Unknown,Transcriptomics,Genomics,Proteomics

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Molecular and cellular characterization of Cord Blood derived, IDO expressing human fibrocystic Myeloid Derived Suppressor Cells

ABSTRACT: By restraining T cell activation and promoting regulatory T cell (Treg) expansion, myeloid-derived suppressor cells (MDSC) and tolerogenic dendritic cells (DC) (tDC) can control self-reactive and anti-graft effector T cells in autoimmunity and transplantation. Their therapeutic use and characterization, however, is limited by their scarce availability in the peripheral blood of tumor-free donors. In the present study we describe and characterize a novel population of myeloid suppressor cells, named fibrocytic MDSC (f-MDSC), that are differentiated from umbilical cord blood (UCB) precursors by a 4 day culture with FDA approved cytokines (rh-GM-CSF and rh-G-CSF). This MDSC subset, characterized by the expression of MDSC-, DC-, and fybrocyte-s associated markers, promotes Treg regulatory cells expansion and promote normoglycemia in a xenogeneic model of Type 1 Diabetes (T1D). In order to exert their pro-tolerogenic function, fibrocytic MDSC require direct contact with activated T cells, which leads to the induction and secretion of indoleamine 2,3 deoxygenase (IDO). This new myeloid subset may represent an important tool for the in vitro and in vivo production of Treg for the treatment of autoimmune diseases, and either prevention or control of allograft rejection. Keywords: expression profiling by array Human Umbilical cord blood (UCB) samples were cultured for 4 days with GM-CSF and G-CSF after Ficoll-Paque gradient separation and red cell lysis. CD33+/IL-4RM-NM-1+ cells were sterilely sorted and part stored in Trizol for RNA extraction (BEFORE CONTACT), and part co-cultured with autologous PHA activated CD3+ T cells either in cell to cell contact (CONTACT) or using a transwell device to separate the two popyulations (TRANSWELL). Three days later, T cells were removed from the co-culture either by repeated washes with PBS or by simply removing the transwell device. Adherent cells were lysed with Trizol and stored at -80M-bM-^@M-^YC. RNA extraction was carry on with miRNAeasy mini kit from Quiagen; RNA QC was assessed by Agilent BioAnalyzer 6000 and samples were analyzed with Affymetrix Human 2.0 ST array

ORGANISM(S): Homo sapiens

SUBMITTER: Silvio Bicciato

PROVIDER: E-GEOD-52376 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Human fibrocytic myeloid-derived suppressor cells express IDO and promote tolerance via Treg-cell expansion.

Zoso Alessia A Mazza Emilia M C EM Bicciato Silvio S Mandruzzato Susanna S Bronte Vincenzo V Serafini Paolo P Inverardi Luca L

European journal of immunology 20141018 11

By restraining T-cell activation and promoting Treg-cell expansion, myeloid-derived suppressor cells (MDSCs) and tolerogenic DCs can control self-reactive and antigraft effector T cells in autoimmunity and transplantation. Their therapeutic use and characterization, however, is limited by their scarce availability in the peripheral blood of tumor-free donors. In the present study, we describe and characterize a novel population of human myeloid suppressor cells, named fibrocytic MDSC, which are ...[more]

PMID: 25113564

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Project description:We have demonstrated therapeutic efficacy of MDSC in mouse Alopecia Areata (AA). In the same AA model, we now asked whether MDSC exosomes (MDSC-Exo) can replace MDSC. MDSC-Exo from bone marrow cells (BMC) cultures of healthy donors could substantially facilitate treatment. With knowledge on MDSC-Exo being limited, their suitability needs to be verified in advance. Protein marker profiles suggest comparability of BMC- to ex vivo collected inflammatory MDSC/MDSC-Exo in mice with a chronic contact dermatitis, which is a therapeutic option in AA. Proteome analyses substantiated a large overlap of function-relevant molecules in MDSC and MDSC-Exo. Furthermore, MDSC-Exo are taken up by T cells, macrophages, NK, and most avidly by Treg and MDSC-Exo uptake exceeds binding of MDSC themselves. In AA mice, MDSC-Exo preferentially target skin-draining lymph nodes and cells in the vicinity of remnant hair follicles. MDSC-Exo uptake is accompanied by a strong increase in Treg, reduced T helper proliferation, mitigated cytotoxic activity, and a slight increase in lymphocyte apoptosis. Repeated MDSC-Exo application in florid AA prevented progression and sufficed for partial hair regrowth. Deep sequencing of lymphocyte mRNA from these mice revealed a significant increase in immunoregulatory mRNA, including FoxP3 and arginase 1. Downregulated mRNA was preferentially engaged in prohibiting T cell hyperreactivity. Taken together, proteome analysis provided important insights into potential MDSC-Exo activities, these Exo preferentially homing into AA-affected organs. Most importantly, changes in leukocyte mRNA seen after treatment of AA mice with MDSC-Exo sustainably supports the strong impact on the adaptive and the non-adaptive immune system, with Treg expansion being a dominant feature. Thus, MDSC-Exo could potentially serve as therapeutic agents in treating AA and other autoimmune diseases. These shotgun data represent GeLC-MS/MS analyses of murine MDSC and CDMS+ cells and of the exosomes from both cell types. Three pulldown RPLC-MS/MS experiments are also included. Cysteines were carbamidomethylated via iodoacetamide. Tandem mass spectrometry took place in an LTQ Orbitrap XL, with scans acquired in the linear ion trap.

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Dataset Information

Molecular and cellular characterization of Cord Blood derived, IDO expressing human fibrocystic Myeloid Derived Suppressor Cells

Publications

Human fibrocytic myeloid-derived suppressor cells express IDO and promote tolerance via Treg-cell expansion.

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