHED: 09-02-2014
Show Abstract
Hide Abstract
Kaposi's sarcoma (KS) is an unusual neoplasia wherein the tumor consists primarily of endothelial cells infected with human herpesvirus 8 (HHV-8; Kaposi's sarcoma-associated herpesvirus) that are not fully transformed but are instead driven to excess proliferation by inflammatory and angiogenic factors. This oncogenic process has been postulated but unproven to depend on a paracrine effect of an abnormal excess of host cytokines and chemokines produced by HHV-8-infected B lymphocytes. Using newly developed measures for intracellular detection of lytic cycle proteins and expression of cytokines and chemokines, we show that HHV-8 targets a range of naive B cell, IgM memory B cell, and plasma cell-like populations for infection and induction of interleukin-6, tumor necrosis factor alpha, macrophage inhibitory protein 1?, macrophage inhibitory protein 1?, and interleukin-8 in vitro and in the blood of HHV-8/HIV-1-coinfected subjects with KS. These B cell lineage subsets that support HHV-8 infection are highly polyfunctional, producing combinations of 2 to 5 of these cytokines and chemokines, with greater numbers in the blood of subjects with KS than in those without KS. Our study provides a new paradigm of B cell polyfunctionality and supports a key role for B cell-derived cytokines and chemokines produced during HHV-8 infection in the development of KS.
Related JoVE Video
Dendritic cells restore CD8+ T cell reactivity to autologous HIV-1.
J. Virol.
PUBLISHED: 06-18-2014
Show Abstract
Hide Abstract
Recall T cell responses to HIV-1 antigens are used as a surrogate for endogenous cellular immune responses generated during infection. Current methods of identifying antigen-specific T cell reactivity in HIV-1 infection use bulk peripheral blood mononuclear cells (PBMC) yet ignore professional antigen-presenting cells (APC) that could reveal otherwise hidden responses. In the present study, peptides representing autologous variants of major histocompatibility complex (MHC) class I-restricted epitopes from HIV-1 Gag and Env were used as antigens in gamma interferon (IFN-?) enzyme-linked immunosorbent spot (ELISpot) and polyfunctional cytokine assays. Here we show that dendritic cells (DC) enhanced T cell reactivity at all stages of disease progression but specifically restored T cell reactivity after combination antiretroviral therapy (cART) to early infection levels. Type 1 cytokine secretion was also enhanced by DC and was most apparent late post-cART. We additionally show that DC reveal polyfunctional T cell responses after many years of treatment, when potential immunotherapies would be implemented. These data underscore the potential efficacy of DC immunotherapy that aims to awaken a dormant, autologous, HIV-1-specific CD8+ T cell response.
Related JoVE Video
Alterations in cholesterol metabolism restrict HIV-1 trans infection in nonprogressors.
MBio
PUBLISHED: 05-01-2014
Show Abstract
Hide Abstract
ABSTRACT HIV-1-infected nonprogressors (NP) inhibit disease progression for years without antiretroviral therapy. Defining the mechanisms for this resistance to disease progression could be important in determining strategies for controlling HIV-1 infection. Here we show that two types of professional antigen-presenting cells (APC), i.e., dendritic cells (DC) and B lymphocytes, from NP lacked the ability to mediate HIV-1 trans infection of CD4(+) T cells. In contrast, APC from HIV-1-infected progressors (PR) and HIV-1-seronegative donors (SN) were highly effective in mediating HIV-1 trans infection. Direct cis infection of T cells with HIV-1 was comparably efficient among NP, PR, and SN. Lack of HIV-1 trans infection in NP was linked to lower cholesterol levels and an increase in the levels of the reverse cholesterol transporter ABCA1 (ATP-binding cassette transporter A1) in APC but not in T cells. Moreover, trans infection mediated by APC from NP could be restored by reconstitution of cholesterol and by inhibiting ABCA1 by mRNA interference. Importantly, this appears to be an inherited trait, as it was evident in APC obtained from NP prior to their primary HIV-1 infection. The present study demonstrates a new mechanism wherein enhanced lipid metabolism in APC results in remarkable control of HIV-1 trans infection that directly relates to lack of HIV-1 disease progression. IMPORTANCE HIV-1 can be captured by antigen-presenting cells (APC) such as dendritic cells and transferred to CD4 helper T cells, which results in greatly enhanced viral replication by a mechanism termed trans infection. A small percentage of HIV-1-infected persons are able to control disease progression for many years without antiretroviral therapy. In our study, we linked this lack of disease progression to a profound inability of APC from these individuals to trans infect T cells. This effect was due to altered lipid metabolism in their APC, which appears to be an inherited trait. These results provide a basis for therapeutic interventions to control of HIV-1 infection through modulation of cholesterol metabolism.
Related JoVE Video
Stuck in the middle: longitudinal HIV-related health disparities among men who have sex with men and women.
J. Acquir. Immune Defic. Syndr.
PUBLISHED: 03-26-2014
Show Abstract
Hide Abstract
Men who have sex with men and women (MSMW) have been shown in cross-sectional studies to suffer HIV-related health disparities above and beyond those found among men who have sex with men only (MSMO). We conducted a secondary data analysis over a 7-year time frame of participants in the Multicenter AIDS Cohort Study, a long-standing prospective cohort study, to examine whether MSMW had persistently higher rates of depression symptoms, polydrug use, and (among HIV-positive men who have sex with men) HIV viral load levels compared with MSMO.
Related JoVE Video
Influenza and other respiratory virus infections in outpatients with medically attended acute respiratory infection during the 2011-12 influenza season.
Influenza Other Respir Viruses
PUBLISHED: 02-26-2014
Show Abstract
Hide Abstract
Respiratory tract infections are a major cause of outpatient visits, yet only a portion is tested to determine the etiologic organism. Multiplex reverse transcriptase polymerase chain reaction (MRT-PCR) assays for detection of multiple viruses are being used increasingly in clinical settings.
Related JoVE Video
The impact of viral evolution and frequency of variant epitopes on primary and memory human immunodeficiency virus type 1-specific CD8? T cell responses.
Virology
PUBLISHED: 02-08-2014
Show Abstract
Hide Abstract
It is unclear if HIV-1 variants lose the ability to prime naïve CD8(+) cytotoxic T lymphocytes (CTL) during progressive, untreated infection. We conducted a comprehensive longitudinal analysis of viral evolution and its impact on primary and memory CD8(+) T cell responses pre-seroconversion (SC), post-SC, and during combination antiretroviral therapy (cART). Memory T cell responses targeting autologous virus variants reached a nadir by 8 years post-SC with development of AIDS, followed by a transient enhancement of anti-HIV-1 CTL responses upon initiation of cART. We show broad and high magnitude primary T cell responses to late variants in pre-SC T cells, comparable to primary anti-HIV-1 responses induced in T cells from uninfected persons. Despite evolutionary changes, CD8(+) T cells could still be primed to HIV-1 variants. Hence, vaccination against late, mutated epitopes could be successful in enhancing primary reactivity of T cells for control of the residual reservoir of HIV-1 during cART.
Related JoVE Video
Human herpesvirus 8 glycoprotein B binds the entry receptor DC-SIGN.
Virus Res.
PUBLISHED: 02-03-2014
Show Abstract
Hide Abstract
We have previously shown that human herpesvirus 8 (HHV-8) uses DC-SIGN as an entry receptor for dendritic cells, macrophages and B cells. The viral attachment protein for DC-SIGN is unknown. HHV-8 virions contain five conserved herpesvirus glycoproteins, a single unique glycoprotein, and two predicted glycoproteins. Previous studies have shown that DC-SIGN binds highly mannosylated glycoproteins. The HHV-8 glycoprotein B (gB) has been reported to be highly mannosylated, and therefore we hypothesized that gB will bind to DC-SIGN. In this report we confirm that gB has a high mannose carbohydrate structure and demonstrate for the first time that it binds DC-SIGN in a dose-dependent manner. We also identify key amino acids in the DC-SIGN carbohydrate recognition domain that are required for HHV-8 infection and compare these results with published binding regions for ICAM-2/3 and HIV-1 gp120. These results clarify some of the initial events in HHV-8 entry and can be used for the design of targeted preventive therapies.
Related JoVE Video
Transmission and evolution of hepatitis C virus in HCV seroconverters in HIV infected subjects.
