Project description:We examined the gene expression profiles in ex vivo human CD4+ and CD8+ T cells from untreated HIV-infected individuals at different clinical stages and rates of disease progression. Profiles of pure CD4+ and CD8+ T cells subsets from HIV-infected nonprogressors who controlled viremia were indistinguishable from HIV-uninfected individuals. Similarly, no gene clusters could distinguish T cells from individuals with early from chronic progressive HIV infection, whereas differences were observed between uninfected or nonprogressors versus early or chronic progressors. In early/chronic HIV infection, three characteristic gene expression signatures were observed: (1) CD4+ and CD8+ T cells showed increased expression of interferon stimulated genes (ISGs). However, some ISGs including CXCL9, CXCL10, and CXCL11, and the IL15R? in both CD4+ and CD8+ T cells and the anti-HIV ISG APOBEC3G in CD4+ T cells, were not upregulated. (2) CD4+ and CD8+ T cells showed a cluster similar to that observed in thymocytes, and (3) more genes were differentially regulated in CD8+ T cells than in CD4+ T cells, including a cluster of genes downregulated exclusively in CD8+ T cells. In conclusion, HIV infection induces a persistent T cell transcriptional profile, early in infection, characterized by a dramatic but potentially aberrant interferon response, and a profile suggesting an active thymic output. We studied a cohort of HIV infected individuals with various clinical stages of HIV infection and healthy uninfected volunteers as a control group (Table 1). We included 5 individuals with early HIV infection (A), five with chronic progressive HIV infection (C), five individuals with non-progressive HIV infection with low or undetectable viral loads (L) and five HIV uninfected individuals (N). The HIV infected individuals were never on therapy prior to entering the study. Samples were taken once from each donor.
Project description:Genome wide DNA methylation profiling of CD4 T cells from uninfected and HIV-infected individuals (viremic, ART-suppressed and elite controllers [EC]) The Illumina Infinium 450k Human DNA methylation Beadchip v1.2 was used to obtain DNA methylation profiles across approximately 485,577 CpGs in DNA from peripheral CD4 T cells samples. Samples included: 22 from HIV-uninfected individuals (uninfected group), 42 from HIV-infected individuals (21 from HIV-infected viremic (viremic group) and 21 from the same participants after viral suppression (viral load< 50 copies HIV-1 RNA/plasma) by antiretroviral therapy administrarion (ART group), and 21 from elite controllers (EC group)
Project description:We examined the gene expression profiles in ex vivo human CD4+ and CD8+ T cells from untreated HIV-infected individuals at different clinical stages and rates of disease progression. Profiles of pure CD4+ and CD8+ T cells subsets from HIV-infected nonprogressors who controlled viremia were indistinguishable from HIV-uninfected individuals. Similarly, no gene clusters could distinguish T cells from individuals with early from chronic progressive HIV infection, whereas differences were observed between uninfected or nonprogressors versus early or chronic progressors. In early/chronic HIV infection, three characteristic gene expression signatures were observed: (1) CD4+ and CD8+ T cells showed increased expression of interferon stimulated genes (ISGs). However, some ISGs including CXCL9, CXCL10, and CXCL11, and the IL15Rα in both CD4+ and CD8+ T cells and the anti-HIV ISG APOBEC3G in CD4+ T cells, were not upregulated. (2) CD4+ and CD8+ T cells showed a cluster similar to that observed in thymocytes, and (3) more genes were differentially regulated in CD8+ T cells than in CD4+ T cells, including a cluster of genes downregulated exclusively in CD8+ T cells. In conclusion, HIV infection induces a persistent T cell transcriptional profile, early in infection, characterized by a dramatic but potentially aberrant interferon response, and a profile suggesting an active thymic output. Keywords: disease state analysis
