Cell-free supernatant from B. lactis INL1 modulates immune cell function and in fluences colorectal cancer cell viability

Abstract

Probiotic-derived metabolites have emerged as promising agents for modulating immune responses and shaping the tumor microenvironment. In this study, we evaluated whether the cell-free supernatant (CFS) of Bifidobacterium animalis subsp. lactis INL1 (B. lactis INL1) (agreement UNS-UNL N°REC-1092496-2), a strain isolated from breast milk, can modulate events associated with these effects. Initially, THP-1 monocytic cells, either undifferentiated or differentiated into macrophage-like cells using phorbol 12-myristate 13-acetate (PMA), were used to assess phagocytic capacity following pre-treatment with B. lactis INL1. Using a gentamicin protection assay with E. coli Nissle 1917 constitutively expressing the red fluorescent protein mKate (EcN-mKate) as the phagocytic target, we observed that PMA-differentiated THP-1 cells exhibited enhanced phagocytic uptake of EcN-mKate following exposure to CFS. This was confirmed by light microscopy and drop-CFU assays. Additionally, we evaluated phagocytosis in Peripheral Blood Mononuclear Cells (PBMCs) isolated from healthy donors, observing a comparable increase in their phagocytic index. We next investigated whether CFS exposure altered the cytokine profile of THP-1 cells and PBMCs. Using a multiplex immunoassay, we found that pre-treatment with CFS under LPS-induced inflammatory conditions modulated the release of key immune cytokines, including IL-6, IL-1β, and IL-10, while others, such as TNF-α and CCL2 showed no significant changes. These effects were cell-type-specific. Finally, conditioned media obtained from THP-1 cells and PBMCs pre-treated with CFS and exposed to LPS reduced the viability of colorectal carcinoma MC38 cells. This suggests that immune cell function and immune-tumor cell interactions may be modulated by metabolites derived from B. lactis INL1