Production of tumor necrosis factor alpha in human T lymphocytes by staphylococcal enterotoxin B correlates with toxin-induced proliferation and is regulated through protein kinase C

The superantigen staphylococcal enterotoxin B (SEB) simultaneously binds bo th the major histocompatibility complex (MHC) class II receptor on monocyte s and the T-cell receptor (TCR) on T lymphocytes, resulting in a range of c ell responses including induction of tumor necrosis factor alpha (TNF-alpha ). In this study, we have used mixed cultures of human peripheral blood mon ocytes and lymphocytes to investigate biochemical events controlling SEE in duction of TNF-alpha. TNF-alpha production induced by SEE in mixed cultures is more closely associated with T cells than with monocytes: (i) a TCR-bin ding-site mutant of SEE (N23F) is less active in TNF-alpha. induction than an MHC class II receptor-binding-site mutant (P44R), and (ii) flow cytometr ic analysis indicated that SEE induced TNF-alpha production in T cells but not in monocytes. Pretreatment of cells with inhibitors of signal transduct ion pathways was employed to further define events in SEE-induced TNF-alpha production. Neither protein kinase A inhibitors nor two protein tyrosine k inase inhibitors altered SEE-induced TNF-alpha production. In contrast, SEE induced protein kinase C (PI(C) translocation, and pretreatment of culture s with inhibitors of PKC blocked TNF-alpha induction, Alteration of levels of diacylglycerol (DAG), an activator of PKC, by treatment with inhibitors of phospholipase C or DAG kinase also altered SEE-induced TNF-alpha product ion. These data suggest that PKC activation plays a critical role in SEE-in duced TNF-alpha production in human T cells.