title: Increased peripheral CD4+regulatory T cells persist after successful direct-acting antiviral treatment of chronic hepatitis C.
abstract: CD4+regulatory T cells (Tregs) expand during chronic hepatitis C virus (HCV) infection, inhibit antiviral immunity and promote fibrosis. Direct-acting antiviral agents (DAA) have revolutionized HCV therapy. However, it is unclear if Tregs are normalized after DAA-induced HCV elimination.
  
  
  
  We analyzed Tregs before (baseline), at end of therapy (EOT), 12 and 24weeks (SVR12, SVR24) and long-term (51±14weeks) after EOT in 26 genotype-1-infected patients who were successfully treated with sofosbuvir (SOF) plus interferon (IFN)/ribavirin (n=12) and IFN-free DAA regimens (SOF plus daclatasvir or simeprevir; n=14). Frequency, phenotype and suppressor function of peripheral Foxp3+CD25+CD4+T cells were studied by multi-color flow cytometry and co-culture inhibition assays.
  
  
  
  Frequencies and activation status of Foxp3+CD25+CD4+T cells remained elevated above those of normal controls in both treatment groups even long-term after HCV elimination. Co-culture assays indicated a dose-response relationship for functional inhibition of autologous CD4+effector T cells and confirmed that activation of Tregs remained largely unchanged over the observation period. Unlike IFN-free regimens, SOF plus IFN/ribavirin induced a transiently increased frequency of Foxp3+CD25+CD4+T cells at EOT (5.0% at baseline to 6.1% at EOT; p=0.001). These Foxp3+CD25+CD4+T cells co-expressed the activation markers glycoprotein A repetitions predominant (GARP; p=0.012) and tumor necrosis factor receptor superfamily, member 4 (OX-40; p=0.001) but showed unchanged in vitro inhibitory activity.
  
  
  
  Although IFN-based DAA therapy induced transient expansion of activated Foxp3+CD25+CD4+T cells, neither IFN-based nor IFN-free DAA regimens normalized frequencies and activation status of Tregs one year after viral elimination. Persistence of immunosuppressive Tregs may thus contribute to complications of liver disease even long-term after HCV cure.
  
  
  
  In chronic hepatitis C virus (HCV) infection, CD4+regulatory T cells (Tregs) can reduce antiviral immune responses, promote liver fibrosis and may increase the risk for liver cancer, because they gradually expand during disease. Modern direct-acting antiviral agents (DAA) can "cure" hepatitis C in almost all treated patients. However, our study shows that DAA do not normalize the increased frequency and activation status of Tregs even long-term after HCV elimination. Tregs may persistently modulate functions of the immune system even after "cure" of hepatitis C.
authors.affiliation: Department of Internal Medicine I, University of Bonn, Bonn, Germany, and the German Center for Infection Research (DZIF), Partner Site Cologne-Bonn, Bonn, Germany. Electronic address: bettina.langhans@ukb.uni-bonn.de.
authors.forename: Bettina
authors.initials: B
authors.lastname: Langhans
authors.affiliation: Department of Internal Medicine I, University of Bonn, Bonn, Germany, and the German Center for Infection Research (DZIF), Partner Site Cologne-Bonn, Bonn, Germany.
authors.forename: Hans Dieter
authors.initials: HD
authors.lastname: Nischalke
authors.affiliation: Department of Internal Medicine I, University of Bonn, Bonn, Germany, and the German Center for Infection Research (DZIF), Partner Site Cologne-Bonn, Bonn, Germany.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Krämer
authors.affiliation: Department of Internal Medicine I, University of Bonn, Bonn, Germany, and the German Center for Infection Research (DZIF), Partner Site Cologne-Bonn, Bonn, Germany.
authors.forename: Annekristin
authors.initials: A
authors.lastname: Hausen
authors.affiliation: Department of Internal Medicine I, University of Bonn, Bonn, Germany, and the German Center for Infection Research (DZIF), Partner Site Cologne-Bonn, Bonn, Germany.
authors.forename: Leona
authors.initials: L
authors.lastname: Dold
authors.affiliation: Department of Internal Medicine I, University of Bonn, Bonn, Germany, and the German Center for Infection Research (DZIF), Partner Site Cologne-Bonn, Bonn, Germany.
authors.forename: Peer
authors.initials: P
authors.lastname: van Heteren
authors.affiliation: Department of Internal Medicine I, University of Bonn, Bonn, Germany, and the German Center for Infection Research (DZIF), Partner Site Cologne-Bonn, Bonn, Germany.
authors.forename: Robert
authors.initials: R
authors.lastname: Hüneburg
authors.affiliation: Department of Internal Medicine I, University of Bonn, Bonn, Germany, and the German Center for Infection Research (DZIF), Partner Site Cologne-Bonn, Bonn, Germany.
authors.forename: Jacob
authors.initials: J
authors.lastname: Nattermann
authors.affiliation: Department of Internal Medicine I, University of Bonn, Bonn, Germany, and the German Center for Infection Research (DZIF), Partner Site Cologne-Bonn, Bonn, Germany.
authors.forename: Christian P
authors.initials: CP
authors.lastname: Strassburg
authors.affiliation: Department of Internal Medicine I, University of Bonn, Bonn, Germany, and the German Center for Infection Research (DZIF), Partner Site Cologne-Bonn, Bonn, Germany.
authors.forename: Ulrich
authors.initials: U
authors.lastname: Spengler
mesh_terms: D000328:Adult; D000368:Aged; D000998:Antiviral Agents / Q000627:therapeutic use*; D005260:Female; D051858:Forkhead Transcription Factors / Q000032:analysis; D037161:Galectins / Q000097:blood; D019698:Hepatitis C, Chronic / Q000188:drug therapy* / Q000276:immunology; D006801:Humans; D008297:Male; D008875:Middle Aged; D050378:T-Lymphocytes, Regulatory / Q000276:immunology*
chemical_list: D000998:Antiviral Agents; C418974:FOXP3 protein, human; D051858:Forkhead Transcription Factors; D037161:Galectins; C105032:LGALS9 protein, human
keywords: CD4(+) regulatory T cells; Chronic hepatitis C virus infection; Directly acting antivirals; Tim-3/galectin-9 pathway