Donna S. Stone-Wolff

Stimulation of lymphokine production by teleocidin, aplysiatoxin, and debromoaplysiatoxin

Previous studies showed that the tumor-promoting phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA) and several structurally related tumor-promoting compounds stimulate lymphocytes to produce immune interferon (IFN-gamma) and interleukin 2 (IL-2). This study shows that three compounds structurally unrelated to TPA, previously shown to mimic TPA in some other biological activities, are similar to TPA in stimulating IFN-gamma and Il-2 production in cultures of human peripheral blood lymphocytes. The production of another lymphokine, termed lymphotoxin (LT), was also enhanced by TPA and the other three compounds examined. Maximal enhancement of lymphokine production was observed in cultures costimulated with TPA or one of the other tested compounds and phytohemagglutinin (PHA). TPA was separated from IFN-gamma during a multistep purification procedure.

STRUCTURE-FUNCTION STUDIES WITH HUMAN INTERFERON-GAMMA

ABSTRACT Interferon-gamma, produced in cultures of human white blood cells by combined stimulation with a phorbol ester and phytohemagglutinin (PHA), was purified about 1,000-fold in a simple 3-step procedure. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) showed that interferon activity not destroyed by SDS treatment was recovered mainly from two peaks with apparent molecular weights of 20,000 and 25,000. The two molecular weight subspecies, isolated in pure form, were antigenically indistinguishable. Highly purified interferon-gamma was iodinated with the 125-I-Bolton-Hunter reagent. This material bound with a high affinity (Kd 7.5 × 10 −11 M) to human fibroblasts in culture. Unlabeled human interferon-gamma or -beta, but not interferon-alpha, competitively inhibited the specific binding of 125I-interferon-gamma. Crude interferon-gamma preparations had a pronounced cytotoxic effect on HeLa and FS-4 cells, while highly purified interferon-gamma was effective only at high concentrations. Furthermore, only in HeLa cells did it show a greater antiproliferative effect than interferon-alpha or -beta.