Extended Data Fig. 8: Circulating PD1+ T cells and subclinical atherosclerosis.
(a-g) Blood from 65–72-year-old individuals (n = 675) recruited from the general population that had previously been enrolled in the Swedish Cardiopulmonary Bioimage Study (SCAPIS) was sampled. (a) Study population characteristics, metabolic profile, and levels of PD1+ T cells by flow cytometry. (b-e) Peripheral blood mononuclear cells were isolated and cultured in complete media with anti-CD28/anti-CD49d antibodies for 24 h and flow cytometry was performed. (b) Gating strategy to identify live CD4 and CD8 T cells. (c) Fluorescent minus-one (FMO) staining control for anti-PD1-BV421. (d-e) Correlation of %PD1+ T cells before and after 24-hour cell culture. (f) Correlation analysis of PD1 subsets and metabolic parameters (Spearman’s rank correlation co-efficient). (g) Beta co-efficients derived from linear regression analysis of association between PD1 subsets and Duke score (n = 611), unadjusted or adjusted for age and sex (Model A) or Model A + glucose, HDL, LDL, total cholesterol and triglycerides. Bars denote 95% CI and data was log transformed. (h-i) PBMCs cultured for 24 h in wells containing either anti-CD3 and co-stimulatory antibodies (anti-CD28/anti-CD49d) or stimulating cytokines (IL-12, IL-15, and IL-18) to study IFN-γ-production of human PBMCs were assessed by intracellular flow cytometry. (h) Negative controls (medium + Brefeldin A only). (i) Frequency of PD1 expression of CD4 (top-down, ****p = 5.2E-7, 1.7E-5) and CD8 T cells (****p = 1.2E-6, **p = 0.0067) in response to the different stimulation conditions. (d-f) Analyzed with two-sided Spearman’s rank correlation coefficients. (i) Bars denote median, CD4 T cells analyzed with two-sided one-way ANOVA and CD8 T cells analyzed with two-sided Mann-Whitney U test.
