Effect of Lactobacillus GG and conditioned media on IL-1beta-induced IL-8 production in Caco-2 cells

Abstract

OBJECTIVE: To evaluate the anti-inflammatory effect of Lactobacillus casei rhamnosus GG (LGG) and its conditioned media (CM) in stimulated Caco-2 cells and to characterize the components of LGG that have the anti-inflammatory effect.

MATERIAL AND METHODS: Caco-2 cells were stimulated with IL-1beta with or without LGG or LGG-CM. Production of IL-8 was measured by enzyme-linked immunosorbent assay (ELISA). The transcriptional activities of the IL-8 gene and the NF-kappaB-responsive gene were evaluated by a transient transfection of the luciferase reporter gene. The effect on IkappaBalpha degradation was evaluated by Western blot analysis. To determine the nature of the immunomodulatory molecules, the LGG was modified to the following: treated with antibiotics, 4% formaldehyde, incubation at 95 degrees C, or sonication.

RESULTS: We demonstrated that the pretreatment of Caco-2 cells with LGG significantly inhibited IL-1beta-induced IL-8 production. Furthermore, LGG attenuated the IL-1beta-induced transcriptional activation of the IL-8 gene and the NF-kappaB-responsive gene, and attenuated the IL-1beta-induced IkappaBalpha degradation. Formaldehyde-fixed or antibiotics-treated LGG maintained the inhibitory effect, but heated LGG lost this effect. Sonicated LGG debris had a similar inhibitory effect with whole bacterial cells. LGG-CM attenuated IL-1beta-induced IL-8 production. This effect was maintained even when the conditioned media were heated.

CONCLUSIONS: LGG inhibited IL-1beta-induced IL-8 production in Caco-2 and this effect occurred at the transcriptional level, at least in part, by inhibition of the NF-kappaB signaling pathway. Both the structural material of LGG and the soluble factor secreted from LGG inhibited the IL-1beta-induced IL-8 production, and thus different substances may cause the effects.