Phorbol ester induces a differential effect on the effector function of human allospecific cytotoxic T lymphocyte and natural killer clones.

Abstract

Six-day allosensitized human peripheral blood lymphocytes treated for 12-18 hr with phorbol 12-myristate 13-acetate (PMA) were found to lose the capacity to kill the specific target in a standard cell-mediated lympholysis (CML) assay, but they were still effective in their ability to kill the tumor cell line K562. We investigated which antigens on the cell membrane involved in alloimmune recognition might be modified by PMA, since it was found that in a lectin-dependent CML assay, the lytic mechanism was not impaired. For this purpose, T3V, T4' allospecific cytotoxic T lymphocyte (CTL) clones and either T3+ or T3-natural killer (NK) clones were generated from 6-day allostimulated lymphocytes. The results indicated that PMA inhibited the cytolytic function of both alloimmune CTL and NK T3+ clones. In contrast, PMA did not modify the effector cell function of T3-NK clones. Phenotypic analysis of T-cell surface antigens from T3+ clones showed that T3 molecule expression on the cell membrane was reduced by 80-90% after PMA treatment, whereas expression of both accessory T4 molecules, involved in antigen recognition, and receptor for interleukin 2 was increased. Moreover, the loss of function was transitory and could be restored 4 days after PMA treatment when the T3 molecules were fully reexpressed at the cell surface. Taken together, these data strongly suggest that (i) PMA prevents cell-mediated cytotoxicity by modulating the disulfide-linked heterodimer associated to T3 and described as the receptor for antigen on the cell surface of major histocompatibility complex (MHC) and non-MHC specific CTL clones, without affecting the lytic mechanism per se, and (ii) the expression of the receptor for the antigen present on the tumor cell line K562 is not decreased on T3-NK clones after PMA treatment and must be different from that on T3+ T-cell clones.Tumor-promoting phorbol esters are chemical agents known to induce tumors in vivo when applied to animals that have received low doses of a carcinogen (1). When used in vitro on malignant cells or normal cell lines, they were found to act on cell differentiation (2, 3) and or activation (4). Recently, it has been shown that phorbol esters in the presence of anti-T3 antibodies were able to induce interleukin 2 (IL-2) and interferon y (IFN-y) production in some human T-cell lines (5) and also peripheral blood T-cell proliferation in the absence of monocytes (6). Specific proliferation of normal human T lymphocytes is regulated via the cell-surface receptor expression for the antigen. The basic antigen/major histocompatibility complex (MHC) recognition unit of the T cell has been defined as a clonotypic, disulfide-linked heterodimer (Ti) associated with T3 molecules. T3-Ti receptor triggering by the specific antigen leads to IL-2 receptor (IL-2R) expression on the cell membrane and IL-2 production by some T-cell subsets (7). Following IL-2 molecule-IL-2R interaction, DNA synthesis and cell division occur (8). During the process of lymphocyte activation by specific a...

The surface phenotype of activated T lymphocytes

“…The amount of complex expressed on the surface of a developing T cell is often used as a gauge of maturity, with cells expressing high levels considered mature, and those expressing little or no CD3 considered immature. The transient decrease in CD3 expression observed in the present study is consistent with published reports of CD3 expression on activated human T cells (4,(6)(7)(8). This observation means that low level expression of CD3 may indicate an activated cell and not necessarily an immature one.…”

“…double positive state on activation (2), but in a recent study this was found not to occur (3). Several recent publications have suggested that CD3, CD4 and CD8 may be lost from the cell surface of human T cells upon activation in vitro (4)(5)(6)(7)(8)(9). Since T cells have been reported to be capable of returning to the thymus from the secondary lymphoid organs (10), this raises questions about the source of the minor subpopulations of double negative cells in the thymus (11,12).…”

A Soluble Form of the MHC Class I-Specific CD160 Receptor Is Released from Human Activated NK Lymphocytes and Inhibits Cell-Mediated Cytotoxicity

“…PB-NK cell purity was shown to be Ͼ90%. The selection of in vitro allogeneic-MHC-I-restricted effector T lymphocytes was performed as previously reported (31). Briefly, PBMC were cocultured for 6 days with irradiated EBV-transformed B cells in RPMI 1640 medium supplemented with penicillin (100 IU/ml), streptomycin (100 g/ml), L-glutamine (2 mM), and 10% heat-inactivated human serum (Jacques Boy Institute, Lyon, France).…”

Interferon: a cytotoxic T lymphocyte differentiation signal

“…After a 6-day culture the primed cells were restimulated for an additional 6 days with the specific stimulating cells in the presence of 10% human serum (HS)/RPMI containing 10% IL 2-conditioned medium [16]. The primed cells were then cloned by micromanipulation.…”