title: [Transgenic expression of T cell receptors on HBV-specific cytotoxic T lymphocytes].
abstract: OBJECTIVE
  To express T lymphocyte receptors (TCRs) on hepatitis B virus (HBV)-specific cytotoxic T lymphocytes (CTLs) mediated by retrovirus and investigate their binding affinity.
  
  
  METHODS
  Peripheral blood mononuclear cells were isolated from acute hepatitis B patients with HLA-A2⁺; phenotype, and after induced, HBV-specific CTLs were sorted out followed by cloning and proliferating. Cell RNAs were extracted. The sequences of TCR's α and β chains were obtained by means of RT-PCR, 5'RACE and OVER-LAP PCR, for constructing TCR retrovirus vectors. Through retrovirus-mediated transduction, HBV-specific TCRs were expressed on Jurkat cells and CD8⁺; T cells from HLA-A⁺; healthy subjects.
  
  
  RESULTS
  Two paired TCR Vα and Vβ, respectively named α21β13 and α15β13, were obtained from one patient with acute hepatitis B and HLA-A2⁺;. The titers of packaged recombinant retroviruses were 1.5×10;-5.0×10⁵; IU/mL. Immunofluorescence staining by anti-Vβ13 TCR-PE targeting the specific TCR and HLA-A2 restricted epitope-specific pentamer showed a positive expression of reconstructed TCR on T cells. The positive cells accounted for 1.06%-2.25% for Vβ13 on Jurkat cells, 1.03%-2.06% for Vβ13 chain and 1.05%-1.12% for the epitope-specific pentamer on T cells from healthy HLA-A2⁺; subjects respectively. By contrast, only less than 0.05% cells from healthy HLA-A2⁻ ;subjects were positive for either Vβ13 or the pentamer.
  
  
  CONCLUSIONS
  TCRs on HBV-specific CTLs could be expressed by TCR gene transfer mediated by retrovirus, and they were proved with binding affinity to HLA-A2-restricted epitope.
authors: Guilin Medical University, China. wujingreference@q63.com
authors: Jing
authors: J
authors: Wu
authors: Lin
authors: L
authors: Wang
authors: Yan
authors: Y
authors: Liu
authors: Haiyan
authors: H
authors: Ye
authors: Ning
authors: N
authors: Ding
authors: Yongming
authors: Y
authors: Liu
authors: Dongping
authors: D
authors: Xu
mesh_terms: D000595:Amino Acid Sequence; D018984:Epitopes, T-Lymphocyte; D015870:Gene Expression; D023061:Gene Order; D005822:Genetic Vectors / Q000235:genetics; D006515:Hepatitis B virus / Q000276:immunology*; D006801:Humans; D016130:Immunophenotyping; D019169:Jurkat Cells; D008969:Molecular Sequence Data; D011948:Receptors, Antigen, T-Cell / Q000235:genetics* / Q000378:metabolism; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000737:chemistry / Q000235:genetics; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology* / Q000378:metabolism*
chemical_list: D018984:Epitopes, T-Lymphocyte; D011948:Receptors, Antigen, T-Cell; D016693:Receptors, Antigen, T-Cell, alpha-beta