Interleukin-12 primes human CD4 and CD8 T cell clones for high production of both interferon-gamma and interleukin-10.

Interleukin-12 (IL-12) induces differentiation of T helper 1 (Th1) cells, primarily through its ability to prime T cells for high interferon-gamma (IFN-gamma) production. We now report that the presence of IL-12 during the first several days of in vitro clonal expansion in limiting dilution cultures of polyclonally stimulated human peripheral blood CD4+ and CD8+ T cells also induces stable priming for high IL-10 production. This effect was demonstrated with T cells from both healthy donors and HIV+ patients. Priming for IL-4 production, which requires IL-4, was maximum in cultures containing both IL-12 and IL-4. IL-4 modestly inhibited the IL-12-induced priming for IFN-gamma, but almost completely suppressed the priming for IL-10 production. A proportion of the clones generated from memory CD45RO+ cells, but not those generated from naive CD45RO- CD4+ T cells, produced some combinations of IFN-gamma, IL-10, and IL-4 even in the absence of IL-12 and IL-4, suggesting in vivo cytokine priming; virtually all CD4+ clones generated from either CD45RO(-) or (+) cells, however, produced high levels of both IFN-gamma and IL-10 when IL-12 was present during expansion. These results indicate that each Th1-type (IFN-gamma) and Th2-type (IL-4 and IL-10) cytokine gene is independently regulated in human T cells and that the dichotomy between T cells with the cytokine production pattern of Th1 and Th2 cells is not due to a direct differentiation-inducing effect of immunoregulatory cytokines, but rather to secondary selective mechanisms. Particular combinations of cytokines induce a predominant generation of T cell clones with anomalous patterns of cytokine production (e.g., IFN-gamma and IL-4 or IFN-gamma and IL-10) that can also be found in a proportion of fresh peripheral blood T cells with "memory" phenotype or clones generated from them and that may identify novel Th subsets with immunoregulatory functions.

In vivo modification of major histocompatibility complex class II DRA promoter occupancy mediated by the AIR-1 trans-activator.

RJ 2.2.5 is a human B cell mutant derived from the Burkitt lymphoma Raji cell which is defective in the AIR-1 locus function. This locus encodes a transcriptional trans-activator required for the constitutive expression of major histocompatibility complex (MHC) class II genes. Here we show, by in vivo DNase I footprinting, that the AIR-1 locus defect correlates with changes in the DRA promoter occupancy. Interestingly, reexpression of human MHC class II genes in RJ 2.2.5 x mouse spleen cell hybrids is associated with partial reversion of DRA promoter occupancy to the Raji pattern. DRA promoter occupancy in other class II-negative B cell lines, derived from patients with bare lymphocyte syndrome, is drastically different from the one observed in RJ 2.2.5 and Raji cells. Moreover, the use of the DNase I as an in vivo footprinting agent reveals that the patients' cell lines do not display a completely "bare promoter" as previously reported using dimethyl sulfate as the footprinting agent. Thus, the use of DNase I allowed us, for the first time, to correlate the AIR-1 locus defect with class II promoter occupancy alterations and distinguish these alterations from the ones observed in phenotypically similar but genetically distinct MHC class II-negative cells.

Staphylococcal enterotoxin-dependent lysis of MHC class II negative target cells by cytolytic T lymphocytes.

The enterotoxins of Staphylococcus aureus (SE) are extremely potent activators of human and mouse T lymphocytes. In general, T cell responses to SE are MHC class II dependent (presumably reflecting the ability of SE to bind directly to MHC class II molecules) and restricted to responding cells expressing certain T cell receptor beta-chain variable (TCR V beta) domains. Recently we demonstrated that CD8+ CTL expressing appropriate TCR V beta could recognize SE presented on MHC class II-bearing target cells. We now show that MHC class II expression is not strictly required for T cell recognition of SE. Both human and mouse MHC class II negative target cells could be recognized (i.e., lysed) in a SE-dependent fashion by CD8+ mouse CTL clones and polyclonal populations, provided that the CTL expressed appropriate TCR V beta elements. SE-dependent lysis of MHC class II negative targets by CTL was inhibited by mAb directed against CD3 or LFA-1, suggesting that SE recognition was TCR and cell contact dependent. Furthermore, different SE were recognized preferentially by CTL on MHC class II+ vs MHC class II- targets. Taken together, our data raise the possibility that SE binding structures distinct from MHC class II molecules may exist.

Inducible and constitutive MHC class II gene expression. Distinct tissue-specific genetic controls.

B cells express MHC class II Ag in a constitutive fashion, whereas macrophages can do so only after induction by a variety of exogenous stimuli. In this study we describe interspecies somatic cell hybrids between the human B cell Raji and the murine macrophage cell P388 D1. This murine cell line does not express detectable levels of class II mRNA. Phenotypic, molecular, and karyotype analysis of a series of hybrids showed that murine macrophage class II genes can be expressed in a constitutive fashion under the control of the human B cell genome. This event is the consequence of de novo accumulation of class II specific mRNA and thus probably reflects activation of transcription. In certain cases the amount of murine class II Ag expressed on the surface of the hybrid cell was significantly higher than the one observed in the parental macrophage cells after induction with IFN-gamma and was not further modified by treatment with the murine lymphokine. Reversion from a murine class II-positive to class II-negative cell surface phenotype in the hybrids correlated with reduced expression of human markers and more important with segregation of human chromosomes. Interestingly, in this case certain hybrids still expressed detectable levels of murine class II mRNA and increased levels of murine invariant chain mRNA when compared with parental P388 D1 murine macrophage cells. These results indicate that constitutive class II gene expression behaves as a dominant trait in B cell x macrophage somatic cell hybrids. Possible mechanisms responsible of the different control of class II gene expression during cell type differentiation are discussed.