title: The primary immune response to Vaccinia virus vaccination includes cells with a distinct cytotoxic effector CD4 T-cell phenotype.
abstract: Smallpox was eradicated by a global program of inoculation with Vaccinia virus (VV). Robust VV-specific CD4 T-cell responses during primary infection are likely essential to controlling VV replication. Although there is increasing interest in cytolytic CD4 T-cells across many viral infections, the importance of these cells during acute VV infection is unclear.
  
  
  
  We undertook a detailed functional and genetic characterization of CD4 T-cells during acute VV-infection of humans. VV-specific T-cells were identified by up-regulation of activation markers directly ex vivo and through cytokine and co-stimulatory molecule expression. At day-13-post primary inoculation with VV, CD38highCD45RO+ CD4 T-cells were purified by cell sorting, RNA isolated and analysed by microarray. Differential expression of up-regulated genes in activated CD4 T-cells was confirmed at the mRNA and protein levels. We compared analyses of VV-specific CD4 T-cells to studies on 12 subjects with primary HIV infection (PHI). VV-specific T-cells lines were established from PBMCs collected post vaccination and checked for cytotoxicity potential.
  
  
  
  A median 11.9% CD4 T-cells were CD38highCD45RO+ at day-13 post-VV inoculation, compared to 3.0% prior and 10.4% during PHI. Activated CD4 T-cells had an up-regulation of genes related to cytolytic function, including granzymes K and A, perforin, granulysin, TIA-1, and Rab27a. No difference was seen between CD4 T-cell expression of perforin or TIA-1 to VV and PHI, however granzyme k was more dominant in the VV response. At 25:1 effector to target ratio, two VV-specific T-cell lines exhibited 62% and 30% cytotoxicity respectively and CD107a degranulation.
  
  
  
  We show for the first time that CD4 CTL are prominent in the early response to VV. Understanding the role of CD4 CTL in the generation of an effective anti-viral memory may help develop more effective vaccines for diseases such as HIV.
authors.affiliation: The Kirby Institute for infection and immunity in society, UNSW Australia, Sydney, NSW, Australia. Electronic address: cmunier@kirby.unsw.edu.au.
authors.forename: C Mee Ling
authors.initials: CML
authors.lastname: Munier
authors.affiliation: The Kirby Institute for infection and immunity in society, UNSW Australia, Sydney, NSW, Australia.
authors.forename: David
authors.initials: D
authors.lastname: van Bockel
authors.affiliation: The Kirby Institute for infection and immunity in society, UNSW Australia, Sydney, NSW, Australia.
authors.forename: Michelle
authors.initials: M
authors.lastname: Bailey
authors.affiliation: The Kirby Institute for infection and immunity in society, UNSW Australia, Sydney, NSW, Australia.
authors.forename: Susanna
authors.initials: S
authors.lastname: Ip
authors.affiliation: The Kirby Institute for infection and immunity in society, UNSW Australia, Sydney, NSW, Australia.
authors.forename: Yin
authors.initials: Y
authors.lastname: Xu
authors.affiliation: Department of Microbiology and Immunology, Peter Doherty Institute, University of Melbourne, Melbourne, VIC, Australia.
authors.forename: Sheilajen
authors.initials: S
authors.lastname: Alcantara
authors.affiliation: The Garvan Institute, Sydney, NSW, Australia.
authors.forename: Sue Min
authors.initials: SM
authors.lastname: Liu
authors.affiliation: School of Molecular Bioscience, Faculty of Science, The University of Sydney, NSW, Australia.
authors.forename: Gareth
authors.initials: G
authors.lastname: Denyer
authors.affiliation: The Garvan Institute, Sydney, NSW, Australia.
authors.forename: Warren
authors.initials: W
authors.lastname: Kaplan
authors.affiliation: The Kirby Institute for infection and immunity in society, UNSW Australia, Sydney, NSW, Australia; St Vincent's Hospital, Sydney, NSW, Australia.
authors.forename: Kazuo
authors.initials: K
authors.lastname: Suzuki
authors.affiliation: Infection and Immunity Program, Department of Biochemistry and Molecular Biology, Biomedicine Discovery Institute, Monash University, Clayton, VIC, Australia.
authors.forename: Nathan
authors.initials: N
authors.lastname: Croft
authors.affiliation: Infection and Immunity Program, Department of Biochemistry and Molecular Biology, Biomedicine Discovery Institute, Monash University, Clayton, VIC, Australia.
authors.forename: Anthony
authors.initials: A
authors.lastname: Purcell
authors.affiliation: John Curtin School of Medical Research, The Australian National University, Canberra, ACT, Australia.
authors.forename: David
authors.initials: D
authors.lastname: Tscharke
authors.affiliation: The Kirby Institute for infection and immunity in society, UNSW Australia, Sydney, NSW, Australia; St Vincent's Hospital, Sydney, NSW, Australia.
authors.forename: David A
authors.initials: DA
authors.lastname: Cooper
authors.affiliation: Department of Microbiology and Immunology, Peter Doherty Institute, University of Melbourne, Melbourne, VIC, Australia; Melbourne Sexual Health Centre and Department of Infectious Diseases, Alfred Health, Central Clinical School, Monash University Melbourne, VIC, Australia; ARC Centre of Excellence in Convergent Bio-Nano Science and Technology, University of Melbourne, Parkville, VIC, Australia.
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Kent
authors.affiliation: The Kirby Institute for infection and immunity in society, UNSW Australia, Sydney, NSW, Australia; St Vincent's Hospital, Sydney, NSW, Australia.
authors.forename: John J
authors.initials: JJ
authors.lastname: Zaunders
authors.affiliation: The Kirby Institute for infection and immunity in society, UNSW Australia, Sydney, NSW, Australia; St Vincent's Hospital, Sydney, NSW, Australia.
authors.forename: Anthony D
authors.initials: AD
authors.lastname: Kelleher
mesh_terms: D051997:ADP-ribosyl Cyclase 1 / Q000235:genetics; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology; D016207:Cytokines / Q000235:genetics; D003602:Cytotoxicity, Immunologic; D053804:Granzymes / Q000235:genetics; D015658:HIV Infections / Q000276:immunology; D006801:Humans; D017493:Leukocyte Common Antigens / Q000235:genetics; D008213:Lymphocyte Activation; D054353:Perforin / Q000235:genetics; D010641:Phenotype; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*; D046888:Tissue Array Analysis; D015854:Up-Regulation; D014616:Vaccinia virus / Q000276:immunology*; D014765:Viral Vaccines / Q000008:administration & dosage / Q000276:immunology*
chemical_list: D016207:Cytokines; D014765:Viral Vaccines; D054353:Perforin; D017493:Leukocyte Common Antigens; C516769:PTPRC protein, human; D051997:ADP-ribosyl Cyclase 1; D053804:Granzymes
keywords: CD4; CD8; Cytotoxic T-cells; Granzyme K; HIV; Microarray; Vaccinia