Ehretia dicksonii Hance var. japonica Nakai

原著和名: マルバチシャノキ科名: ムラサキ科 = Boraginaceae採集地: 千葉県 安房郡 天津小湊町 実入 (安房 天津小湊町 実入)採集日: 1966/6/26採集者: 萩庭丈壽整理番号: JH001226国立科学博物館整理番号: TNS-VS-95122

MOESM2 of Identification of novel HIV-1 dependency factors in primary CCR4+CCR6+Th17 cells via a genome-wide transcriptional approach

Additional file 2: Table S2. The complete list of genes down regulated in Th17 versus Th1 subsets are included and classified based on p-values < 0.05 and FC cut-off 1.3

MOESM6 of Identification of novel HIV-1 dependency factors in primary CCR4+CCR6+Th17 cells via a genome-wide transcriptional approach

Additional file 6: Figure S3. A meta-analysis for transcripts included in the NCBI HIV Interaction data base that were identified as top differentially expressed between Th17 and Th1 subsets

CD160 and PD-1 Co-Expression on HIV-Specific CD8 T Cells Defines a Subset with Advanced Dysfunction

<div><p>Chronic viral infections lead to persistent CD8 T cell activation and functional exhaustion. Expression of programmed cell death-1 (PD-1) has been associated to CD8 T cell dysfunction in HIV infection. Herein we report that another negative regulator of T cell activation, CD160, was also upregulated on HIV-specific CD8 T lymphocytes mostly during the chronic phase of infection. CD8 T cells that expressed CD160 or PD-1 were still functional whereas co-expression of CD160 and PD-1 on CD8 T cells defined a novel subset with all the characteristics of functionally exhausted T cells. Blocking the interaction of CD160 with HVEM, its natural ligand, increased HIV-specific CD8 T cell proliferation and cytokine production. Transcriptional profiling showed that CD160⁻PD-1⁺CD8 T cells encompassed a subset of CD8⁺ T cells with activated transcriptional programs, while CD160⁺PD-1⁺ T cells encompassed primarily CD8⁺ T cells with an exhausted phenotype. The transcriptional profile of CD160⁺PD-1⁺ T cells showed the downregulation of the NFκB transcriptional node and the upregulation of several inhibitors of T cell survival and function. Overall, we show that CD160 and PD-1 expressing subsets allow differentiating between activated and exhausted CD8 T cells further reinforcing the notion that restoration of function will require multipronged approaches that target several negative regulators.</p> </div

Changes in gene expression in DC-induced latently infected CD4⁺ T cells.

<p>(<b>A</b>) Fold change scatterplot comparing gene expression between HIV T (+DC) (CD4⁺ T cells cultured with DC and HIV), and Mock T (+DC) (CD4⁺ T cells cultured with only DC) relative to their controls, HIV T (CD4⁺ T cells cultured with HIV) and Mock T (CD4⁺ T cells cultured in media alone) respectively. The solid line indicates absolute 2-fold change. (<b>B</b> and <b>C</b>) Top two gene interaction networks as ranked by Ingenuity Pathway Analysis. The networks were built from the list of differentially expressed genes induced by HIV T (+DC), relative to Mock T (+DC) after subtracting HIV T and Mock T from each group respectively. Genes highlighted in red were up-regulated and those in blue were down-regulated. The different node shapes indicate genes in different functional categories according to the legend. The interactions between the different nodes are shown as solid (direct interaction) or dashed (indirect interaction) lines (edges). (<b>D</b>) Fold change in gene expression values for selected genes from the Illumina BeadArrays plotted against Real-Time PCR (qPCR) deltaCt values for each target gene. PCR targets were mapped to BeadArray probes by matching the official gene symbols.</p

Soluble factors in DC-induced HIV latency.