Virology
PUBLISHED: 01-15-2014
Show Abstract
Hide Abstract
HIV/HCV co-infection provides a model to determine the role of immunity on HCV transmission and evolution. In this study HCV transmission and evolution were evaluated in 6 HCV seroconverters in HIV-infected subjects with a wide range of CD4 cell count. The HCV envelope E1/E2 sequences were analyzed for transmission bottleneck, viral diversity/divergence, immune pressure, and mutations of HLA class I/II restricted epitopes. HCV infection started with transmission bottleneck in all HIV-infected individuals. During the 1.0-2.0 years of infection there was a shift of viral quasispecies in majority of the subjects from one to next visit. However, HCV diversity, divergence, mutations in HLA class I/II restricted and virus neutralizing epitopes were similar in all subjects regardless of CD4 cell count at the time of HCV infection. Our results suggest that HCV transmission and evolution in HIV-infected subjects may not be influenced by host CD4 cell count at the time of infection.
Related JoVE Video
Dendritic cells: key players in human herpesvirus 8 infection and pathogenesis.
Front Microbiol
PUBLISHED: 01-01-2014
Show Abstract
Hide Abstract
Human herpesvirus 8 (HHV-8; Kaposi's sarcoma-associated herpesvirus) is an oncogenic gammaherpesvirus that primarily infects cells of the immune and vascular systems. HHV-8 interacts with and targets professional antigen presenting cells and influences their function. Infection alters the maturation, antigen presentation, and immune activation capabilities of certain dendritic cells (DC) despite non-robust lytic replication in these cells. DC sustains a low level of antiviral functionality during HHV-8 infection in vitro. This may explain the ability of healthy individuals to effectively control this virus without disease. Following an immune compromising event, such as organ transplantation or human immunodeficiency virus type 1 infection, a reduced cellular antiviral response against HHV-8 compounded with skewed DC cytokine production and antigen presentation likely contributes to the development of HHV-8 associated diseases, i.e., Kaposi's sarcoma and certain B cell lymphomas. In this review we focus on the role of DC in the establishment of HHV-8 primary and latent infection, the functional state of DC during HHV-8 infection, and the current understanding of the factors influencing virus-DC interactions in the context of HHV-8-associated disease.
Related JoVE Video
Circulating mediators of inflammation and immune activation in AIDS-related non-hodgkin lymphoma.
PLoS ONE
PUBLISHED: 01-01-2014
Show Abstract
Hide Abstract
Non-Hodgkin lymphoma (NHL) is the most common AIDS-related malignancy in developed countries. An elevated risk of developing NHL persists among HIV-infected individuals in comparison to the general population despite the advent of effective antiretroviral therapy. The mechanisms underlying the development of AIDS-related NHL (A-NHL) are not fully understood, but likely involve persistent B-cell activation and inflammation.
Related JoVE Video
Baseline natural killer and T cell populations correlation with virologic outcome after regimen simplification to atazanavir/ritonavir alone (ACTG 5201).
PLoS ONE
PUBLISHED: 01-01-2014
Show Abstract
Hide Abstract
Simplified maintenance therapy with ritonavir-boosted atazanavir (ATV/r) provides an alternative treatment option for HIV-1 infection that spares nucleoside analogs (NRTI) for future use and decreased toxicity. We hypothesized that the level of immune activation (IA) and recovery of lymphocyte populations could influence virologic outcomes after regimen simplification.
Related JoVE Video
Influenza Vaccine Effectiveness in the 2011-2012 Season: Protection Against Each Circulating Virus and the Effect of Prior Vaccination on Estimates.
Clin. Infect. Dis.
PUBLISHED: 11-13-2013
Show Abstract
Hide Abstract
Background.?Each year, the US Influenza Vaccine Effectiveness Network examines the effectiveness of influenza vaccines in preventing medically attended acute respiratory illnesses caused by influenza. Methods.?Patients with acute respiratory illnesses of ?7 days duration were enrolled at ambulatory care facilities in 5 communities. Specimens were collected and tested for influenza by real-time reverse-transcriptase polymerase chain reaction. Receipt of influenza vaccine was defined based on documented evidence of vaccination in medical records or immunization registries. Vaccine effectiveness was estimated in adjusted logistic regression models by comparing the vaccination coverage in those who tested positive for influenza with those who tested negative. Results.?The 2011-2012 season was mild and peaked late, with circulation of both type A viruses and both lineages of type B. Overall adjusted vaccine effectiveness was 47% (95% confidence interval [CI], 36-56) in preventing medically attended influenza; vaccine effectiveness was 65% (95% CI, 44-79) against type A (H1N1) pdm09 but only 39% (95% CI, 23-52) against type A (H3N2). Estimates of vaccine effectiveness against both type B lineages were similar (overall, 58%; 95% CI, 35-73). An apparent negative effect of prior year vaccination on current year effectiveness estimates was noted, particularly for A (H3N2) outcomes. Conclusions.?Vaccine effectiveness in the 2011-2012 season was modest overall, with lower effectiveness against the predominant A (H3N2) virus. This may be related to antigenic drift, but past history of vaccination might also play a role.
Related JoVE Video
Safety, tolerability, and immunogenicity of repeated doses of dermavir, a candidate therapeutic HIV vaccine, in HIV-infected patients receiving combination antiretroviral therapy: results of the ACTG 5176 trial.
J. Acquir. Immune Defic. Syndr.
PUBLISHED: 10-31-2013
Show Abstract
Hide Abstract
HIV-specific cellular immune responses are associated with control of viremia and delayed disease progression. An effective therapeutic vaccine could mimic these effects and reduce the need for continued antiretroviral therapy. DermaVir, a topically administered plasmid DNA-nanomedicine expressing HIV (CladeB) virus-like particles consisting of 15 antigens, induces predominantly central memory T-cell responses.
Related JoVE Video
Identification of conserved and HLA promiscuous DENV3 T-cell epitopes.
PLoS Negl Trop Dis
PUBLISHED: 10-01-2013
Show Abstract
Hide Abstract
Anti-dengue T-cell responses have been implicated in both protection and immunopathology. However, most of the T-cell studies for dengue include few epitopes, with limited knowledge of their inter-serotype variation and the breadth of their human leukocyte antigen (HLA) affinity. In order to expand our knowledge of HLA-restricted dengue epitopes, we screened T-cell responses against 477 overlapping peptides derived from structural and non-structural proteins of the dengue virus serotype 3 (DENV3) by use of HLA class I and II transgenic mice (TgM): A2, A24, B7, DR2, DR3 and DR4. TgM were inoculated with peptides pools and the T-cell immunogenic peptides were identified by ELISPOT. Nine HLA class I and 97 HLA class II novel DENV3 epitopes were identified based on immunogenicity in TgM and their HLA affinity was further confirmed by binding assays analysis. A subset of these epitopes activated memory T-cells from DENV3 immune volunteers and was also capable of priming naïve T-cells, ex vivo, from dengue IgG negative individuals. Analysis of inter- and intra-serotype variation of such an epitope (A02-restricted) allowed us to identify altered peptide ligands not only in DENV3 but also in other DENV serotypes. These studies also characterized the HLA promiscuity of 23 HLA class II epitopes bearing highly conserved sequences, six of which could bind to more than 10 different HLA molecules representing a large percentage of the global population. These epitope data are invaluable to investigate the role of T-cells in dengue immunity/pathogenesis and vaccine design.
Related JoVE Video
Related JoVE Video
Selective induction of CTL helper rather than killer activity by natural epitope variants promotes dendritic cell-mediated HIV-1 dissemination.
J. Immunol.
PUBLISHED: 08-02-2013
Show Abstract
Hide Abstract
The ability of HIV-1 to rapidly accumulate mutations provides the virus with an effective means of escaping CD8(+) CTL responses. In this study, we describe how subtle alterations in CTL epitopes expressed by naturally occurring HIV-1 variants can result in an incomplete escape from CTL recognition, providing the virus with a selective advantage. Rather than paralyzing the CTL response, these epitope modifications selectively induce the CTL to produce proinflammatory cytokines in the absence of target killing. Importantly, instead of dampening the immune response through CTL elimination of variant Ag-expressing immature dendritic cells (DC), a positive CTL-to-DC immune feedback loop dominates whereby the immature DC differentiate into mature proinflammatory DC. Moreover, these CTL-programmed DC exhibit a superior capacity to mediate HIV-1 trans-infection of T cells. This discordant induction of CTL helper activity in the absence of killing most likely contributes to the chronic immune activation associated with HIV-1 infection, and can be used by HIV-1 to promote viral dissemination and persistence. Our findings highlight the need to address the detrimental potential of eliciting dysfunctional cross-reactive memory CTL responses when designing and implementing anti-HIV-1 immunotherapies.
