Neutrophilic myeloid-derived suppressor cells in cord blood modulate innate and adaptive immune responses

Abstract

Neonates show an impaired anti-microbial host defence, but the underlying immune mechanisms are not understood fully. Myeloid-derived suppressor cells (MDSCs) represent an innate immune cell subset characterized by their capacity to suppress T cell immunity. In this study we demonstrate that a distinct MDSC subset with a neutrophilic/granulocytic phenotype (Gr-MDSCs) is highly increased in cord blood compared to peripheral blood of children and adults. Functionally, cord blood isolated Gr-MDSCs suppressed T cell proliferation efficiently as well as T helper type 1 (Th1), Th2 and Th17 cytokine secretion. Beyond T cells, cord blood Gr-MDSCs controlled natural killer (NK) cell cytotoxicity in a cell contact-dependent manner. These studies establish neutrophilic Gr-MDSCs as a novel immunosuppressive cell subset that controls innate (NK) and adaptive (T cell) immune responses in neonates. Increased MDSC activity in cord blood might serve as key fetomaternal immunosuppressive mechanism impairing neonatal host defence. Gr-MDSCs in cord blood might therefore represent a therapeutic target in neonatal infections.

Keywords: Gr-MDSC; MDSC; NK cells; Th17; cord blood; myeloid-derived suppressor cells; neonatal; neutrophilic.

Fig. 1

Characterization and quantification of human granulocytic/neutrophilic myeloid-derived suppressor cells (Gr-MDSCs) in cord blood and different age groups. (a) Gr-MDSCs are characterized in the forward-/side-scatter (FSC/SCC) area as a granulocytic cell population in the peripheral blood mononuclear cells (PBMC) fraction (low-density neutrophils) (R1), as published previously [12,14,42]. This population was distinct from lymphocytes, monocytes, erythrocytes or debris. Representative scatter-plots are shown. (b) Within R1, Gr-MDSCs were identified as CD66b^highCD33^highinterleukin IL-4Rα^interhuman leucocyte antigen D-related (HLA-DR)^neg expressing neutrophilic MDSC population. Representative histograms are shown. (c) Log scale illustration of human Gr-MDSC numbers in cord blood and different age groups of healthy children and adults.

Fig. 2

Cord blood granulocytic/neutrophilic myeloid-derived suppressor cells (CB-Gr-MDSCs) suppress T cell proliferation. The T cell suppressive capacities of cord blood Gr-MDSCs are shown. For proliferation assays, peripheral blood mononuclear cells (PBMC) were stimulated with interleukin (IL)-2 (100 U/ml) and muromonab-CD3 (OKT3) (1 μg/ml). The suppressive effects of CD66b⁺-magnetic-activated cell sorting (MACS)-isolated Gr-MDSCs or as a control CD66b⁺-MACS-isolated conventional high-density polymorphonuclear leucocytes (PMNs) were analysed on CD4⁺ and CD8⁺ T cell subsets using the carboxyfluorescein succinimidyl ester (CFSE) polyclonal proliferation assay. A minimum of three independent CFSE assays was conducted. Asterisks indicate significant differences in proliferation indices under addition of Gr-MDSCs compared to target cells only (a–e). (a) Representative CFSE histograms. (b) Effect of cord blood and adult Gr-MDSCs on CD4⁺ T cell proliferation. (c) Effect of cord blood and adult Gr-MDSCs on CD8⁺ T cell proliferation. (d) Transwell assays indicating cell contact-dependent effects of Gr-MDSCs. (e) No effect of conventional cord blood or adult PMNs on lymphocyte proliferation. CB: cord blood; TW: Transwell.

Fig. 3

Cord blood granulocytic/neutrophilic myeloid-derived suppressor cells (Gr-MDSCs) inhibit T helper cell cytokine secretion. Cord blood Gr-MDSCs were added to interleukin (IL)-2 (100 U/ml) and muromonab-CD3 (OKT3) (1 μg/ml)-stimulated peripheral blood mononuclear cells (PBMC) in a 1 : 2 ratio and the effect on interferon (IFN)-γ, IL-5 and IL-17 concentrations in supernatants was measured using a Bioplex system.

Fig. 4

Cord blood granulocytic/neutrophilic myeloid-derived suppressor cells (Gr-MDSCs) suppress natural killer (NK cell cytotoxicity in a cell contact-dependent manner. NK cell cytotoxicity against K562 tumour cell line with or without addition of cord-blood Gr-MDSCs in a 1:1 ratio was measured in a europium release assay. Effector : target (E : T) ratio was 2·5:1 (NK cells : K562 cells). CB: cord blood; TW: Transwell.