<p>(<b>A</b>) Resting CD4⁺ T cells were cultured alone (light grey) or with sorted pDC (grey) or mDC (dark grey). At day 5 post-infection, cytokines and chemokines were quantified in culture supernatants using cytometric bead arrays, n = 3. *<i>P</i>&lt;0.05; **<i>P</i>&lt;0.01 (paired t-test). (<b>B</b>) Latent infection was quantified in eFluor670^hiEGFP⁻ resting CD4⁺ T cells that were cultured either alone or with sorted pDC in the presence of media alone (light grey), anti-IgG (grey) or anti-IFN-alpha (dark grey) following stimulation with anti-CD3/CD28 in the presence of L8, n = 5. (<b>C</b>) Resting CD4⁺ T cells were co-cultured with mDC with (grey) or without (light grey) the addition of equal numbers of pDC. Productive infection was determined at day 5 post-infection. Latent infection was determined in sorted eFluor670^hiEGFP⁻ CD4⁺ T cells following stimulation with anti-CD3/CD28 in the presence of L8, n = 5. (<b>D</b>) Latent infection was quantified in eFluor670^hiEGFP⁻ resting memory CD4⁺ T cells that were cultured either alone or with sorted mDC in the presence of media alone (light grey), anti-IgG (grey) or neutralising antibodies (dark grey) to IL-6, IL-10-receptor, CXCR3 or CCL19, n = 5. (<b>E</b>) eFluor670-labelled resting CD4⁺ T cells were cultured either alone (light grey) or with blood mDC (dark grey). Virus was added to (<b>i</b>) CD4⁺ T cells cultured alone; (<b>ii</b>) CD4⁺ T cells co-cultured with mDC; (<b>iii</b>) CD4⁺ T cells cultured with mDC in the presence of a 0.4 µm membrane transwell and latency determined at day 5 post-infection, n = 5. (<b>F</b>) eFluor670-labelled resting CD4⁺ T cells were cultured either (<b>i</b>) alone (light grey) or (<b>ii</b>) with blood mDC (dark grey) and infected. (<b>iii</b>) Following 24 hours, supernatant from infected mDC-T cell co-cultures was added to uninfected resting CD4⁺ T cells and these cells were then infected, n = 3. Columns represent the median of 3–5 donors and error bars indicate the interquartile range. *<i>P</i>&lt;0.05; **<i>P</i>&lt;0.01 (Wilcoxon signed-rank test).</p

The frequency of HIV-specific CD8 T cells with a CD160⁺PD-1⁺ phenotype increases with disease progression.

<p>(A) Representative flow cytometry plots of CMV and HIV-specific CD8 T cells (A*02 CMV and A*03 Gag) detected during AHI (<3 months) and CHI (>6 months) within the same individual. Frequencies of 4 distinct CD8 T cell subsets expressing CD160 and/or PD-1 were measured at both time points. Figures represent the frequency of CD160 and/or PD-1 expressing subsets over time for (B) HIV and (C) CMV/EBV-specific responses in 4 HIV-infected subjects. <i>P</i>-values were determined by the Wilcoxon matched pairs test.</p

Blocking the interaction between CD160 and HVEM enhances CMV and HIV-specific CD8 T cell proliferation and cytokine production.

<p>HIV-infected individuals(n = 11) were stimulated with HLA-restricted CMV and HIV peptides in the presence of blocking antibodies to HVEM and/or PD-L1. Dying cells were eliminated with an amine-reactive viability dye and PBMCs were stained at day 6 with HLA class I matched-tetramers and mAbs to CD3 and CD8. (A) Representative flow cytometry plots of an HIV-infected patient stimulated for 6 days with a CMV and HIV peptide in the presence of isotype, αPD-L1 and/or αHVEM blocking antibodies. Scatter plots represent the median fold increase in (B) CMV and (C) HIV-specific proliferation (Tetramer⁺/CFSE_low) compared to the isotype control. Each dot represents a CMV or HIV tetramer-specific response. <i>P</i>-values were determined by the Wilcoxon matched pairs test.</p

Summary of clinical characteristics.

<p>Summary of clinical characteristics.</p

CD160⁺PD-1⁺CD8 T cells represent a distinct subset with a unique transcriptional profile.

<p>PBMCs were stained with 7AAD (cell viability dye), αCD3, αCD8, αPD-1, αCD160 and total CD8 T cells were sorted using a FACS ARIA based on CD160 and PD-1 expression. (A) Heatmap illustrating the differentially expressed genes (39 genes; p<0.05) between DP and SP-PD-1 sorted subsets. (B) Network analysis of significantly inferred genes and predicted targets. Node colors indicate fold change of gene expression between ex-vivo sorted CD160⁺PD-1⁺ and CD160⁻PD-1⁺ CD8 T cells sorted from 4 HIV viremic patients. The different shapes indicate genes in the different functional categories (see legend). (C) Histogram and scatter plot showing the MFI of KIR2DL2 and KIR2DL3 expression on DP and SP-PD-1 subsets in 6 HIV viremic patients.</p