Immunomodulatory Effects of Bacillus toyonensis BCT-7112T on Macrophage Cell Line RAW 264.7

Abstract

Abstract Bacillus toyonensis is a spore-forming bacterium widely used as a probiotic for animals and humans. It has been reported for its immunomodulatory properties, particularly in enhancing innate and acquired immune responses. One mechanism involved in immune modulation is the capacity of B. toyonensis to stimulate immune cells to produce cytokines. This study investigated the immunomodulatory effects of Bacillus toyonensis BCT-7112T on the macrophage-like cell line RAW 264.7. Cells were stimulated with B. toyonensis vegetative cells (live and killed), spores (viable and non-viable), DNA, and conditioned media for 24, 48, and 72 hours to evaluate proliferation. Additionally, cells were stimulated for 1, 8, and 24 hours and analyzed by qPCR for the transcription of Tnf-α, Il4, Il10, Nuclear Factor kappa Beta (Nfkβ), and Toll-like receptor 2 (Tlr-2). All B. toyonensis stimuli induced cell proliferation, with spores exhibiting the most significant increase (p < 0.05) across all time points. B. toyonensis also induced the transcription of Tnf-α, Il4, Il10 cytokines, Nfkβ, and Tlr-2, with the most pronounced effects (p<0.05) observed following stimulation with viable spores. Our findings suggest that B. toyonensis BCT-7112T has immunomodulatory effects through the activation of murine RAW 264.7 cells, supporting its potential as a model for studying probiotic immunomodulation.