Project description:Assessment of the extent to which the altered profiles of miRNA expression influence viral replication and latency, as well as the efficiency of host defenses, may be useful for understanding the basis of the HIV-1-related alterations in cellular physiology and immunologic control. To this end, three patient groups were enrolled. One group consisted of subjects who were classified as M-CM-)lite control long-term non-progressors (M-CM-)LTNP). A second study group was HIV-1-positive subjects, who were antiretroviral therapy naive. A third group was multiply exposed to HIV-1, but uninfected (MEU). This study allowed us to investigate the existence of a CD4+T- lymphocytes miRNAs signature able to discriminate among different stages of HIV-1 infection, and to evaluate whether the exposure to HIV-1 antigen is sufficient to change the miRNA profile. MicroRNAs inhibit HIV-1 expression by either modulating host innate immunity or by directly interfering with viral mRNAs. We evaluated the expression of 377 miRNAs in CD4+ T cells from HIV-1 M-CM-)lite LTNP (M-CM-)LTNP), naive and multiply exposed uninfected individuals (MEU) and we observed that the M-CM-)LTNP patients clustered with naive, whereas all MEU subjects grouped together. The discriminatory power of miRNAs showed that 21 miRNAs significantly differentiated M-CM-)LTNP from MEU and 23 miRNAs distinguished naive from MEU, while only 1 (miR-155) discriminated M-CM-)LTNP from naive. We proposed that miRNA expression may discriminate between HIV-1 infected and exposed but negative individuals. Analysis of miRNAs expression after exposure of healthy CD4+T cells to gp120 in vitro confirmed our hypothesis that a miRNA profile could be the result not only of a productive infection, but also of the exposure to HIV-1 products that leave a signature in immune cells. The comparison of normalized Dicer and Drosha expression in ex vivo and in vitro conditions revealed that these enzymes did not affect the change of miRNA profiles, supporting the existence of a Dicer-independent biogenesis pathway. We have compared miRNA profiles of CD4+ T-lymphocytes from 18 HIV-1-exposed subjects with healthy CD4+ T-lymphocytes following exposure to gp120 using a RT-qPCR assay.
Project description:Assessment of the extent to which the altered profiles of miRNA expression influence viral replication and latency, as well as the efficiency of host defenses, may be useful for understanding the basis of the HIV-1-related alterations in cellular physiology and immunologic control. To this end, three patient groups were enrolled. One group consisted of subjects who were classified as élite control long-term non-progressors (éLTNP). A second study group was HIV-1-positive subjects, who were antiretroviral therapy naive. A third group was multiply exposed to HIV-1, but uninfected (MEU). This study allowed us to investigate the existence of a CD4+T- lymphocytes miRNAs signature able to discriminate among different stages of HIV-1 infection, and to evaluate whether the exposure to HIV-1 antigen is sufficient to change the miRNA profile. MicroRNAs inhibit HIV-1 expression by either modulating host innate immunity or by directly interfering with viral mRNAs. We evaluated the expression of 377 miRNAs in CD4+ T cells from HIV-1 élite LTNP (éLTNP), naive and multiply exposed uninfected individuals (MEU) and we observed that the éLTNP patients clustered with naive, whereas all MEU subjects grouped together. The discriminatory power of miRNAs showed that 21 miRNAs significantly differentiated éLTNP from MEU and 23 miRNAs distinguished naive from MEU, while only 1 (miR-155) discriminated éLTNP from naive. We proposed that miRNA expression may discriminate between HIV-1 infected and exposed but negative individuals. Analysis of miRNAs expression after exposure of healthy CD4+T cells to gp120 in vitro confirmed our hypothesis that a miRNA profile could be the result not only of a productive infection, but also of the exposure to HIV-1 products that leave a signature in immune cells. The comparison of normalized Dicer and Drosha expression in ex vivo and in vitro conditions revealed that these enzymes did not affect the change of miRNA profiles, supporting the existence of a Dicer-independent biogenesis pathway.
Project description:We have investigated the dynamic host response to HIV-1 infection by systematically measuring transcriptome, proteome and phosphoproteome expression changes in infected and uninfected SupT1 CD4+ T cells at 5 time-points throughout the HIV-1 replication cycle (from 2h to 24h).
