title: Helios expression and Foxp3 TSDR methylation of IFNy+ and IFNy- Treg from kidney transplant recipients with good long-term graft function.
abstract: BACKGROUND
  There is circumstantial evidence that IFNy+ Treg might have clinical relevance in transplantation. IFNy+ Treg express IFNy receptors and are induced by IFNy. In the present study we investigated in kidney transplant recipients with good long-term stable graft function the absolute cell counts of IFNy+ Treg subsets and whether their expression of Foxp3 is stable or transient.
  
  
  METHODS
  Helios expression determined by eight-color-fluorescence flow cytometry and methylation status of the Foxp3 Treg specific demethylation region (TSDR) served as indicators for stability of Foxp3 expression. Methylation status was investigated in enriched IFNy+ and IFNy- Treg preparations originating from peripheral blood using high resolution melt analysis. A total of 136 transplant recipients and 52 healthy controls were studied.
  
  
  RESULTS
  Proportions of IFNy+ Treg were similar in patients and healthy controls (0.05% and 0.04% of all CD4+ lymphocytes; p = n.s.). Patients also had similar absolute counts of IFNy producing Helios+ and Helios- Treg (p = n.s.). Most of the IFNy+ and IFNy- Treg in transplant recipients had a methylated Foxp3 TSDR, however, there was a sizeable proportion of IFNy+ and IFNy- Treg with demethylated Foxp3 TSDR. Male and female patients showed more frequently methylated IFNy+ and IFNy- Treg than male and female controls (all p<0.05).
  
  
  CONCLUSIONS
  Kidney transplant recipients with good long-term stable graft function have similar levels of IFNy+ Treg as healthy controls. IFNy+ and IFNy- Treg subsets in patients consist of cells with stable and cells with transient Foxp3 expression; however, patients showed more frequently methylated IFNy+ and IFNy- Treg than controls. The data show increased levels of Treg subsets with stable as well as transient Foxp3 expression in patients with stable allograft acceptance compared to healthy controls.
authors.affiliation: Transplantation-Immunology, Institute of Immunology, University Hospital Heidelberg, Heidelberg, Germany.
authors.forename: Karina
authors.initials: K
authors.lastname: Trojan
authors.affiliation: Transplantation-Immunology, Institute of Immunology, University Hospital Heidelberg, Heidelberg, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Unterrainer
authors.affiliation: Department of Internal Medicine, University of Giessen, Giessen, Germany.
authors.forename: Rolf
authors.initials: R
authors.lastname: Weimer
authors.affiliation: Department of Internal Medicine, University of Giessen, Giessen, Germany.
authors.forename: Nuray
authors.initials: N
authors.lastname: Bulut
authors.affiliation: Department of Nephrology, University of Heidelberg, Heidelberg, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Morath
authors.affiliation: Transplantation-Immunology, Institute of Immunology, University Hospital Heidelberg, Heidelberg, Germany.|Nephrology unit, Internal Medicine Department, Assiut University, Assiut, Egypt.
authors.forename: Mostafa
authors.initials: M
authors.lastname: Aly
authors.affiliation: Transplantation-Immunology, Institute of Immunology, University Hospital Heidelberg, Heidelberg, Germany.|Department of Hematology, Tongji Hospital, Huazhong University of Science and Technology, Wuhan, China.
authors.forename: Li
authors.initials: L
authors.lastname: Zhu
authors.affiliation: Transplantation-Immunology, Institute of Immunology, University Hospital Heidelberg, Heidelberg, Germany.
authors.forename: Gerhard
authors.initials: G
authors.lastname: Opelz
authors.affiliation: Transplantation-Immunology, Institute of Immunology, University Hospital Heidelberg, Heidelberg, Germany.
authors.forename: Volker
authors.identifier: http://orcid.org/0000-0001-8345-1031
authors.initials: V
authors.lastname: Daniel
mesh_terms: D000328:Adult; D000368:Aged; D016022:Case-Control Studies; D019175:DNA Methylation; D005260:Female; D051858:Forkhead Transcription Factors / Q000378:metabolism*; D006801:Humans; D051740:Ikaros Transcription Factor / Q000378:metabolism*; D007371:Interferon-gamma / Q000276:immunology*; D007372:Interferons; D007668:Kidney / Q000503:physiopathology*; D016030:Kidney Transplantation; D016131:Lymphocyte Subsets; D008297:Male; D008875:Middle Aged; D050378:T-Lymphocytes, Regulatory / Q000276:immunology*
chemical_list: C418974:FOXP3 protein, human; D051858:Forkhead Transcription Factors; C111431:IKZF2 protein, human; D051740:Ikaros Transcription Factor; D007371:Interferon-gamma; D007372:Interferons