title: Human CD4+ CD39+ regulatory T cells produce adenosine upon co-expression of surface CD73 or contact with CD73+ exosomes or CD73+ cells.
abstract: While murine CD4(+) CD39(+) regulatory T cells (T(reg)) co-express CD73 and hydrolyze exogenous (e) adenosine triphosphate (ATP) to immunosuppressive adenosine (ADO), surface co-expression of CD73 on human circulating CD4(+) CD39(+) T(reg) is rare. Therefore, the ability of human T(reg) to produce and utilize ADO for suppression remains unclear. Using mass spectrometry, we measured nucleoside production by subsets of human CD4(+) CD39(+) and CD4(+) CD39(-)CD73(+) T cells or CD19(+) B cells isolated from blood of 30 volunteers and 14 cancer patients. CD39 and CD73 expression was evaluated by flow cytometry, Western blots, confocal microscopy or reverse transcription-polymerase chain reaction (RT-PCR). Circulating CD4(+) CD39(+) T(reg) which hydrolyzed eATP to 5'-AMP contained few intracytoplasmic granules and had low CD73 mRNA levels. Only ∼1% of these T(reg) were CD39(+) CD73(+) . In contrast, CD4(+) CD39(neg) CD73(+) T cells contained numerous CD73(+) granules in the cytoplasm and strongly expressed surface CD73. In vitro-generated T(reg) (Tr1) and most B cells were CD39(+) CD73(+) . All these CD73(+) T cell subsets and B cells hydrolyzed 5'-AMP to ADO. Exosomes isolated from plasma of normal control (NC) or cancer patients carried enzymatically active CD39 and CD73(+) and, when supplied with eATP, hydrolyzed it to ADO. Only CD4(+) CD39(+) T(reg) co-incubated with CD4(+) CD73(+) T cells, B cells or CD39(+) CD73(+) exosomes produced ADO. Thus, contact with membrane-tethered CD73 was sufficient for ADO production by CD4(+) CD39(+) T(reg). In microenvironments containing CD4(+) CD73(+) T cells, B cells or CD39(+) CD73(+) exosomes, CD73 is readily available to CD4(+) CD39(+) CD73(neg) T(reg) for the production of immunosuppressive ADO.
authors.affiliation: University of Pittsburgh Cancer Institute, Pittsburgh, PA, USA; Department of Otolaryngology, University of Ulm, Ulm, Germany.
authors.forename: P J
authors.initials: PJ
authors.lastname: Schuler
authors.forename: Z
authors.initials: Z
authors.lastname: Saze
authors.forename: C-S
authors.initials: CS
authors.lastname: Hong
authors.forename: L
authors.initials: L
authors.lastname: Muller
authors.forename: D G
authors.initials: DG
authors.lastname: Gillespie
authors.forename: D
authors.initials: D
authors.lastname: Cheng
authors.forename: M
authors.initials: M
authors.lastname: Harasymczuk
authors.forename: M
authors.initials: M
authors.lastname: Mandapathil
authors.forename: S
authors.initials: S
authors.lastname: Lang
authors.forename: E K
authors.initials: EK
authors.lastname: Jackson
authors.forename: T L
authors.initials: TL
authors.lastname: Whiteside
mesh_terms: D015720:5'-Nucleotidase / Q000235:genetics / Q000378:metabolism*; D000241:Adenosine / Q000096:biosynthesis*; D015703:Antigens, CD / Q000378:metabolism; D001081:Apyrase / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D055354:Exosomes / Q000378:metabolism*; D015870:Gene Expression; D006801:Humans; D016130:Immunophenotyping; D007963:Leukocytes, Mononuclear / Q000276:immunology / Q000378:metabolism; D008213:Lymphocyte Activation; D009369:Neoplasms / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D010641:Phenotype; D016176:T-Lymphocyte Subsets / Q000276:immunology / Q000378:metabolism; D050378:T-Lymphocytes, Regulatory / Q000276:immunology* / Q000378:metabolism*
chemical_list: D015703:Antigens, CD; D015720:5'-Nucleotidase; D001081:Apyrase; C067869:CD39 antigen; D000241:Adenosine
keywords: adenosine; ectonucleotidases; exosomes; human tTreg; immune suppression; pTreg
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