Related JoVE Video
MicroRNA regulation and its effects on cellular transcriptome in human immunodeficiency virus-1 (HIV-1) infected individuals with distinct viral load and CD4 cell counts.
BMC Infect. Dis.
PUBLISHED: 05-16-2013
Show Abstract
Hide Abstract
Disease progression in the absence of therapy varies significantly in HIV-1 infected individuals. Both viral and host cellular molecules are implicated; however, the exact role of these factors and/or the mechanism involved remains elusive. To understand how microRNAs (miRNAs), which are regulators of transcription and translation, influence host cellular gene expression (mRNA) during HIV-1 infection, we performed a comparative miRNA and mRNA microarray analysis using PBMCs obtained from infected individuals with distinct viral load and CD4 counts.
Related JoVE Video
Associations of T cell activation and inflammatory biomarkers with virological response to darunavir/ritonavir plus raltegravir therapy.
J. Antimicrob. Chemother.
PUBLISHED: 04-18-2013
Show Abstract
Hide Abstract
One of the goals of antiretroviral therapy (ART) is to attenuate HIV-induced systemic immune activation and inflammation. We determined the dynamics of biomarkers of immune activation, microbial translocation and inflammation during initial ART with a nucleos(t)ide-sparing regimen of darunavir/ritonavir plus raltegravir. We also evaluated associations between these biomarkers and the virological response to the regimen.
Related JoVE Video
Fifty percent tissue culture infective dose assay for determining the titer of infectious human herpesvirus 8.
J. Clin. Microbiol.
PUBLISHED: 04-03-2013
Show Abstract
Hide Abstract
We have developed a human herpesvirus 8 (HHV-8) 50% tissue culture infective dose (TCID50) assay using the T1H6-DC-SIGN cell line. Infection of T1H6-DC-SIGN cells with HHV-8 induces expression of ?-galactosidase, which was used to determine TCID50 levels. Validation of TCID50 values was performed by immunofluorescence assay of HHV-8 infection of immature dendritic cells at various TCID50 doses.
Related JoVE Video
Incident hepatitis C virus infection in men who have sex with men: a prospective cohort analysis, 1984-2011.
Clin. Infect. Dis.
PUBLISHED: 03-26-2013
Show Abstract
Hide Abstract
Prospective characterization of hepatitis C virus (HCV) transmission in both human immunodeficiency virus (HIV)-infected and -uninfected men who have sex with men (MSM) over the entire HIV epidemic has not been comprehensively conducted.
Related JoVE Video
HIV-1 Trans Infection of CD4(+) T Cells by Professional Antigen Presenting Cells.
Scientifica (Cairo)
PUBLISHED: 02-28-2013
Show Abstract
Hide Abstract
Since the 1990s we have known of the fascinating ability of a complex set of professional antigen presenting cells (APCs; dendritic cells, monocytes/macrophages, and B lymphocytes) to mediate HIV-1 trans infection of CD4(+) T cells. This results in a burst of virus replication in the T cells that is much greater than that resulting from direct, cis infection of either APC or T cells, or trans infection between T cells. Such APC-to-T cell trans infection first involves a complex set of virus subtype, attachment, entry, and replication patterns that have many similarities among APC, as well as distinct differences related to virus receptors, intracellular trafficking, and productive and nonproductive replication pathways. The end result is that HIV-1 can sequester within the APC for several days and be transmitted via membrane extensions intracellularly and extracellularly to T cells across the virologic synapse. Virus replication requires activated T cells that can develop concurrently with the events of virus transmission. Further research is essential to fill the many gaps in our understanding of these trans infection processes and their role in natural HIV-1 infection.
Related JoVE Video
Value of a quality assessment program in optimizing cryopreservation of peripheral blood mononuclear cells in a multicenter study.
Clin. Vaccine Immunol.
PUBLISHED: 02-13-2013
Show Abstract
Hide Abstract
Cryopreservation of peripheral blood mononuclear cells (PBMC) allows assays of cellular function and phenotype to be performed in batches at a later time on PBMC at a central laboratory to minimize assay variability. The Multicenter AIDS Cohort Study (MACS) is an ongoing prospective study of the natural and treated history of human immunodeficiency virus (HIV) infection that stores cryopreserved PBMC from participants two times a year at four study sites. In order to ensure consistent recovery of viable PBMC after cryopreservation, a quality assessment program was implemented and conducted in the MACS over a 6-year period. Every 4 months, recently cryopreserved PBMC from HIV-1-infected and HIV-1-uninfected participants at each MACS site were thawed and evaluated. The median recoveries of viable PBMC for HIV-1-infected and -uninfected participants were 80% and 83%, respectively. Thawed PBMC from both HIV-1-infected and -uninfected participants mounted a strong proliferative response to phytohemagglutinin, with median stimulation indices of 84 and 120, respectively. Expression of the lymphocyte surface markers CD3, CD4, and CD8 by thawed PBMC was virtually identical to what was observed on cells measured in real time using whole blood from the same participants. Furthermore, despite overall excellent performance of the four participating laboratories, problems were identified that intermittently compromised the quality of cryopreserved PBMC, which could be corrected and monitored for improvement over time. Ongoing quality assessment helps laboratories improve protocols and performance on a real-time basis to ensure optimal cryopreservation of PBMC for future studies.
Related JoVE Video
The impact of HAART on the respiratory complications of HIV infection: longitudinal trends in the MACS and WIHS cohorts.
PLoS ONE
PUBLISHED: 02-07-2013
Show Abstract
Hide Abstract
To review the incidence of respiratory conditions and their effect on mortality in HIV-infected and uninfected individuals prior to and during the era of highly active antiretroviral therapy (HAART).
Related JoVE Video
Identification of class I HLA T cell control epitopes for West Nile virus.
PLoS ONE
PUBLISHED: 01-01-2013
Show Abstract
Hide Abstract
The recent West Nile virus (WNV) outbreak in the United States underscores the importance of understanding human immune responses to this pathogen. Via the presentation of viral peptide ligands at the cell surface, class I HLA mediate the T cell recognition and killing of WNV infected cells. At this time, there are two key unknowns in regards to understanding protective T cell immunity: 1) the number of viral ligands presented by the HLA of infected cells, and 2) the distribution of T cell responses to these available HLA/viral complexes. Here, comparative mass spectroscopy was applied to determine the number of WNV peptides presented by the HLA-A*11:01 of infected cells after which T cell responses to these HLA/WNV complexes were assessed. Six viral peptides derived from capsid, NS3, NS4b, and NS5 were presented. When T cells from infected individuals were tested for reactivity to these six viral ligands, polyfunctional T cells were focused on the GTL9 WNV capsid peptide, ligands from NS3, NS4b, and NS5 were less immunogenic, and two ligands were largely inert, demonstrating that class I HLA reduce the WNV polyprotein to a handful of immune targets and that polyfunctional T cells recognize infections by zeroing in on particular HLA/WNV epitopes. Such dominant HLA/peptide epitopes are poised to drive the development of WNV vaccines that elicit protective T cells as well as providing key antigens for immunoassays that establish correlates of viral immunity.
Related JoVE Video
Factors associated with incorrect identification of recent HIV infection using the BED capture immunoassay.
AIDS Res. Hum. Retroviruses
PUBLISHED: 12-01-2011
Show Abstract
Hide Abstract
The BED capture enzyme immunoassay (BED-CEIA) was developed for estimating HIV incidence from cross-sectional data. This assay misclassifies some individuals with nonrecent HIV infection as recently infected, leading to overestimation of HIV incidence. We analyzed factors associated with misclassification by the BED-CEIA. We analyzed samples from 383 men who were diagnosed with HIV infection less than 1 year after a negative HIV test (Multicenter AIDS Cohort Study). Samples were collected 2-8 years after HIV seroconversion, which was defined as the midpoint between the last negative and first positive HIV test. Samples were analyzed using the BED-CEIA with a cutoff of OD-n ? 0.8 for recent infection. Logistic regression was used to identify factors associated with misclassification. Ninety-one (15.1%) of 603 samples were misclassified. In multivariate models, misclassification was independently associated with highly active antiretroviral treatment (HAART) for >2 years, HIV RNA <400 copies/ml, and CD4 cell count <50 or <200 cells/mm(3); adjusted odds ratios (OR) and 95% confidence intervals (CI) were 4.72 (1.35-16.5), 3.96 (1.53-10.3), 6.85 (2.71-17.4), and 11.5 (3.64-36.0), respectively. Among 220 men with paired samples, misclassification 2-4 years after seroconversion was significantly associated with misclassification 6-8 years after seroconversion [adjusted OR: 25.8 (95% CI: 8.17-81.5), p<0.001] after adjusting for race, CD4 cell count, HIV viral load, and HAART use. Low HIV viral load, low CD4 cell count, and >2 years of HAART were significantly associated with misclassification using the BED-CEIA. Some men were persistently misclassified as recently infected up to 8 years after HIV seroconversion.
