Purification and characterization of cytokine-inducing protein of seed extract from Aeginetia indica L., a parasitic plant

We have isolated 55 kDa protein from the seed extract of Aeginetia indica L . (AIL), a parasitic plant, by an affinity chromatography on N-hydroxysucci nimide (NHS)-activated Sepharose High Performance column bound F3 monoclona l antibody which neutralizes cytokine-inducing and antitumor effect of AIL. In in vitro model using human peripheral blood mononuclear cells (PBMC), t he 55 kDa protein (AILb-A) induced multiple cytokines, such as IFN-gamma, t umor necrosis factor (TNF)-alpha, granulocyte macrophage-colony stimulating factor (GM-CSF), IL-2, IL-6, IL-IO, IL-12 and IL-18, and also accelerated killer cell activities of PBMC, When compared with a commonly used immunoth erapeutic agent OK-432, AILb-A induced Th1 cytokines are greater than OK-43 2. Of the Th2 cytokines, the amounts of IL-6 and IL-10 induced by AILb-A we re lower than those by OK-432. No significant induction of IL-4 and IL-13 w as observed in AILb-A-stimulated PBMC. TNF family including TNF-alpha, TNF- beta, Fas ligand (FasL) and TNF-related apoptosis-inducing ligand (TRAIL) w ere suggested to be important fur AILb-A-induced killing activity of PBMC b y reverse transcription-polymerase chain reaction (RT-PCR) analysis. Furthe rmore, the neutralizing test using cytokine-specific antibodies demonstrate d that IL-18 plays a most significant role for IFN-gamma- and killer cell-i nducing ability of AILb-A among the cytokines tested. These findings dearly indicated that AILb-A, a 55 kDa protein of AIL, is a potent Th1 cytokine i nducer and may be a useful immunotherapeutic agent for the patients with ma lignancies. (C) 2000 Elsevier Science B.V. All rights reserved.