Figure 2. CD8⁺ T-cell profiles in convalescent coronavirus disease-19 (COVID-19) patients at 3 months post admission.

(A–D) Percentage of CD8⁺ T-cells within the CD3⁺ gate (A), absolute number of CD8⁺ T-cells (cells/mm³) (B), and percentages of CD8⁺ T-cells co-expressing the activation markers HLA-DR/CD38 (C) or granzyme B (D, shown as mean fluorescence intensity [MFI]) are shown in mild, moderate, and severe patients. (E) Percentages of naïve (CCR7⁺ CD45RA⁺), T central memory (T_CM, CCR7⁺ CD45RA^-), T effector memory (T_EM, CCR7^- CD45RA^-), and T effector memory RA re-expressing (T_EMRA, CCR7^- CD45RA⁺) CD8⁺ T-cells in patients with mild, moderate, and severe disease. (F) Flow cytometry plot with a representative staining from a mild, moderate, and severe patient (overlaid and shown respectively in blue, black, and red) of HLA-DR and CD38 expression in CD8⁺ T_EM cells. (G) Percentages of activated HLA-DR⁺ CD38⁺ CD8⁺ T-cells within naïve, T_CM, T_EM, and T_EMRA cells. (H) Flow cytometry plot with a representative staining from a mild, moderate, and severe patient of HLA-DR and granzyme B (GrzmB) expression in CD8⁺ T_EM cells. (I) Percentages of proliferating HLA-DR⁺ GrzmB⁺ CD8⁺ T-cells within naïve, T_CM, T_EM, and T_EMRA cells. (J) Unsupervised uniform manifold approximation and projection (UMAP) analysis showing the FlowSOM clusters in mild (N=17), moderate (N=25), and severe (N=14) patients. Plots are gated on CD8⁺ T-cells. (K) Heatmap with MFI levels for each analysed marker within the FlowSOM populations. (L) Summary of percentage of CD8⁺ T-cells within the indicated FlowSOM populations in mild, moderate, and severe patients. Data in the graphs are shown as mean ± SEM. Statistics were calculated by one-way ANOVA (Kruskal-Wallis test) with Dunn’s correction for multiple testing.

Figure 2—figure supplement 1. Resolution of T-cell activation at 12 months.

(A–E) CD4⁺ T-cells in convalescent coronavirus disease-19 (COVID-19) patients at 3 and 12 months. Shown are the percentages of CD4⁺ T-cells within the CD3⁺ gate (A) and the percentages of CD4⁺ T-cells that are Ki67⁺ CD38⁺ (B), CXCR3⁺ (C), HLA-DR⁺CD38⁺ (D) and granzyme B⁺ (shown as mean fluorescence intensity [MFI]) (E).(F–J) CD8⁺ T-cells in convalescent COVID-19 patients at 3 and 12 months. Shown are the percentages of CD8⁺ T-cells within the CD3⁺ gate (F) and the percentages of CD8⁺ T-cells that are Ki67⁺ CD38⁺ (G), CXCR3⁺ (H), HLA-DR⁺CD38⁺ (I), and granzyme B⁺ (shown as MFI) (J). (K) Unsupervised uniform manifold approximation and projection (UMAP) analysis showing the density plot of cell distribution in mild (N=17), moderate (N=25), and severe (N=14) patients at 3 months and in matched mild (N=8), moderate (N=17), and severe (N=8) patients at 12 months post infection. Data in A–J are shown as mean ± SEM. Statistics in A–J were calculated by Mann-Whitney t-test.

Figure 2—figure supplement 2. Immune cell populations in coronavirus disease-19 (COVID-19) patients at 3 months post admission.

(A–F) Shown are the percentages of CD56^dim (A), CD56^bright NK cells (B), Ki67⁺CD38⁺ CD56^dim (C), and Ki67⁺CD38⁺CD56^bright NK cells (D). Granzyme B expression on CD56^dim (E) and CD56^bright (F) NK cells is shown as mean florescence intensity (MFI). (G–K) Shown are the percentages of classical (G: CD14⁺CD16^-), intermediate (H: CD14⁺CD16⁺), and non-classical (I: CD14^-CD16⁺) monocytes, activated CD14⁺CD80⁺CD86⁺ (J) and CD14⁺CD86⁺CD163⁺ cells (K). (L–N) TCR-γδ T-cells: frequencies of TCR-γδ T-cells (L), activated HLA-DR⁺CD38⁺ (M), and activated/proliferating Ki67⁺CD38⁺ TCR-γδ T-cells (N).(O–R) Shown are the percentages of CD3^-CD19⁺ B cells (O), activated CD80⁺ (P), and HLA-DR⁺CD38⁺ B cells (Q) and proliferating Ki67⁺CD38⁺ B cells (R). Data are shown as mean ± SD in patients with mild (N=16), moderate (N=21) and severe (N=14) disease. Statistics were by calculated one-way ANOVA test (Kruskal-Wallis test) with Dunn’s correction for multiple testing.