Related JoVE Video
Clinical evaluation of multiplex real-time PCR panels for rapid detection of respiratory viral infections.
J. Med. Virol.
PUBLISHED: 06-27-2011
Show Abstract
Hide Abstract
Respiratory viral infections are one of the leading causes of morbidity and mortality, particularly in children, the elderly and immunocompromised persons. Rapid identification of viral etiology is critical in ruling out non-viral infections, initiating antiviral treatment and limiting the spread of the infection. Multiplex assays of more than one viral gene target in a single tube have the advantage of rapid screening of a large number of potential viral pathogens in a short time. A multiplex real-time PCR assay was used in this study for detection of respiratory RNA and DNA viral infections in 728 specimens received from 585 adult and pediatric patients comprised of symptomatic and asymptomatic organ transplant recipients and non-recipients for diagnosis of respiratory illnesses and for routine clinical monitoring. Multiplex PCR was more sensitive than the multiplex immunofluoresence culture assay (R-mix) and also detected additional respiratory viruses that were not covered by the R-mix panel. The number of respiratory viruses detected in symptomatic patients was significantly higher than asymptomatic patients in both adult and pediatric patients. Herpesviral infections were the predominant cause of lower respiratory tract infection in the organ transplant recipients, whereas respiratory syncytial virus was the most common pathogen in non-transplant patients particularly children. Multiplex real-time PCR for detection of respiratory viruses has the potential for rapid identification of viral pathogens. In this era of emerging viral infections, addition of newer viral targets to the multiplex PCR panels will be beneficial in determining both patient management and public health epidemiology.
Related JoVE Video
Multisite comparison of high-sensitivity multiplex cytokine assays.
Clin. Vaccine Immunol.
PUBLISHED: 06-22-2011
Show Abstract
Hide Abstract
The concentrations of cytokines in human serum and plasma can provide valuable information about in vivo immune status, but low concentrations often require high-sensitivity assays to permit detection. The recent development of multiplex assays, which can measure multiple cytokines in one small sample, holds great promise, especially for studies in which limited volumes of stored serum or plasma are available. Four high-sensitivity cytokine multiplex assays on a Luminex (Bio-Rad, BioSource, Linco) or electrochemiluminescence (Meso Scale Discovery) platform were evaluated for their ability to detect circulating concentrations of 13 cytokines, as well as for laboratory and lot variability. Assays were performed in six different laboratories utilizing archived serum from HIV-uninfected and -infected subjects from the Multicenter AIDS Cohort Study (MACS) and the Womens Interagency HIV Study (WIHS) and commercial plasma samples spanning initial HIV viremia. In a majority of serum samples, interleukin-6 (IL-6), IL-8, IL-10, and tumor necrosis factor alpha were detectable with at least three kits, while IL-1? was clearly detected with only one kit. No single multiplex panel detected all cytokines, and there were highly significant differences (P < 0.001) between laboratories and/or lots with all kits. Nevertheless, the kits generally detected similar patterns of cytokine perturbation during primary HIV viremia. This multisite comparison suggests that current multiplex assays vary in their ability to measure serum and/or plasma concentrations of cytokines and may not be sufficiently reproducible for repeated determinations over a long-term study or in multiple laboratories but may be useful for longitudinal studies in which relative, rather than absolute, changes in cytokines are important.
Related JoVE Video
Reduction in the incidence of influenza A but not influenza B associated with use of hand sanitizer and cough hygiene in schools: a randomized controlled trial.
Pediatr. Infect. Dis. J.
PUBLISHED: 06-22-2011
Show Abstract
Hide Abstract
Laboratory-based evidence is lacking regarding the efficacy of nonpharmaceutical interventions (NPIs) such as alcohol-based hand sanitizer and respiratory hygiene to reduce the spread of influenza.
Related JoVE Video
CD4+ T-cell counts and plasma HIV-1 RNA levels beyond 5 years of highly active antiretroviral therapy.
J. Acquir. Immune Defic. Syndr.
PUBLISHED: 05-24-2011
Show Abstract
Hide Abstract
The heterogeneity of CD4 T-cell counts and HIV-1 RNA at 5-12 years after the initiation of highly active antiretroviral therapy (HAART) remains largely uncharacterized.
Related JoVE Video
Effect of highly active antiretroviral therapy on biomarkers of B-lymphocyte activation and inflammation.
AIDS
PUBLISHED: 05-17-2011
Show Abstract
Hide Abstract
Chronic inflammation and B-cell hyperactivation are seen in HIV infection, contributing to an increased risk for the accrual of genetic errors that may result in B-cell lymphoma. The primary objective of this study was to determine the effect of highly active antiretroviral therapy (HAART) on serum levels of molecules that are associated with immune activation and/or inflammation, including several that are associated with B-cell activation, specifically IL-6, sCD30, sCD27, IgG, IgA, CXCL13 (B lymphocyte chemoattractant, BLC), a B-lymphocyte chemokine involved in B-cell trafficking, as well as C-reactive protein, an acute-phase protein.
Related JoVE Video
An optimized sensitive method for quantitation of DNA/RNA viruses in heparinized and cryopreserved plasma.
J. Virol. Methods
PUBLISHED: 05-02-2011
Show Abstract
Hide Abstract
Sodium heparin, an anticoagulant used widely for blood collection, has been known to inhibit DNA polymerase activity in polymerase chain reaction (PCR) assays. However, all cryopreserved plasma samples collected in the 1980s and early 1990s at the Multicenter AIDS Cohort Study were from heparin-treated blood, which poses a problem in quantifying the target nucleic acids contained in those samples by PCR assay. In this study, a nucleic acid extraction procedure was optimized to remove the heparin from extracted nucleic acids. Using this optimized method, similar human immunodeficiency virus 1 (HIV-1) and cytomegalovirus (CMV) loads of these viruses that were added to normal donor blood from ethylenediaminetetraacetic acid (EDTA), acid citrate dextrose (ACD) or sodium heparin tubes were detected by reverse transcriptase (RT) real-time PCR and real-time PCR. Comparable HIV-1 and CMV loads were also detected in the blood of persons with active HIV-1 and CMV infections collected in EDTA-, ACD- or sodium heparin-treated tubes by RT real-time and real-time PCR. The findings showed that the optimized nucleic acid extraction procedure efficiently removes the heparin inhibition effect on the performance of real-time PCR. This method could be used to extract nucleic acids from archived, heparinized plasma for PCR based quantitation of target molecules.
Related JoVE Video
B-cell stimulatory cytokines and markers of immune activation are elevated several years prior to the diagnosis of systemic AIDS-associated non-Hodgkin B-cell lymphoma.
Cancer Epidemiol. Biomarkers Prev.
PUBLISHED: 04-28-2011
Show Abstract
Hide Abstract
The risk of developing non-Hodgkin lymphoma (NHL) is greatly increased in HIV infection. The aim of this study was to determine whether elevated serum levels of molecules associated with B-cell activation precede the diagnosis of AIDS-associated NHL (AIDS-NHL).
Related JoVE Video
"In vitro systems to characterize the immune response to HIV-1 and HIV-1 vaccine candidates", NIAID Workshop Report, Bethesda, August 4, 2010.
Vaccine
PUBLISHED: 03-04-2011
Show Abstract
Hide Abstract
Although clinical trials are the ultimate way to prove vaccine safety and efficacy, the complexity, cost and time required to develop a product to enter human trials demand a serious, long-term investment. Lack of knowledge on immune correlates of protection from HIV infections makes investments in HIV vaccine research significantly risky. Preclinical testing of HIV vaccines is routinely carried out in non-human primate models however these studies have a significant cost and their predictive value is still questionable. The potential value of screening new HIV-1 vaccine candidates on human cells and tissues via high throughput in vitro systems that allow rapid, cost-effective and accurate predictions of in vivo immune responses would be enormous. A one-day workshop was convened by Division of AIDS, National Institutes of Health on August 4, 2010 to address the benefits and challenges of assessing HIV-1 vaccine responses in alternative ways. Consideration was given to the use of various in vitro model systems, human mucosal tissue explants and humanized mouse models as ways to predict immunogenicity and efficacy of HIV-1 vaccines early in the development process, and support decisions on whether a product may be worthy of moving into non-human primates or human trials. This report summarizes the outcome of the workshop.
