title: IFNy+ and IFNy- Treg subsets with stable and unstable Foxp3 expression in kidney transplant recipients with good long-term graft function.
abstract: BACKGROUND
  Treg are a heterogenous cell population. In the present study we attempted to identify Treg subsets that might contribute to stable and good long-term graft function.
  
  
  METHODS
  Lymphocyte and Treg subsets were studied in 136 kidney transplant recipients with good long-term graft function and in 52 healthy control individuals using eight-color-fluorescence flow cytometry. Foxp3 TSDR methylation status was investigated in enriched IFNy+ and IFNy- Treg preparations using high resolution melt analysis.
  
  
  RESULTS
  Compared with healthy controls, patients showed strong associations of IFNy secreting Helios+ and Helios- Treg with Treg that co-expressed perforin and/or CTLA4 (CD152; p<0.01). Moreover they showed associations of IFNy-Helios+ Treg with Treg that produced TGFβ and/or perforin and of IFNy-Helios- Treg with TGFβ production (all p<0.01). Only in patients, but not in healthy controls, were IFNy- Helios+ and Helios- Treg associated with higher CD45+, CD3+, (CD4+), CD19+ lymphocyte counts (p<0.001). In addition IFNy-Helios+ Treg were associated with CD16+56+ lymphocytes (p<0.001). Enriched IFNy- Treg from female but not male patients showed an association of Foxp3 methylation with higher total Treg and higher Helios+IFNy-, CXCR3+Lselectin+ (CD183+CD62L+), CXCR3-Lselectin+ and CD28+HLADR+ Treg subsets (p<0.01). Enriched IFNy+ Treg from male patients showed an association of demethylated Foxp3 with total Treg and IL10-TFGβ+ Treg counts, and in enriched IFNy- Treg an association of methylated Foxp3 with APO1/FasR+FasL+ (CD95+CD178+) Treg (p<0.01).
  
  
  CONCLUSIONS
  Kidney recipients with good long-term graft function possess IFNy+ and IFNy- Treg with stable and unstable Foxp3 expression in the blood. They co-express CD28, HLADR, CTLA4, CXCR3, Lselectin, TGFβ, perforin and FasL and might contribute to the establishment and maintenance of good long-term graft function.
authors.affiliation: Transplantation-Immunology, Institute of Immunology, University Hospital Heidelberg, Im Neuenheimer Feld 305, 69120 Heidelberg, Germany. Electronic address: Karina.Trojan@med.uni-heidelberg.de.
authors.forename: Karina
authors.initials: K
authors.lastname: Trojan
authors.affiliation: Transplantation-Immunology, Institute of Immunology, University Hospital Heidelberg, Im Neuenheimer Feld 305, 69120 Heidelberg, Germany. Electronic address: Christian.Unterrainer@med.uni-heidelberg.de.
authors.forename: Christian
authors.initials: C
authors.lastname: Unterrainer
authors.affiliation: Transplantation-Immunology, Institute of Immunology, University Hospital Heidelberg, Im Neuenheimer Feld 305, 69120 Heidelberg, Germany. Electronic address: Mostafa.Aly@med.uni-heidelberg.de.
authors.forename: Mostafa
authors.initials: M
authors.lastname: Aly
authors.affiliation: Transplantation-Immunology, Institute of Immunology, University Hospital Heidelberg, Im Neuenheimer Feld 305, 69120 Heidelberg, Germany. Electronic address: Li.Zhu@med.uni-heidelberg.de.
authors.forename: Li
authors.initials: L
authors.lastname: Zhu
authors.affiliation: Department of Internal Medicine, University of Giessen, Klinikstrasse 33, D-35385 Giessen, Germany. Electronic address: Rolf.Weimer@innere.med.uni-giessen.de.
authors.forename: Rolf
authors.initials: R
authors.lastname: Weimer
authors.affiliation: Department of Internal Medicine, University of Giessen, Klinikstrasse 33, D-35385 Giessen, Germany. Electronic address: Nuray.Bulut@innere.med.uni-giessen.de.
authors.forename: Nuray
authors.initials: N
authors.lastname: Bulut
authors.affiliation: Department of Nephrology, University of Heidelberg, Heidelberg, Germany. Electronic address: Christian.Morath@med.uni-heidelberg.de.
authors.forename: Christian
authors.initials: C
authors.lastname: Morath
authors.affiliation: Transplantation-Immunology, Institute of Immunology, University Hospital Heidelberg, Im Neuenheimer Feld 305, 69120 Heidelberg, Germany. Electronic address: Gerhard.Opelz@med.uni-heidelberg.de.
authors.forename: Gerhard
authors.initials: G
authors.lastname: Opelz
authors.affiliation: Transplantation-Immunology, Institute of Immunology, University Hospital Heidelberg, Im Neuenheimer Feld 305, 69120 Heidelberg, Germany. Electronic address: Volker.Daniel@med.uni-heidelberg.de.
authors.forename: Volker
authors.initials: V
authors.lastname: Daniel
keywords: Foxp3 TSDR methylation; Good long-term graft function; IFNy+ Treg; IFNy− Treg; Kidney transplant recipients; Stable Foxp3 expression; Unstable Foxp3 expression