Decidua-derived granulocyte macrophage colony-stimulating factor induces polymorphonuclear myeloid-derived suppressor cells from circulating CD15+ neutrophils

Abstract

STUDY QUESTION Do decidua-derived factors stimulate the conversion of circulating neutrophils to polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) in early human pregnancy? SUMMARY ANSWER Circulating neutrophils can acquire PMN-MDSC-like phenotypes and function via phosphorylated signal transducer and activator of transcription 5/programmed death ligand 2 (pSTAT5/PD-L2) signalling after stimulation with decidua-derived granulocyte macrophage colony-stimulating factor (GM-CSF). WHAT IS KNOWN ALREADY PMN-MDSCs are an important immunoregulatory cell type in early pregnancy. Neutrophils are of high heterogeneity and plasticity and can polarize to immunosuppressive PMN-MDSCs upon stimulation. STUDY DESIGN, SIZE, DURATION For analysis of myeloid-derived suppressor cell (MDSC) subset proportions, 12 endometrium tissues and 12 peripheral blood samples were collected from non-pregnant women, and 40 decidua tissues and 16 peripheral blood samples were obtained from women with normal early pregnancy undergoing elective surgical pregnancy termination for nonmedical reasons with gestation age of 6–10 weeks. Twenty-nine decidua tissues were collected for isolation of CD15+ PMN-MDSCs. Twenty endometrium tissues and 30 decidua tissues were collected for cytokine analysis, immunohistochemistry or neutrophil stimulation. Peripheral blood samples were obtained from 36 healthy donors for isolation of CD3+ T cells and CD15+ neutrophils. PARTICIPANTS/MATERIALS, SETTING, METHODS The proportion of MDSC subsets in the decidua and peripheral blood of normal early pregnancy, endometrium and peripheral blood of non-pregnant women was analysed by flow cytometry. The phenotypes and function of decidual PMN-MDSCs and circulating neutrophils were compared by flow cytometry. Circulating neutrophils were stimulated with decidual explant supernatant (DES) and the phenotypes were measured by flow cytometry and immunofluorescence. The suppressive capacity of decidual PMN-MDSCs and DES-conditioned neutrophils was analysed by flow cytometry with or without anti-programmed cell death-1 (PD-1) antibody. Cytokines from DES and endometrial explant supernatant (EES) were detected by a Luminex assay. GM-CSF expression was determined by ELISA and immunohistochemistry. Neutrophils were stimulated with DES, EES, DES with anti-GM-CSF antibody or EES with GM-CSF. CD11b, lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1), PD-L2 and pSTAT5 expression were measured by flow cytometry. MAIN RESULTS AND THE ROLE OF CHANCE The frequency of PMN-MDSCs was significantly increased in the decidua of early pregnancy compared with peripheral blood of non-pregnant women, the endometrium of non-pregnant women or peripheral blood during early pregnancy. Decidual PMN-MDSCs suppressed T-cell proliferation and cytokine production. Phenotypes of decidual PMN-MDSCs were similar to mature activated neutrophils. DES-induced CD11b, LOX-1, PD-L2 expression and STAT5 phosphorylation in neutrophils. The PD-L2 expression in neutrophils was dependent on STAT5 phosphorylation. Both decidual PMN-MDSCs and DES-conditioned neutrophils suppressed T-cell proliferation via PD-1 signalling. GM-CSF was up-regulated in the decidua and induced CD11b, LOX-1 and PD-L2 expression on neutrophils. DES significantly induced CD11b, LOX-1, PD-L2 expression and STAT5 phosphorylation. Anti-GM-CSF antibody remarkably blocked such stimulation in neutrophils. EES did not induce CD11b, LOX-1, PD-L2 expression or STAT5 phosphorylation, while GM-CSF treatment sufficiently stimulated CD11b, LOX-1, PD-L2 expression and STAT5 phosphorylation in neutrophils. LARGE SCALE DATA N/A LIMITATIONS, REASONS FOR CAUTION The study was based on in vitro experiments and we were not able to evaluate neutrophils differentiation to PMN-MDSCs in other sites before entering the maternal–foetal interface due to the limited availability of human samples. This needs to be explored using murine models. WIDER IMPLICATIONS OF THE FINDINGS This is the first study demonstrating that decidual PMN-MDSCs are a group of immunoregulatory cells with mature status, and that neutrophils can be induced to a PMN-MDSC-like phenotype with decidua-derived GM-CSF via pSTAT5/PD-L2 signalling. This study indicates that GM-CSF can facilitate immune tolerance of early pregnancy through regulating PMN-MDSCs and further provides a potential role of GM-CSF in prevention and treatment for pregnancy complications. STUDY FUNDING/COMPETING INTEREST(S) This work was supported by the National Natural Science Foundation of China (81671481) and National Natural Science Foundation of China (81871179). All authors have no competing interests to declare.

Neutrophils in pregnancy: New insights into innate and adaptive immune regulation

“…In early pregnancy, it is believed that circulating neutrophils can acquire a MDSC phenotype upon migration to the maternal–fetal interface following stimulation with decidua‐derived GM‐CSF. Stimulation of CD15 + neutrophils with GM‐CSF activates STAT5 signalling, leading to the induction PD‐L2 expression which consequently is used by the neutrophil to restrain T‐cell proliferation though PD‐1 signalling [ 97 ]. CXCR2 has also been implicated with the recruitment of G‐MDSCs in decidual tissue and induces arginase I, an effector molecule used by MDSCs to regulate T‐cell activity [ 98 ].…”

Myeloid-Derived Suppressor Cells Alleviate Renal Fibrosis Progression via Regulation of CCL5-CCR5 Axis

“…Recent studies have indicated that G-CSF pretreatment attenuates acute ischemia-reperfusion injury and subsequent renal fibrosis by increasing MDSC infiltration into the kidney (12). GM-CSF also stimulates neutrophils to MDSCs conversion in early human pregnancy (14). Since there is also an evidence that bone marrow (BM)-induced MDSCs exhibit more potent suppressive function than mice-isolated MDSCs (13), here raises new questions: is there a difference in the function of MDSCs induced in vitro or in vivo?…”

Regulation of the innate immune cells during pregnancy: An immune checkpoint perspective

“… 72 Compared to peripheral blood, elevated PD‐L1 expression by CD4 + T, Treg, NKT‐like and CD56 + NK cells were also found with the increased PD‐1 expression by decidual CD8 + T, CD4 + T and NKT‐like cells. 77 PD‐L2 is expressed on the decidual polymorphonuclear myeloid‐derived suppressor cells (PMN‐MDSCs) and neutrophils 76 , 80 and is also expressed in syncytiotrophoblasts. 81 T‐cell proliferation is suppressed by both decidual PMN‐MDSCs and decidual explant supernatant (DES)‐conditioned neutrophils via PD‐1 signalling.…”

“…A shift from circulating neutrophils to PMN‐MDSC‐like phenotypes requires stimulation with decidua‐derived granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) by pSTAT5/PD‐L2 signaling. 80 Overall, the PD‐1‐PD‐L1 axis not only regulates T cells including promoting a shift to Type 2 helper T‐cell (Th2) bias 82 and Treg cell proliferation, 83 , 84 as well as reducing CD8 + T accompanied by NKG2D activating receptor, 77 , 85 , 86 , 87 , 88 , 89 , 90 Th17 83 , 91 and TFH 92 , 93 responses but also plays an important role in innate immune cells for pregnancy maintenance. 94 …”