Related JoVE Video
Influenza-like illness among university students: symptom severity and duration due to influenza virus infection compared to other etiologies.
J Am Coll Health
PUBLISHED: 02-11-2011
Show Abstract
Hide Abstract
University students with influenza-like illness (ILI) were assessed to determine whether symptom severity, duration, or missed days of school or work varied according to etiology.
Related JoVE Video
The dual impact of HIV-1 infection and aging on naïve CD4 T-cells: additive and distinct patterns of impairment.
PLoS ONE
PUBLISHED: 01-26-2011
Show Abstract
Hide Abstract
HIV-1-infected adults over the age of 50 years progress to AIDS more rapidly than adults in their twenties or thirties. In addition, HIV-1-infected individuals receiving antiretroviral therapy (ART) present with clinical diseases, such as various cancers and liver disease, more commonly seen in older uninfected adults. These observations suggest that HIV-1 infection in older persons can have detrimental immunological effects that are not completely reversed by ART. As naïve T-cells are critically important in responses to neoantigens, we first analyzed two subsets (CD45RA(+)CD31(+) and CD45RA(+)CD31(-)) within the naïve CD4(+) T-cell compartment in young (20-32 years old) and older (39-58 years old), ART-naïve, HIV-1 seropositive individuals within 1-3 years of infection and in age-matched seronegative controls. HIV-1 infection in the young cohort was associated with lower absolute numbers of, and shorter telomere lengths within, both CD45RA(+)CD31(+)CD4(+) and CD45RA(+)CD31(-)CD4(+) T-cell subsets in comparison to age-matched seronegative controls, changes that resembled seronegative individuals who were decades older. Longitudinal analysis provided evidence of thymic emigration and reconstitution of CD45RA(+)CD31(+)CD4(+) T-cells two years post-ART, but minimal reconstitution of the CD45RA(+)CD31(-)CD4(+) subset, which could impair de novo immune responses. For both ART-naïve and ART-treated HIV-1-infected adults, a renewable pool of thymic emigrants is necessary to maintain CD4(+) T-cell homeostasis. Overall, these results offer a partial explanation both for the faster disease progression of older adults and the observation that viral responders to ART present with clinical diseases associated with older adults.
Related JoVE Video
Demographic and clinical factors associated with persistent symptoms after West Nile virus infection.
Am. J. Trop. Med. Hyg.
PUBLISHED: 11-02-2010
Show Abstract
Hide Abstract
Prognosis varies among persons with West Nile virus (WNV) infection, but the most important factors associated with persistent symptoms are not clear. In this cross-sectional study, 265 persons with symptomatic WNV infection during 2006-2008 completed a survey a mean of 7.7 months after diagnosis. We determined the association of demographic and clinical characteristics to the most common symptoms. Of 214 persons infected ? 6 months, 53% reported one or more persistent symptoms, including fatigue, muscle aches, decreased activity, difficulty with memory, and difficulty concentrating. Persons with neuroinvasive disease, hypertension, or diabetes were significantly more likely to report persistent symptoms, whereas age, sex, and time since infection were not associated with persistent symptoms. In conclusion, persistent symptoms persisted in most persons for more than six months after symptomatic WNV infection. Improved strategies for prevention and treatment are needed.
Related JoVE Video
Sensitivity and specificity of rapid influenza testing of children in a community setting.
Influenza Other Respir Viruses
PUBLISHED: 10-01-2010
Show Abstract
Hide Abstract
Rapid influenza testing (RFT) allows for a rapid point-of-care diagnosis of influenza. The Quidel QuickVue Influenza A+B test (QuickVue) has a reported manufacturers sensitivity and specificity of 73% and 96%, respectively, with nasal swabs. However, investigators have shown sensitivities ranging from 22% to 77% in community settings.
Related JoVE Video
Monofunctional and polyfunctional CD8+ T cell responses to human herpesvirus 8 lytic and latency proteins.
Clin. Vaccine Immunol.
PUBLISHED: 08-18-2010
Show Abstract
Hide Abstract
Human herpesvirus 8 (HHV-8) is the etiological agent of Kaposis sarcoma, primary effusion lymphoma, and multicentric Castlemans disease. It is postulated that CD8(+) T cell responses play an important role in controlling HHV-8 infection and preventing development of disease. In this study, we investigated monofunctional and polyfunctional CD8(+) T cell responses to HHV-8 lytic proteins gB (glycoprotein B) and K8.1 and latency proteins LANA-1 (latency-associated nuclear antigen-1) and K12. On the basis of our previous findings that dendritic cells (DC) reveal major histocompatibility complex (MHC) class I epitopes in gB, we used a DC-based system to identify 2 novel epitopes in gB, 2 in K8.1, 5 in LANA-1, and 1 in K12. These new HHV-8 epitopes activated monofunctional and polyfunctional CD8(+) T cells that produced various combinations of gamma interferon, interleukin 2, tumor necrosis factor alpha, macrophage inhibitory protein 1?, and cytotoxic degranulation marker CD107a in healthy HHV-8-seropositive individuals. We were also able to detect HHV-8-specific CD8(+) T cells in peripheral blood samples using HLA A*0201 pentamer complexes for one gB epitope, one K8.1 epitope, two LANA-1 epitopes, and one K12 epitope. These immunogenic regions of viral lytic and latency proteins could be important in T cell control of HHV-8 infection.
Related JoVE Video
Surface phenotype and functionality of WNV specific T cells differ with age and disease severity.
PLoS ONE
PUBLISHED: 08-13-2010
Show Abstract
Hide Abstract
West Nile virus (WNV) infection can result in severe neuroinvasive disease, particularly in persons with advanced age. As rodent models demonstrate that T cells play an important role in limiting WNV infection, and strong T cell responses to WNV have been observed in humans, we postulated that inadequate antiviral T cell immunity was involved in neurologic sequelae and the more severe outcomes associated with age. We previously reported the discovery of six HLA-A*0201 restricted WNV peptide epitopes, with the dominant T cell targets in naturally infected individuals being SVG9 (Env) and SLF9 (NS4b). Here, memory phenotype and polyfunctional CD8+ T cell responses to these dominant epitopes were assessed in 40 WNV seropositive patients displaying diverse clinical symptoms. The patients PBMC were stained with HLA-I multimers loaded with the SVG9 and SLF9 epitopes and analyzed by multicolor flow cytometry. WNV-specific CD8+ T cells were found in peripheral blood several months post infection. The number of WNV-specific T cells in older individuals was the same, if not greater, than in younger members of the cohort. WNV-specific T cells were predominantly monofunctional for CD107a, MIP-1?, TNF?, IL-2, or IFN?. When CD8+ T cell responses were stratified by disease severity, an increased number of terminally differentiated, memory phenotype (CD45RA+ CD27- CCR7- CD57+) T cells were detected in patients suffering from viral neuroinvasion. In conclusion, T cells of a terminally differentiated/cytolytic profile are associated with neuroinvasion and, regardless of age, monofunctional T cells persist following infection. These data provide the first indication that particular CD8+ T cell phenotypes are associated with disease outcome following WNV infection.
Related JoVE Video
Comparison of immunologic assays for detecting immune responses in HIV immunotherapeutic studies: AIDS Clinical Trials Group Trial A5181.
Clin. Vaccine Immunol.
PUBLISHED: 07-14-2010
Show Abstract
Hide Abstract
This study was designed to evaluate which of several T-cell-specific, immune response assays are the most relevant in measuring the key characteristics of an effective immune response to HIV-1. Using 5 HIV-1 antigens as stimulants, we assessed lymphocyte proliferation, supernatant gamma interferon (IFN-gamma) cytokine production (CP), single-cell IFN-gamma production by enzyme-linked immunospot (ELISPOT) assay, with and without Epstein-Barr virus-transformed B-lymphoblastoid cell lines (B-LCLs), and intracellular cytokine production (ICC) for IFN-gamma and interleukin 2 (IL-2) by flow cytometry. We used these to compare specimens from HIV-1-infected subjects who were virally suppressed with a stable antiretroviral therapy (ART) regimen (group A) with specimens from subjects not on ART but with HIV-1 viremia of <3,000 copies/ml (group B). The lymphocyte proliferation assay (LPA) did not significantly differentiate between the two groups. Using fresh peripheral blood mononuclear cells (PBMCs), the CP and ELISPOT assays for IFN-gamma detected the greatest differences between the two groups, specific for three of the five HIV-1 antigens, whereas significant differences were seen only in response to one antigen when cryopreserved cells were used. The strongest correlations were seen between the CP and ELISPOT assays. The ELISPOT B-LCL assay showed a cell concentration-dependent increase in IFN-gamma production compared to that shown by the standard ELISPOT assay but did not differentiate between the groups. In the ICC assay, greater numbers of IFN-gamma-producing T cells were seen in group B, and little or no detectable IL-2 production was seen in both groups. These studies highlight complexities of immunologic monitoring of T-cell responses in multisite clinical trials in HIV infection and outline considerations for optimizing these efforts.
