RT Journal Article
SR Electronic
T1 TNFa and IL-6 promote ex-vivo proliferation of lineage-committed human regulatory T cells
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2021.08.09.455690
DO 10.1101/2021.08.09.455690
A1 Nikolaos Skartsis
A1 Yani Peng
A1 Leonardo M.R. Ferreira
A1 Vinh Nguyen
A1 Yannick Muller
A1 Flavio Vincenti
A1 Qizhi Tang
YR 2021
UL http://biorxiv.org/content/early/2021/08/10/2021.08.09.455690.abstract
AB Treg therapy is being tested in clinical trials in transplantation and autoimmune diseases, however, the impact of inflammation on Tregs is unclear. In this study, we challenged human Tregs ex-vivo with pro-inflammatory cytokines, TNFα and IL-6. These cytokines enhanced Treg proliferation induced by anti-CD3 and anti-CD28 or CD28 superagonist (CD28SA) while maintaining high expression of FOXP3 and HELIOS, demethylated FOXP3 enhancer, and low expression of cytokines IFNg, IL-4 and IL-17. Blocking TNF receptor signaling using etanercept or deletion of TNF receptor 2 using CRISPR/Cas9 blunted Treg proliferation and attenuated FOXP3 and HELIOS expression, revealing the importance of TNFR2 signaling in Treg proliferation and lineage stability. The robust proliferation induced by CD28SA with IL-6 and TNFα may be adopted for the expansion of therapeutic Tregs. Metabolomics analysis showed that Tregs expanded with CD28SA plus cytokines had more active glycolysis and oxidative phosphorylation, increased energy production, and higher antioxidant potential. Finally, CD28SA plus cytokine-expanded Tregs had comparable suppressive activity in vitro and in vivo in a humanized mouse model of graft-versus-host-disease when compared to Tregs expanded using the conventional protocol. These results demonstrate that human Tregs positively respond to proinflammatory cytokines with enhanced proliferation without compromising their lineage identity or function.Competing Interest StatementQT is a co-founder and scientific advisor of Sonoma Biotherapeutics. QT, NS and FV are coinventors of a patent on manufacturing Tregs based on results from this work. The remaining authors have no relevant competing interests to disclose.aCD3/28anti-CD3 and anti-CD28 dynabeadsCD28SACD28 super agonistCNSCentral Nervous SystemFAOFatty acid oxidationFOXP3Forkhead box proteinP3 GVHDGraft-versus-host-diseaseGSHglutathioneGSSGglutathione disulfideLTαlymphotoxin αNSGNOD.Cg-PrkdcscidIl2rgtm1Wjl/SzJOXPHOSoxidative phosphorylationRORγTRetinoic acid-related orphan receptor γTTconvconventional CD4+ T cellTeffeffector T cellTNFR1Tumor necrosis factor receptor 1TNFR2Tumor necrosis factor receptor 2TNFSFTumor necrosis factor superfamilyTCRT cell receptorTSDRTreg-specific demethylated region