title: Use of retroviral vectors for the analysis of SIV/HIV-specific CD8 T cell responses.
abstract: CD8+ T cell responses and particularly cytotoxic T lymphocyte (CTL) activity are critical factors in controlling SHIV, SIV or HIV replication during natural infection and represent key parameters which need to be monitored during vaccine development. In order to improve the methodology for measuring CD8+ T cell responses, retroviral vectors expressing the full-length SIV-Gag or HIV-Env proteins were constructed and used to transduce B lymphoblastoid cell lines (BLCL) from cynomolgus monkeys infected with SHIV89.6P. Continuous expression of Gag and Env proteins was detected in stably transduced BLCL, which induced Gag- or Env-specific T cell responses, as measured by both IFNgamma-ELISPOT and chromium release assays, upon in vitro stimulation of PBMC from the SHIV89.6P-infected monkeys. Moreover, induction of Gag-specific CTL using BLCL transduced with retroviral vector expressing the SIV-Gag protein was more efficient and specific compared to that obtained using BLCL infected with a recombinant vaccinia virus (rVV) encoding for the same antigen. Assays on purified CD4+ and CD8+ T cells indicated that both populations specifically produced IFNgamma, but only the CD8+ T cells mediated Gag- and Env-specific cytotoxicity, indicating preferential expansion of these effector cells. Thus, this method represents an alternative tool for the analysis of CTL responses during vaccination protocols in those animal models where little information is available on MHC class I alleles or CTL epitopes.
authors.affiliation: Laboratory of Virology, Retrovirus Division, Istituto Superiore di Sanità, Viale Regina Elena, 299, 00161 Rome, Italy.
authors.forename: Donatella R M
authors.initials: DR
authors.lastname: Negri
authors.forename: Viviana
authors.initials: V
authors.lastname: Buffa
authors.forename: Pasqualina
authors.initials: P
authors.lastname: Leone
authors.forename: Roberta
authors.initials: R
authors.lastname: Bona
authors.forename: Martina
authors.initials: M
authors.lastname: Borghi
authors.forename: Francesca
authors.initials: F
authors.lastname: Carlini
authors.forename: Fabiana
authors.initials: F
authors.lastname: De Angelis
authors.forename: Stefano
authors.initials: S
authors.lastname: Indraccolo
authors.forename: Barbara
authors.initials: B
authors.lastname: Ensoli
authors.forename: Andrea
authors.initials: A
authors.lastname: Cara
mesh_terms: D000818:Animals; D000956:Antigens, Viral / Q000235:genetics / Q000276:immunology; D018414:CD8-Positive T-Lymphocytes / Q000166:cytology / Q000276:immunology*; D002455:Cell Division; D002478:Cells, Cultured; D015683:Gene Products, gag / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D005822:Genetic Vectors / Q000235:genetics*; D006678:HIV / Q000235:genetics* / Q000276:immunology*; D015488:HIV Antigens / Q000235:genetics / Q000276:immunology; D015699:HIV Envelope Protein gp120 / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D008252:Macaca fascicularis; D008297:Male; D012680:Sensitivity and Specificity; D015302:Simian Immunodeficiency Virus / Q000235:genetics* / Q000276:immunology*; D013602:T-Lymphocytes, Cytotoxic / Q000166:cytology / Q000276:immunology
chemical_list: D000956:Antigens, Viral; D015683:Gene Products, gag; D015488:HIV Antigens; D015699:HIV Envelope Protein gp120