Clonal analysis of CD4-CD8- human thymocytes expressing a T cell receptor gamma/delta chain. Direct evidence for the de novo expression of CD8 surface antigen and of cytolytic activity against tumor targets.
CD4-CD8- human thymocytes were obtained by treating total thymocyte suspensions with anti-CD4 and anti-CD8 monoclonal antibodies (mAb) and complement. The resulting cell populations contained virtually no CD4+, CD8+ or WT31+ cells and 17-65% CD3+ cells. In addition, analysis of cell reactivity with delta-TCS-1 mAb (specific for the C gamma 2-encoded, nondisulfide-linked form of TcR gamma/delta), revealed the presence of a variable proportion of delta-TCS-1+ cells (the % of delta-TCS-1+ cells were lower than the percentage of CD3+ cells). Upon culture in recombinant interleukin 2 (IL2, in the presence of irradiated mononuclear cells), CD4-CD8- thymocytes underwent extensive proliferation. In addition, a progressive increase of CD8+ cells (but not of CD4+ or WT31+ cells) could be detected. Cells also progressively acquired cytolytic activity against K-562 or fresh melanoma cells. Fresh CD4-CD8- thymocytes were cloned under limiting dilution conditions. The cloning efficiencies were relatively high (1/3 cells); in addition, virtually all the clonal progenies obtained displayed cytolytic activity and expressed the CD3+WT31-delta-TCS-1+ surface phenotype. About half of the clones analyzed were CD8+, whereas none expressed CD4 antigens. We conclude that (a) only delta-TCS-1-reactive, TcR gamma/delta+ cells can be isolated from CD4-CD8- thymocytes cultured in IL2, and (b) the expression of CD8 antigen and of cytolytic activity reflects a true in vitro phenotypic change of CD8-, noncytolytic precursors (and not the preferential growth of few contaminating cells).
Mingari
M C
MC
Istituto Nazionale per la Ricerca sul Cancro, Genova, Italy.
Varese
P
P
Bottino
C
C
Melioli
G
G
Moretta
A
A
Moretta
L
L
D000945:Antigens, Differentiation, T-Lymphocyte / Q000032:analysis* / Q000502:physiology*; D016827:CD8 Antigens; D002454:Cell Differentiation; D003602:Cytotoxicity, Immunologic; D006801:Humans; D011948:Receptors, Antigen, T-Cell / Q000032:analysis*; D016692:Receptors, Antigen, T-Cell, gamma-delta; D013601:T-Lymphocytes / Q000166:cytology* / Q000276:immunology; D013950:Thymus Gland / Q000166:cytology*
D000945:Antigens, Differentiation, T-Lymphocyte; D016827:CD8 Antigens; D011948:Receptors, Antigen, T-Cell; D016692:Receptors, Antigen, T-Cell, gamma-delta