Figure 4.

Total blood was stained with antibodies against CD45, CD3, CD4, CD8, CD14, CD16, CD33, CD38 and HLADR and in a second staining antibodies used were anti-CD3, CD4, CD8, CD45RA, CD27, CD28, CD31 and CCR7. (A) Graphs indicate the percentage of total CD3⁺, CD3⁺CD4⁺ and CD3⁺CD8⁺ T cells. (B) FACS plots depict the gating strategy to measure the frequency of CD4⁺ and CD8⁺ T cells expressing HLADR in three representative patients (asymptomatic, mild and severe). Percentage of CD4⁺HLADR⁺ or CD8⁺HLADR⁺ T cells is shown in the relative graphs (at the first sample collected after diagnosis). (C) Pseudocolor plots show CD31 expression in CD4⁺ and CD8⁺ naïve T cells (CD3⁺CCR7⁺CD45RA⁺). CD31⁺ are recent thymic emigrants and CD31⁻ are naïve T cells. Graphs show the percentage of CD31⁺ and CD31⁻ T cells in all patients (at the first sample collected after diagnosis). (D) FACS plots show CD4⁺ or CD8⁺ central memory (CD3⁺CCR7⁺CD45RA⁻), effector memory (CD3⁺CCR7⁻CD45RA⁻) T cells and TEMRA (CD3⁺CCR7⁻CD45RA⁺) T cells. Scatter plots depict the percentage of CD4⁺ and CD8⁺ TEMRA (at the first sample collected after diagnosis). Median is shown as midline. Statistical significances were determined using unpaired, two-tailed Mann–Whitney U-tests. *p ≤ 0.05, **p < 0.01, ***p < 0.001.