Related JoVE Video
Dendritic cells reveal a broad range of MHC class I epitopes for HIV-1 in persons with suppressed viral load on antiretroviral therapy.
PLoS ONE
PUBLISHED: 06-25-2010
Show Abstract
Hide Abstract
HIV-1 remains sequestered during antiretroviral therapy (ART) and can resume high-level replication upon cessation of ART or development of drug resistance. Reactivity of memory CD8(+) T lymphocytes to HIV-1 could potentially inhibit this residual viral replication, but is largely muted by ART in relation to suppression of viral antigen burden. Dendritic cells (DC) are important for MHC class I processing and presentation of peptide epitopes to memory CD8(+) T cells, and could potentially be targeted to activate memory CD8(+) T cells to a broad array of HIV-1 epitopes during ART.
Related JoVE Video
A Twenty-Two-Year-Old Community Advisory Board: Health Research as an Opportunity for Social Change.
J Community Pract
PUBLISHED: 06-05-2010
Show Abstract
Hide Abstract
Conducting health research often requires a partnership between marginalized communities and researchers. Community organizers can broker this partnership in a way that not only produces important scientific discoveries but also brings needed resources to the communities. This article is a description of a community advisory board established in 1984 to advise researchers on a longitudinal study of the natural history of AIDS among gay men. The Board successfully guided the recruitment of more than 3,000 gay and bisexual male volunteers and, at the same time worked as a powerful change agent. An analysis of minutes from all Board meetings between 1984-2006 indicates that significant social change as well as important research findings resulted from Board actions. Community organizers who work to create a mutually beneficial partnership between communities and researchers may find new opportunities to support community growth and social justice.
Related JoVE Video
Regulatory T cells in HIV immunotherapy.
HIV Ther
PUBLISHED: 06-01-2010
Show Abstract
Hide Abstract
Significant research has been conducted on the role of regulatory T cells (Tregs) in HIV infection. To date, however, it is not clear whether Tregs play a detrimental role or a beneficial role in the pathogenesis of HIV infection. In fact, a number of immunotherapeutic strategies to control HIV infection have revealed a possible antagonistic role for Tregs. This necessitates investigating ways to counteract the suppressive function, such as through Treg depletion or blockade of specific Treg immunosuppressive mechanisms, without further increasing the cellular immune activation associated with chronic HIV infection. Simply applying Treg immunotherapeutic strategies used in diseases other than HIV may pose problems due to the complexity of HIV immunopathogenesis. Studies are therefore necessary to elucidate the different immunoregulatory networks in HIV infection in order to determine the specific cellular or molecular pathways that can be altered to boost the bodys immune control of HIV.
Related JoVE Video
Safety, tolerability, and mechanisms of antiretroviral activity of pegylated interferon Alfa-2a in HIV-1-monoinfected participants: a phase II clinical trial.
J. Infect. Dis.
PUBLISHED: 04-28-2010
Show Abstract
Hide Abstract
To our knowledge, the antiviral activity of pegylated interferon alfa-2a has not been studied in participants with untreated human immunodeficiency virus type 1 (HIV-1) infection but without chronic hepatitis C virus (HCV) infection.
Related JoVE Video
The relationship between antibody to R7V and progression of HIV type 1 infection.
AIDS Res. Hum. Retroviruses
PUBLISHED: 04-27-2010
Show Abstract
Hide Abstract
The presence of antibody to R7V (anti-R7VAb), a seven-amino acid sequence derived from beta(2)-microglobulin incorporated into HIV-1 virions from the surface of infected cells, has been proposed as an early marker of nonprogressive HIV-1 infection. The present study was undertaken because no prospective studies have tested this hypothesis. Stored samples collected prospectively from 361 HIV-1 seroconverting men in the Multicenter AIDS Cohort Study (0.44-1.53 years after seroconversion) were assayed for the presence or absence of anti-R7VAb, using a standardized ELISA. Using Cox proportional hazards models, crude and adjusted relative hazards (RH) were determined for the following outcomes: (a) clinically defined AIDS, (b) clinically defined AIDS or CD4 T cell count of <200 cells/microl, and (c) death. A total of 143 (39.6%) men had early anti-R7VAb and 218 (60.4%) did not; 192 (53.2%) developed AIDS. At the visit tested, men with anti-R7VAb had significantly lower CD4 T cell counts and higher plasma HIV-1 viral loads than those without antibody. After adjustment for CD4 T cell count, HIV-1 viral load, CCR5 polymorphism, and use of combined antiretroviral therapy, the presence of anti-R7VAb was associated with a higher risk of progression for all outcomes, but not significantly so. Absence of anti-R7VAb was significantly associated with expression of HLA-B*5701 and -B*2705, two alleles associated with slower progression of HIV-1 disease. The early presence of anti-R7VAb in HIV-1 seroconverters was not associated with slower progression of HIV-1 disease.
Related JoVE Video
Regulatory T cell suppression of Gag-specific CD8 T cell polyfunctional response after therapeutic vaccination of HIV-1-infected patients on ART.
PLoS ONE
PUBLISHED: 02-05-2010
Show Abstract
Hide Abstract
We tested the hypothesis that therapeutic vaccination against HIV-1 can increase the frequency and suppressive function of regulatory, CD4(+) T cells (Treg), thereby masking enhancement of HIV-1-specific CD8(+) T cell response. HIV-1-infected subjects on antiretroviral therapy (N = 17) enrolled in a phase I therapeutic vaccine trial received 2 doses of autologous dendritic cells (DC) loaded with HIV-1 peptides. The frequency of CD4(+)CD25(hi)FOXP3(+) Treg in blood was determined prior to and after vaccination in subjects and normal controls. Polyfunctional CD8(+) T cell responses were determined pre- and post-vaccine (N = 7) for 5 immune mediators after in vitro stimulation with Gag peptide, staphylococcal enterotoxin B (SEB), or medium alone. Total vaccine response (post-vaccine-pre-vaccine) was compared in the Treg(+) and Treg-depleted (Treg-) sets. After vaccination, 12/17 subjects showed a trend of increased Treg frequency (P = 0.06) from 0.74% to 1.2%. The increased frequency did not correlate with CD8(+) T cell vaccine response by enzyme linked immunosorbent assay for interferon gamma production. Although there was no significant change in CD8(+) T cell polyfunctional response after vaccination, Treg depletion increased the polyfunctionality of the total vaccine response (P = 0.029), with a >2-fold increase in the percentage of CD8(+) T cells producing multiple immune mediators. In contrast, depletion of Treg did not enhance polyfunctional T cell response to SEB, implying specificity of suppression to HIV-1 Gag. Therapeutic immunization with a DC-based vaccine against HIV-1 caused a modest increase in Treg frequency and a significant increase in HIV-1-specific, Treg suppressive function. The Treg suppressive effect masked an increase in the vaccine-induced anti-HIV-1-specific polyfunctional response. The role of Treg should be considered in immunotherapeutic trials of HIV-1 infection.
Related JoVE Video
PD-1 blockade: A promising immunotherapy for HIV?
Cellscience
PUBLISHED: 09-09-2009
Show Abstract
Hide Abstract
The progressive loss of effector function in the setting of chronic viral infections has been associated with the upregulation of programmed death 1 (PD-1), a negative regulator of activated T cells. In HIV infection, increased levels of PD-1 expression correlate with CD8(+) T cell exhaustion, which has been shown in vitro to be reversible with PD-1 blockade. Velu and colleagues recently reported the first in vivo study showing enhancement of a virus-specific immune response through PD-1 blockade using an anti-PD-1 antibody in an SIV-macaque model. Their results show an expansion of virus-specific, polyfunctional CD8(+) T cells. Anti-PD1 antagonists show promise as a novel immunotherapy for HIV. However, several issues including development of autoimmunity, regulatory T cells and multiple inhibitory receptors associated with CD8(+) T cell exhaustion should first be addressed to help ensure a successful response in chronic HIV infected patients.
Related JoVE Video
Multiple T-cell responses to human immunodeficiency virus type 1 are enhanced by dendritic cells.
Clin. Vaccine Immunol.