Project description:CD8+ and CD4+ T cells from HIV infected patients with HIV-RNA viremia of >50 copies/ml and CD8+ and CD4+ T cells from healthy controls were isolated by negative selection (Miltenyi Biotech, Auburn, CA). Cell sorting of the CD4 and CD8 T cell subsets were performed based on surface staining of CD3+CD8+ or CD3+CD4+ and na ve CD45RA+CD27+CD127highHLA-DRlow, and CD3+CD8+ or CD3+CD4+ and memory CD45RA-CD27+CD127highHLA-DRlow. Sorted cell populations were spun down and stored as dry pellets at -80M-BM-0C. Samples analyzed by transcript levels of genes related to cytokine signaling were determined by the JAK/STAT Signaling Pathway microarray. CD8+ and CD4+ T cells from HIV infected patients with HIV-RNA viremia of >50 copies/ml and CD8+ and CD4+ T cells from healthy controls were isolated by negative selection (Miltenyi Biotech, Auburn, CA). Cell sorting of the CD4 and CD8 T cell subsets were performed based on surface staining of CD3+CD8+ or CD3+CD4+ and naM-CM-/ve CD45RA+CD27+CD127highHLA-DRlow, and CD3+CD8+ or CD3+CD4+ and memory CD45RA-CD27+CD127highHLA-DRlow. Sorted cell populations were spun down and stored as dry pellets at -80M-BM-0C. Samples analyzed by transcript levels of genes related to cytokine signaling were determined by the JAK/STAT Signaling Pathway microarray (http://www.sabiosciences.com/rt_pcr_product/HTML/PAHS-039A.html, SABiosciences Frederick, MD). Briefly, total RNA was harvested from each individual T cell subset from each patient or healthy control and contaminating DNA was digested with DNase. Messenger RNA was converted to cDNA and loaded onto PCR array plates for quantitative real-time PCR. Quantification of transcript levels was determined by normalizing to 5 housekeeping genes from each individual sample. Relative gene expression levels for each subset were averaged and compared between cell populations from the patient group and healthy controls. Because of the multiple comparisons only p values M-bM-^IM-$ 0.01 were considered significant.
Project description:Background. Previous epigenome-wide association studies have shown that HIV infection can disrupt the host DNA methylation landscape. However, it remains unclear how antiretroviral therapy (ART) affects the HIV-induced epigenetic modifications. Methods. 184 individuals with HIV from the NEAT001/ANRS143 clinical trial (with pre-ART and post-ART samples [96 weeks of follow-up]) and 44 age-and-sex matched individuals without HIV were included. We compared genome-wide DNA methylation profiles in whole blood between groups adjusting for age, sex, batch effects, and leucocyte type heterogeneity. Findings. We identified 430 differentially methylated positions (DMPs) between HIV+ pre-ART individuals and HIV-uninfected controls. In participants with HIV, ART initiation modified the DNA methylation levels at 845 CpG positions and restored 49.3% of the changes found between HIV+ pre-ART and HIV-uninfected individuals. We only found 15 DMPs when comparing DNA methylation profiles between HIV+ post-ART individuals and participants without HIV. The Gene Ontology enrichment analysis of DMPs associated with untreated HIV infection revealed an enrichment in biological processes regulating the immune system and antiviral responses. In participants with untreated HIV infection, DNA methylation levels at top HIV-related DMPs were associated with CD4/CD8 ratios and viral loads. Changes in DNA methylation levels after ART initiation were weakly correlated with changes in CD4+ cell counts and the CD4/CD8 ratio. Interpretation. Control of HIV viraemia after 96 weeks of ART initiation restores most of the host DNA methylation changes that occurred before antiretroviral treatment of HIV infection.
Project description:We compared transcriptional profiles of CD4+ and CD8+ T lymphocytes from HIV infected individuals before and 1 year after interruption of antiretroviral therapy (ART).
Project description:In perinatal HIV infection, early ART initiation is recommended but questions remain regarding infant immune responses to HIV and impact of HIV on immune development. Using single cell transcriptional and phenotypic analysis we evaluated the T cell compartment at pre-ART initiation of infants with perinatally acquired HIV from Maputo, Mozambique (TARA cohort). CD8+ T cell maturation subsets exhibited altered distribution in HIV exposed infected (HEI) infants relative to control infants (HIV exposed uninfected) with reduced naïve, increased effectors, higher frequencies of activated T cells, and lower frequencies of cells with markers of self-renewal. Additionally, a cluster of CD8+ T cells identified in HEI displayed gene profiles consistent with cytotoxic T lymphocytes (CTL) and showed evidence for hyper expansion. Longitudinal phenotypic analysis revealed accelerated maturation of CD8+ T cells was maintained in HEI despite viral control. The results point to a HIV directed immune response that is likely to influence reservoir establishment.