T cell activation and inhibition by cholera toxin and its B subunit

Abstract

Cholera toxin (CT) and its B subunit (CT-B) are potent oral immunogens in vivo, although both strongly inhibit polyclonal lymphocyte activation in vitro. In order to help understand this paradox, we have studied the activation of CT-B specific T cells in vitro. CT-B subunit primed PLN T cells were used as responders, Con A stimulated PEC as antigen presenting cells (APC), and various forms of CT-B as antigen. There was a marked difference in the activation of CT-specific T cells when different forms of CT-B were used. Native CT-B did not stimulate T cell proliferation when co-cultured with primed T cells and APC’s; in contrast, denatured CT-B or CT-B blocked by Gml ganglioside stimulated well. The degree of proliferation was proportional to the dose of antigen and APC’s in the cultures, was antigen-specific, and was H-2 restricted. APC’s from high responder strains to CT were significantly more effective than were APC’s from low responder strains in presenting CT-B to F1 T cells. Addition of native CT-B to co-cultures of primed T cells plus denatured CT-B inhibited the CT-B specific proliferative response. Thus the inhibitory properties of CT-B and CT extends even to T cells specific for CT itself. The nature of this inhibition was further investigated using phorbol myristic acetate (PMA) and ionomycin as polyclonal activators, which trigger the two arms of the phosphatidylinositol system. CT-B inhibited T-cell proliferation induced by PMA and ionomycin and the kinetics of such inhibition indicate that CT-B and CT affect late events in T-cell activation. The inhibition occurred also after relatively brief pulses of CT. The CD8 T cell subset was more sensitive to inhibition than was the CD4. We conclude that, because of these inhibitory properties, processing of CT to non-binding molecular forms or fragments in vivo must be an important prerequisite for the induction of an immune response to CT.