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Cytolytic CD4+ T cells express high levels of T-bet and Eomes in blood.

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posted on 2018-04-13, 17:45 authored by Marcus Buggert, Son Nguyen, Laura M. McLane, Maria Steblyanko, Nadia Anikeeva, Dominic Paquin-Proulx, Perla M. Del Rio Estrada, Yuria Ablanedo-Terrazas, Kajsa Noyan, Morgan A. Reuter, Korey Demers, Johan K. Sandberg, Michael A. Eller, Hendrik Streeck, Marianne Jansson, Piotr Nowak, Anders Sönnerborg, David H. Canaday, Ali Naji, E. John Wherry, Merlin L. Robb, Steven G. Deeks, Gustavo Reyes-Teran, Yuri Sykulev, Annika C. Karlsson, Michael R. Betts

(A) Representative flow cytometry plots of Granzyme B and perforin expression in CD4+ T cells for an HIV-infected and–uninfected subject. The distribution of Granzyme B+perforin+ (red) and Granzyme B-perforin- (blue) CD4+ T cells are shown for T-bet and Eomes expression. (B) Frequency of perforin+ CD4+ T cells within the T-bethiEomes+ and T-betdim/- population (left) and T-bethiEomes+ within the perforin+ or perforin- population for HIV-infected and–uninfected subjects. (C) Correlation between the frequency of perforin+ and T-bethi CD4+ T cells. (D) Imagestream analysis on T-bethi and T-betdim CD4+ T cells. Overlays of fluorescent channels for DAPI (nuclear) and T-bet, showing where in the cells T-bet are localized. The frequency of nuclear, nuclear/cytoplasmic and cytoplasmic localization for T-bethi and T-betdim CD4+ T cells are shown in the before-after graphs. (E) tSNE plots based on 30,000 live CD4+ T cells that were merged from three HIV-uninfected subjects with detectable cytolytic CD4+ T cells. The tSNE clustering is based on CD45RO, CD27, CCR7, T-bet, Eomes, Granzyme A, Granzyme B and perforin expression intensity. The red gate indicates the identified “effector” cluster with overlapped expression of cytolytic markers as well as T-bet and Eomes. (F) Flow plots of MIP-1α production using media (NC) and aCD3-CD28 stimulations for T-bethi and Eomes+ CD4+ T cells, as well as correlation between the frequency of T-bethiEomes+ and MIP-1α+ CD4+ T cells following aCD3-CD28 stimulations. Median and IQR are shown for all scatter plots and Mann-Whitney tests were performed to compare differences between groups; ***P < 0.001. A non-parametric Spearman test was used for the correlations analysis. All data are derived from the North-American cohort.

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