The synergistic effect of phytohemagglutinin and interferon-gamma on the expression of tumor necrosis factor-alpha from RAW 264.7 cells.

Tumor necrosis factor-alpha (TNF-alpha) is a major cytokine of host immune reaction by foreign agents. Phytohemagglutinin (PHA) is a dynamic contributor to mitogenic stimulation and augmentation of host immune defense. Interferon-gamma (IFN-gamma) induces induction of cytokines in macrophages and lymphocytes. The aim of this study was to examine the synergistic effects of PHA plus low dose IFN-gamma on TNF-alpha mRNA production, cytosolic levels, and secretion in RAW 264.7 cells. The cells were stimulated with PHA or IFN-gamma using various concentrations for various times. The effects of PHA on TNF-alpha expression appeared in dose- and time-dependent manners. The maximum doses of PHA and IFN-gamma to produce them were 300 microg/ml PHA and 10 ng/ml IFN-gamma. The optimum time of PHA for the TNF-alpha mRNA production and release were 6 and 7 h after stimulation, respectively, whereas the time of IFN-gamma on them was achieved at 3 and 8 h. Although the TNF-alpha mRNA production, cytosolic levels, and secretion from the cells were slightly detected under 10 microg/ml PHA and 1 ng/ml IFN-gamma, the combination of PHA (10 microg/ml) and IFN-gamma (1 ng/ml) greatly increased them, indicating the synergistic effect of PHA plus low dose IFN-gamma on TNF-alpha expression.