PUBLISHED: 08-19-2009
Show Abstract
Hide Abstract
Human immunodeficiency virus type 1 (HIV-1)-specific T-cell reactivity has been related to protection from disease progression. Optimal T-cell reactivity to HIV-1 presumably requires antigen processing and presentation by professional antigen-presenting cells, particularly dendritic cells (DC). Here we examined whether multiple HIV-1-specific T-cell functions are enhanced by stimulation with HIV-1 peptide-loaded DC derived from HIV-1-infected subjects on antiretroviral therapy. We first found that mature DC increased the number of gamma interferon (IFN-gamma)-producing T cells detected by enzyme-linked immunospot assay to overlapping 15-mer peptides of HIV-1 Gag and Nef, compared to stimulation with peptide-loaded, immature DC or to peptides without DC. IFN-gamma production was lower in response to large pools of the Gag and Nef peptides, regardless of presentation by DC. We further observed that HIV-1 peptide-loaded, mature DC stimulated greater CD8(+) and CD4(+) T-cell proliferation than did the peptides without DC and that T-cell proliferation was lower in response to larger pools of the peptides. The lower T-cell IFN-gamma and proliferation responses to the larger peptide pools were related to lower T-cell viability. Finally, the number of polyfunctional CD8(+) and CD4(+) T cells stimulated by HIV-1 peptide-loaded, mature DC, defined as positive by intracellular staining for more than one immune mediator (IFN-gamma, interleukin 2, tumor necrosis factor alpha, macrophage inhibitory protein 1beta, or CD107a), was greater than that stimulated by the peptides alone. These results indicate that DC can enhance multiple types of HIV-1-specific T-cell functions.
Related JoVE Video
Detection of HIV-1 RNA/DNA and CD4 mRNA in feces and urine from chronic HIV-1 infected subjects with and without anti-retroviral therapy.
AIDS Res Ther
PUBLISHED: 07-07-2009
Show Abstract
Hide Abstract
HIV-1 infects gut associated lymphoid tissues (GALT) very early after transmission by multiple routes. The infected GALT consequently serves as the major reservoir for HIV-1 infection and could constantly shed HIV-1 and CD4+ T cells into the intestinal lumen. To examine this hypothesis, we monitored HIV-1 RNA/DNA and CD4 mRNA in fecal samples of chronically infected subjects with and without antiretroviral therapy (ART). We compared this to levels of HIV-1 RNA/DNA in urine and blood from the same subjects. Our results show that HIV-1 DNA, RNA and CD4 mRNA were detected in 8%, 19% and 31% respectively, of feces samples from infected subjects with detectable plasma viral load, and were not detected in any of subjects on ART with undetectable plasma viral load. In urine samples, HIV-1 DNA was detected in 24% of infected subjects with detectable plasma viral load and 23% of subjects on ART with undetectable plasma viral load. Phylogenetic analysis of the envelope sequences of HIV-1 revealed distinct virus populations in concurrently collected serum, feces and urine samples from one subject. In addition, our study demonstrated for the first time the presence of CD4 mRNA in fecal specimens of HIV-1 infected subjects, which could be used to assess GALT pathogenesis in HIV-1 infection.
Related JoVE Video
Primary human immunodeficiency virus type 1-specific CD8+ T-cell responses induced by myeloid dendritic cells.
J. Virol.
PUBLISHED: 04-08-2009
Show Abstract
Hide Abstract
Induction of an antigenically broad and vigorous primary T-cell immune response by myeloid dendritic cells (DC) in blood and tissues could be important for an effective prophylactic or therapeutic vaccine to human immunodeficiency virus type 1 (HIV-1). Here we show that a primary CD8(+) T-cell response can be induced by HIV-1 peptide-loaded DC derived from blood monocytes of HIV-1-negative adults and neonates (moDC) and by Langerhans cells (LC) and interstitial, dermal-intestinal DC (idDC) derived from CD34(+) stem cells of neonatal cord blood. Optimal priming of single-cell gamma interferon (IFN-gamma) production by CD8(+) T cells required CD4(+) T cells and was broadly directed to multiple regions of Gag, Env, and Nef that corresponded to known and predicted major histocompatibility complex class I epitopes. Polyfunctional CD8(+) T-cell responses, defined as single-cell production of more than one cytokine (IFN-gamma, interleukin 2, or tumor necrosis factor alpha), chemokine (macrophage inhibitory factor 1beta), or cytotoxic degranulation marker CD107a, were primed by moDC, LC, and idDC to HIV-1 Gag and reverse transcriptase epitopes, as well as to Epstein-Barr virus and influenza A virus epitopes. Thus, three major types of blood and tissue myeloid DC targeted by HIV-1, i.e., moDC, LC, and idDC, can prime multispecific, polyfunctional CD8(+) T-cell responses to HIV-1 and other viral antigens.
Related JoVE Video
Fidelity of SNP array genotyping using Epstein Barr virus-transformed B-lymphocyte cell lines: implications for genome-wide association studies.
PLoS ONE
PUBLISHED: 04-07-2009
Show Abstract
Hide Abstract
As availability of primary cells can be limited for genetic studies of human disease, lymphoblastoid cell lines (LCL) are common sources of genomic DNA. LCL are created in a transformation process that entails in vitro infection of human B-lymphocytes with the Epstein-Barr Virus (EBV).
Related JoVE Video
Low sensitivity of rapid diagnostic test for influenza.
Clin. Infect. Dis.
PUBLISHED: 03-28-2009
Show Abstract
Hide Abstract
The QuickVue Influenza A+B Test (Quidel) was used to test nasal swab specimens obtained from persons with influenza-like illness in 3 different populations. Compared with reverse-transcriptase polymerase chain reaction, the test sensitivity was low for all populations (median, 27%; range, 19%-32%), whereas the specificity was high (median, 97%; range, 96%-99.6%).
Related JoVE Video
Production of a dendritic cell-based vaccine containing inactivated autologous virus for therapy of patients with chronic human immunodeficiency virus type 1 infection.
Clin. Vaccine Immunol.
PUBLISHED: 03-10-2009
Show Abstract
Hide Abstract
In preparation for a pilot clinical trial in patients with chronic human immunodeficiency virus type 1 (HIV-1) infection, a novel dendritic cell (DC)-based vaccine is being manufactured. The trial will test the hypothesis that isolated endogenous virus presented by DCs serves as a potent immunogen for activation of CD8(+) and CD4(+) T cells specific for a broad range of autologous HIV-1 antigens. Production of the vaccine under good manufacture practice conditions involves (i) autologous virus isolation; (ii) superinfection of CD4(+) T cells with the virus; (iii) inactivation of the virus in CD4(+) T cells, T-cell apoptosis, and coincubation of T cells with autologous DCs; and (iv) product testing and release. Endogenous virus was isolated from peripheral blood-derived CD4(+) T cells of three HIV-1-positive subjects by coincubation with autologous OKT-3-stimulated CD4(+) T cells. CD4(+) T-cell supernatants were tested for p24 levels by enzyme-linked immunosorbent assay (>25 ng/ml) and for the 50% tissue culture infective doses (TCID(50); which ranged from 4,642 to 46,416/ml on day 19 of culture). Autologous CD4(+) T cells that were separated on immunobeads (>95% purity) and superinfected with virus-expressed p24 (28 to 54%) had TCID(50) of >400/ml on days 5 to 10. Virus inactivation with psoralen (20 microg/ml) and UVB irradiation (312 nm) reduced the TCID(50) of the supernatants from 199,986 to 11/ml (>99%). 7-Amino-actinomycin D-positive, annexin V-positive CD4(+) T cells were fed to autologous DCs generated by using the Elutra cell separation system and the Aastrom system. Flow analysis showed that DC loading was complete in 24 h. On the basis of these translational results and experience with the generation of DCs from HIV-1-infected patients in a previous clinical trial, the Investigational New Drug application for clinical vaccination was submitted and approved by the FDA (application no. BB-IND-13137).
Related JoVE Video
Cytokine production by human herpesvirus 8-infected dendritic cells.
J. Gen. Virol.
PUBLISHED: 02-13-2009
Show Abstract
Hide Abstract
We have shown previously that human herpesvirus 8 (HHV-8)-infected dendritic cells (DCs) undergo incomplete maturation and have a defective antigen-presenting function. Here, we examined the effects of HHV-8 infection on cytokine production, which is critical to the function of DCs. We detected expression of interleukin (IL)-6, tumour necrosis factor (TNF)-alpha, macrophage inflammatory protein (MIP)-1alpha, MIP-1beta, RANTES and IL-12p40 from 2 to 6 h post-infection, and these peaked by 15-24 h. Expression of these factors decreased 24-48 h post-infection, with the exception of TNF-alpha which remained high throughout the entire 72 h. Interestingly, while IL-12p40 expression increased post-infection, bioactive IL-12p70 was not detected in the supernatants. These results suggest an intentional skewing of cytokine production in HHV-8-infected DCs towards induction of a Th2 response.
