title: T-cell receptor beta-chain usage in directly activated alloreactive CD4+ T cells: unrestricted Vbeta gene usage with CDR3 size limitations.
abstract: BACKGROUND
  Alloreactive immune responses may engage both direct and indirect antigen allorecognition. This study focuses on T-cell receptor (TCR) beta-chain usage by in vitro generated alloreactive CD4+ T cells involved in direct allorecognition pathway.
  
  
  RESULTS
  We have established Lewis anti-Brown Norway rat CD4+ T-cell lines and confirmed their reactivities against cell-surface, but not soluble, alloantigens. TCR Vbeta-specific reverse-transcriptase polymerase chain reaction detected all 22 Vbeta genes in these cell lines at all stages, regardless of the lengths of in vitro stimulation. By using spectrotyping, we found that Vbeta complementarity determining region (CDR)3 length distribution pattern altered dramatically after repeated allostimulation. Such a skewed CDR3 distribution occurred in most Vbeta genes without any obvious preference, indicating that expansion occurred in all Vbeta expressing cells by allostimulation. Fluorescence-activated cell sorter analysis of Vbeta expression in alloreactive CD4+ T-cell lines with anti-Vbeta-specific monoclonal antibodies showed, quantitatively, similar percentages of individual Vbeta-expressing cells in the alloreactive pool after repeated allostimulation. To test whether preferential TCR Vbeta gene usage occurred in "high responder" cells, we sorted CD4+ T cells that underwent three or more divisions from primary mixed leukocyte reactions. Unlimited Vbeta usage with CDR3 alterations was observed, as in unsorted alloreactive T cells.
  
  
  CONCLUSIONS
  TCR Vbeta gene usage in directly alloreactive CD4+ T-cell population is unrestricted. Clonal expansion occurs in all Vbeta expressing T cells by allostimulation.
authors.affiliation: Dumont-UCLA Transplant Center, UCLA School of Medicine, Los Angeles, CA 90095, USA.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Zhai
authors.forename: Jerzy W
authors.initials: JW
authors.lastname: Kupiec-Weglinski
mesh_terms: D000818:Animals; D015496:CD4-Positive T-Lymphocytes / Q000166:cytology / Q000276:immunology*; D002460:Cell Line; D022801:Complementarity Determining Regions / Q000276:immunology*; D005434:Flow Cytometry; D007135:Immunoglobulin Variable Region / Q000276:immunology; D051381:Rats; D011914:Rats, Inbred BN; D011917:Rats, Inbred Lew; D011920:Rats, Inbred WF; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000276:immunology*; D013154:Spleen / Q000166:cytology / Q000276:immunology; D014181:Transplantation Immunology / Q000276:immunology*
chemical_list: D022801:Complementarity Determining Regions; D007135:Immunoglobulin Variable Region; D016693:Receptors, Antigen, T-Cell, alpha-beta