Fig 1: Memory populations in SARS-CoV-2 spike-specific CD4+ T cells following BNT162b2 mRNA vaccination in naive subjects and individuals recovered from COVID-19(A and B) PBMCs were labeled with CFSE and stimulated with SARS-CoV-2 spike peptide pool for 5 days. CD4+ T cells were classified according to their proliferative response in samples 2 and 3. Memory subpopulations were analyzed (naive; CM, central memory; EMRA, effector memory cells re-expressing CD45RA; EM, effector memory). Frequencies (A) and mean distribution (B) of memory populations in proliferative (?; CFSEdim) and non-proliferative (?; CFSEbright) CD4+ T cells.(C) Proliferative (CFSEdim) versus non-proliferative (CFSEbright) ratio of CD4+ T cell memory populations in samples 2 (?) and 3 (?).(D) Frequency of IL-2+ cells in gated naive, CM, EMRA, and EM CD4+ T cells stimulated or not with SARS-CoV-2 spike peptide pool.(E) Increment of frequencies of IL-2+ cells comparing SARS-CoV-2 spike pool-stimulated and non-stimulated CD4+ T cell subpopulations.(A, C–E) Each dot represents an individual. (A, C, and D) Paired Student’s t test (ns, not significant; *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001). (E) Mann Whitney test (ns, not significant; *p < 0.05). n = 16 naive, n = 11 recovered from COVID-19.See also Figure S3B, S3C, and S5.
Fig 2: Differential SARS-CoV-2 spike-specific humoral and cellular responses between naive subjects and individuals recovered from COVID-19 after vaccination(A) Heatmap analysis of main humoral and cellular variables of SARS-CoV-2 spike-specific responses. This algorithm showed differential clustering distribution between naive subjects and individuals recovered from COVID-19.(B) Spearman correlation matrix heatmap of the main humoral and cellular variables of SARS-CoV-2 spike-specific responses grouped by functionality.(C) Spearman correlations between different SARS-CoV-2 spike-specific humoral and cellular responses. Top: ratio of CD4+ T cells proliferation versus titers of RBD IgG antibodies; middle: ratio of CD4+ T cells proliferation versus S1 IgG antibodies; bottom: IL-2 production in supernatants versus titers of neutralizing antibodies.R, Spearman’s rank correlation coefficient. p value in Spearman correlation test (ns, not significant; *p < 0.05). White dots represent naive individuals; red dots represent subjects recovered from COVID-19. n = 16 naive, n = 11 recovered from COVID-19.
Fig 3: SARS-CoV-2 spike-specific cellular ex vivo response following BNT162b2 mRNA vaccination in naive subjects and individuals recovered from COVID-19(A) Experimental design of the T cell cellular response ex vivo in PBMCs in samples 2 and 3 after stimulation with SARS-CoV-2 spike peptide pool.(B) CCL-2 and CXCL10 chemokines production.(C) IL-2, IL-4, IL-6, IL-10, TNFa, and IFN? production.(D) Percentage of IL-2+ cells in CD4+ (left panel) and CD8+ (right panel) T cells.(E) Increment of IL-2+ cells comparing SARS-CoV-2 spike peptide pool-stimulated and non-stimulated CD4+ and CD8+ T cells.(F) Frequency of proliferative (CFSEdim) CD4+ and CD8+ T cells.(G) Increment of proliferation comparing SARS-CoV-2 spike peptide pool-stimulated and non-stimulated CD4+ and CD8T cells.(B–G) Each dot represents an individual. (B–D and F) Paired Student’s t test (ns, not significant; *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001). (E and G) Mann Whitney test (ns, not significant; *p < 0.05; **p < 0.01). n = 16 naive, n = 11 recovered from COVID-19.See also Figure S4.
Fig 4: Plasma monocyte chemoattractant (MCP-1) concentration (A) and relative circulating natural killer (NK) cell counts (B) pre-exercise (Pre) and 0 (Post), 24, 48 and 72 h after the bench-stepping exercise. Data are means ± SEM. * denotes significant difference (p < 0.05) from Pre values. † denotes significant difference (p < 0.05) from Post values. GSM, Greenshell™ mussel. ^ indicates significant difference from placebo (p < 0.05).
Fig 5: Functional status of the newly differentiated cells from in vitro U937 cells was investigated in terms of mRNA abundance of the cytokine gene produced by the new cells. The expression amount of each cytokine gene was normalized against ACTB, a house-keeping gene; in the cytokine panel, IL-6, TNF-α, IFN-γ, and GMCSF are M1-macrophage-specific, and IL-4 and IL-10 are M2-specific cytokines; * p < 0.05, ** p < 0.01, *** p < 0.001, as compared to UC.
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