Saccharomyces boulardii (S6) is a non-pathogenic yeast that ameliorates intestinal injury and inflammation caused by a wide variety of enteric pathogens. We hypothesized that Sb may exert its probiotic effects by modulation of host cell signaling and pro-inflammatory gene expression. Human HT-29 colonocytes and THP-1 rnonocytes were stimulated with IL-1 beta, TNF alpha or LPS in the presence or absence of Sb culture supernatant (SbS). IL-8 protein and mRNA levels were measured by ELISA and RT-PCR, respectively. The effect of SbS on I kappa B alpha degradation was studied by Western blotting and on NF-kappa B-DNA binding by EMSA. NF-kappa B-regulated gene expression was evaluated by transient transfection of THP-1 cells with a NF-kappa B-responsive luciferase reporter gene. SbS inhibited IL-8 protein production in IL-1 beta or TNF alpha stimulated HT-29 cells (by 75% and 85%, respectively; P < 0.001) and prevented IL-1 beta-induced up-regulation of IL-8 mRNA. Sbs also inhibited IL-8 production, prevented I kappa B alpha degradation, and reduced both NF-kappa B-DNA binding and NF-kappa B reporter gene up-regulation in IL-1 beta or LPS-stimulated THP-1 Cells. Purification and characterization studies indicate that the S. boulardii anti-inflammatory factor (SAIF) is small (< 1 kDa), heat stable, and water soluble. The probiotic yeast Saccharomyces boulardii exerts an anti-inflammatory effect by producing a low molecular weight soluble factor that blocks NF-kappa B activation and NF-kappa B-mediated IL-8 gene expression in intestinal epithelial cells and monocytes. SAIF may mediate, at least in part, the beneficial effects of Saccharomyces boulardii in infectious and non-infectious human intestinal disease. (c) 2006 Published by Elsevier Inc.