Related JoVE Video
Professional antigen presenting cells in human herpesvirus 8 infection.
Front Immunol
Show Abstract
Hide Abstract
Professional antigen presenting cells (APC), i.e., dendritic cells (DC), monocytes/macrophages, and B lymphocytes, are critically important in the recognition of an invading pathogen and presentation of antigens to the T cell-mediated arm of immunity. Human herpesvirus 8 (HHV-8) is one of the few human viruses that primarily targets these APC for infection, altering their cytokine profiles, manipulating their surface expression of MHC molecules, and altering their ability to activate HHV-8-specific T cells. This could be why T cell responses to HHV-8 antigens are not very robust. Of these APC, only B cells support complete, lytic HHV-8 infection. However, both complete and abortive virus replication cycles in APC could directly affect viral pathogenesis and progression to Kaposis sarcoma (KS) and HHV-8-associated B cell cancers. In this review, we discuss the effects of HHV-8 infection on professional APC and their relationship to the development of KS and B cell lymphomas.
Related JoVE Video
Serum levels of the chemokine CXCL13, genetic variation in CXCL13 and its receptor CXCR5, and HIV-associated non-hodgkin B-cell lymphoma risk.
Cancer Epidemiol. Biomarkers Prev.
Show Abstract
Hide Abstract
CXCL13 and CXCR5 are a chemokine and receptor pair whose interaction is critical for naïve B-cell trafficking and activation within germinal centers. We sought to determine whether CXCL13 levels are elevated before HIV-associated non-Hodgkin B-cell lymphoma (AIDS-NHL), and whether polymorphisms in CXCL13 or CXCR5 are associated with AIDS-NHL risk and CXCL13 levels in a large cohort of HIV-infected men.
Related JoVE Video
Predictors of the isolated hepatitis B core antibody pattern in HIV-infected and -uninfected men in the multicenter AIDS cohort study.
Clin. Infect. Dis.
Show Abstract
Hide Abstract
The significance of hepatitis B core antibody (anti-HBc) without hepatitis B surface antigen (HBsAg) or hepatitis B surface antibody (anti-HBs) is unclear.
Related JoVE Video
HIV and aging: state of knowledge and areas of critical need for research. A report to the NIH Office of AIDS Research by the HIV and Aging Working Group.
J. Acquir. Immune Defic. Syndr.
Show Abstract
Hide Abstract
HIV risk behaviors, susceptibility to HIV acquisition, progression of disease after infection, and response to antiretroviral therapy all vary by age. In those living with HIV, current effective treatment has increased the median life expectancy to >70 years of age. Biologic, medical, individual, social, and societal issues change as one ages with HIV infection, but there has been only a small amount of research in this field. Therefore, the Office of AIDS Research of the National Institutes of Health commissioned a working group to develop an outline of the current state of knowledge and areas of critical need for research in HIV and Aging; the working groups findings and recommendations are summarized in this report. Key overarching themes identified by the group included the following: multimorbidity, polypharmacy, and the need to emphasize maintenance of function; the complexity of assessing HIV versus treatment effects versus aging versus concurrent disease; the inter-related mechanisms of immune senescence, inflammation, and hypercoagulability; the utility of multivariable indices for predicting outcomes; a need to emphasize human studies to account for complexity; and a required focus on issues of community support, caregivers, and systems infrastructure. Critical resources are needed to enact this research agenda and include expanded review panel expertise in aging, functional measures, and multimorbidity, and facilitated use and continued funding to allow long-term follow-up of cohorts aging with HIV.
Related JoVE Video
Early HLA-B*57-restricted CD8+ T lymphocyte responses predict HIV-1 disease progression.
J. Virol.
Show Abstract
Hide Abstract
Although HLA-B*57 (B57) is associated with slow progression to disease following HIV-1 infection, B57 heterozygotes display a wide spectrum of outcomes, including rapid progression, viremic slow progression, and elite control. Efforts to identify differences between B57-positive (B57(+)) slow progressors and B57(+) rapid progressors have largely focused on cytotoxic T lymphocyte (CTL) phenotypes and specificities during chronic stages of infection. Although CTL responses in the early months of infection are likely to be the most important for the long-term rate of HIV-1 disease progression, few data on the early CTL responses of eventual slow progressors have been available. Utilizing the Multicenter AIDS Cohort Study (MACS), we retrospectively examined the early HIV-1-specific CTL responses of 14 B57(+) individuals whose time to development of disease ranged from 3.5 years to longer than 25 years after infection. In general, a greater breadth of targeting of epitopes from structural proteins, especially Gag, as well as of highly conserved epitopes from any HIV-1 protein, correlated with longer times until disease. The single elite controller in the cohort was an outlier on several correlations of CTL targeting and time until disease, consistent with reports that elite control is typically not achieved solely by protective HLA-mediated CTLs. When targeting of individual epitopes was analyzed, we found that early CTL responses to the IW9 (ISPRTLNAW) epitope of Gag, while generally subdominant, correlated with delayed progression to disease. This is the first study to identify early CTL responses to IW9 as a correlate of protection in persons with HLA-B*57.
Related JoVE Video
HIV status, burden of comorbid disease, and biomarkers of inflammation, altered coagulation, and monocyte activation.
Clin. Infect. Dis.
Show Abstract
Hide Abstract
Biomarkers of inflammation, altered coagulation, and monocyte activation are associated with mortality and cardiovascular disease (CVD) in the general population and among human immunodeficiency virus (HIV)-infected people. We compared biomarkers for inflammation, altered coagulation, and monocyte activation between HIV-infected and uninfected people in the Veterans Aging Cohort Study (VACS).
Related JoVE Video
Comparative risk of liver-related mortality from chronic hepatitis B versus chronic hepatitis C virus infection.
Clin. Infect. Dis.
Show Abstract
Hide Abstract
It is not known whether chronic hepatitis B (CH-B) or chronic hepatitis C (CH-C) carries a greater risk of liver-related mortality. This study compared rates of liver-related mortality between these 2 groups in the Multicenter AIDS Cohort Study (MACS).
Related JoVE Video
B-cell activation induced microRNA-21 is elevated in circulating B cells preceding the diagnosis of AIDS-related non-Hodgkin lymphomas.
AIDS
Show Abstract
Hide Abstract
We show that microRNA-21 is significantly elevated in peripheral B cells of HIV-infected individuals who go on to develop AIDS-related non-Hodgkin lymphoma (n=13, <3 years prior to diagnosis) when compared with HIV-negative (n=18) or HIV-positive controls (n=21) (P<0.01). Moreover, miR-21 is overexpressed in activated B cells and can be induced by interleukin 4 alone, or with CD40 or immunoglobulin M co-stimulation, and lipopolysaccharides, suggesting that miR-21 may help maintain B-cell hyperactivation, contributing to lymphomagenesis.
Related JoVE Video
Herpes simplex virus type 2 (HSV-2) as a coronary atherosclerosis risk factor in HIV-infected men: multicenter AIDS cohort study.
Atherosclerosis
Show Abstract
Hide Abstract
We assessed associations of herpes simplex virus types 1 and 2 (HSV-1 and -2), cytomegalovirus (CMV), and human herpesvirus 8 (HHV-8) infection with subclinical coronary atherosclerosis in 291 HIV-infected men in the Multicenter AIDS Cohort Study. Coronary artery calcium (CAC) was measured by non-contrast coronary CT imaging. Markers for herpesviruses infection were measured in frozen specimens collected 10-12 years prior to case identification. Multivariable logistic regression models and ordinal logistic regression models were performed. HSV-2 seropositivity was associated with coronary atherosclerosis (adjusted odds ratio [AOR]=4.12, 95% confidence interval [CI]=1.58-10.85) after adjustment for age, race/ethnicity, cardiovascular risk factors, and HIV infection related factors. Infection with a greater number of herpesviruses was associated with elevated CAC levels (AOR=1.58, 95% CI=1.06-2.36). Our findings suggest HSV-2 may be a risk factor for subclinical coronary atherosclerosis in HIV-infected men. Infection with multiple herpesviruses may contribute to the increased burden of atherosclerosis.
Related JoVE Video

What is Visualize?

JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

How does it work?

We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.

Video X seems to be unrelated to Abstract Y...

In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.