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. 2017 Jun 8;152(1):89–101. doi: 10.1111/imm.12751

Figure 2.

Figure 2

Cord blood myeloid‐derived suppressor cells (CB‐MDSC) inhibit T helper type 1 (Th1) cells in a cell‐contact‐dependent manner. CB‐MDSC were enriched from cord blood mononuclear cells (CBMC) and added to peripheral blood mononuclear cells (PBMC) or CD3+ T cells isolated from a healthy adult control either with direct cell/cell contact or in transwells. After 5 days of culture, surface staining was performed and cells were analysed by flow cytometry. (a) Scatter plot with bars shows the percentage of CXCR3+ CCR4− CCR6− Th1 cells without addition of CB‐MDSC (white bar), with addition of CB‐MDSC in direct cell/cell‐contact (grey bar) or separated by transwells (checked bar). Bars represent pooled data from 11 independent experiments and each point represents an individual sample. ns = not significant; Wilcoxon matched‐pairs signed rank test. (b) Scatter plot with bars shows the percentage of CXCR3+ CCR4− CCR6− Th1 cells after culture of CD3+ T cells without (white bar) or with CB‐MDSC (grey bar). n = 7, P = 0·02, Wilcoxon matched‐pairs signed rank test. (c, d) Scatter plots with bars shows inhibition of Th1 cells by CB‐MDSC alone (white bar) or under addition of inhibitors for ArgI (nor‐NOHA), ROS (catalase), iNOS (L‐NMMA) and IDO (1‐MT) in ascending grey scales. (c) Percentages of Th1 cells without addition of CB‐MDSC (blank bars) and with addition of CB‐MDSC (checked bars). n = 5 or n = 6; *P < 0·05, ns = not significant; Wilcoxon matched‐pairs signed rank test. (d) Inhibition of Th1 cells was determined as ratio between percentages of Th1 cells with addition of CB‐MDSC and percentages of Th1 cells without addition of CB‐MDSC. n = 5 or n = 6; ns = not significant in comparison to without (w/o) inhibitors; Wilcoxon matched‐pairs signed rank test. (e, f) Scatter plots with bars show the percentage of CXCR3+ CCR4− CCR6− Th1 cells without addition of CB‐MDSC (white bars), with addition of CB‐MDSC (grey bars) and with addition of CB‐MDSC and blocking antibodies (dashed bars) against transforming growth factor‐β (TGF‐β) (e) and programmed death 1 (PD‐1) (f). n = 5 or n = 6, ns = not significant; Wilcoxon matched‐pairs signed rank test. (g–i) Bar graphs show the percentage of Annexin V+ cells of all CD4+ T cells (g) and of CXCR3+ CD4+ (h) and CCR4+ CD4+ T cells (i) without the addition of CB‐MDSC (white bars) and with the addition of CD‐MDSC (grey bars). n = 6, *P = 0·02, ns = not significant; Wilcoxon matched‐pairs signed rank test. (j) Scatter plot with bars shows the percentage of CXCR3+ CCR4− CCR6− Th1 cells without addition of CB‐MDSC (white bars), with addition of CB‐MDSC (grey bars) and with addition of CB‐MDSC and blocking antibody (dashed bars) against CD95L, n = 5.