Impact of 5-aza-2'-deoxycytidine and epigallocatechin-3-gallate for induction of human regulatory T cells.
The epigenetic regulation of transcription factor genes is critical for T-cell lineage specification. A specific methylation pattern within a conserved region of the lineage specifying transcription factor gene FOXP3, the Treg-specific demethylated region (TSDR), is restricted to regulatory T (Treg) cells and is required for stable expression of FOXP3 and suppressive function. We analysed the impact of hypomethylating agents 5-aza-2'-deoxycytidine and epigallocatechin-3-gallate on human CD4(+)  CD25(-) T cells for generating demethylation within FOXP3-TSDR and inducing functional Treg cells. Gene expression, including lineage-specifying transcription factors of the major T-cell lineages and their leading cytokines, functional properties and global transcriptome changes were analysed. The FOXP3-TSDR methylation pattern was determined by using deep amplicon bisulphite sequencing. 5-aza-2'-deoxycytidine induced FOXP3-TSDR hypomethylation and expression of the Treg-cell-specific genes FOXP3 and LRRC32. Proliferation of 5-aza-2'-deoxycytidine-treated cells was reduced, but the cells did not show suppressive function. Hypomethylation was not restricted to FOXP3-TSDR and expression of master transcription factors and leading cytokines of T helper type 1 and type 17 cells were induced. Epigallocatechin-3-gallate induced global DNA hypomethylation to a lesser extent than 5-aza-2'-deoxycitidine, but no relevant hypomethylation within FOXP3-TSDR or expression of Treg-cell-specific genes. Neither of the DNA methyltransferase inhibitors induced fully functional human Treg cells. 5-aza-2'-deoxycitidine-treated cells resembled Treg cells, but they did not suppress proliferation of responder cells, which is an essential capability to be used for Treg cell transfer therapy. Using a recently developed targeted demethylation technology might be a more promising approach for the generation of functional Treg cells.
Institute of Medical Microbiology, University Hospital Essen, University of Duisburg-Essen, Essen, Germany.
Jan
J
Kehrmann
Roman
R
Tatura
Michael
M
Zeschnigk
Michael
M
Probst-Kepper
Robert
R
Geffers
Joerg
J
Steinmann
Jan
J
Buer
D001374:Azacitidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D002392:Catechin / Q000031:analogs & derivatives* / Q000494:pharmacology; D000077209:Decitabine; D051858:Forkhead Transcription Factors / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008297:Male; D008745:Methylation / Q000187:drug effects; D020411:Oligonucleotide Array Sequence Analysis; D050378:T-Lymphocytes, Regulatory / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism
C418974:FOXP3 protein, human; D051858:Forkhead Transcription Factors; D000077209:Decitabine; D002392:Catechin; C045651:epigallocatechin gallate; D001374:Azacitidine
5-Aza-dC; decitabine; epigallocatechin-3-gallate; methylation; regulatory T cell