Bifidobacterium bifidum causes an enhancement of the intestinal epithelial tight junction barrier is mediated by TLR-2 dependent increases of Toll-interacting protein (TOLLIP) expression in a MYD88 independent manner
Authors: Abdulqadir Raz , Al-Sadi Rana , Ma Thomas 
Published: May 2023

Background: Bifidobacteria are commonly utilized probiotics that have been shown to protect against intestinal inflammation in part by enhancing the intestinal epithelial tight junction (TJ) barrier function. It has been recently shown that Bifidobacterium bifidum enhances the intestinal epithelial TJ barrier in a strain-specific and toll-like receptor-2 (TLR-2) dependent manner. Toll-interacting protein (TOLLIP) is a potent inhibitory, cytosolic adaptor protein that represses myeloid differentiation primary response 88 (MYD88). However, the association between BB, TLR-2, and TOLLIP in mediating the BB enhancement of intestinal epithelial TJ barrier remains unclear. Aims: The major aim of this study was to delineate the mechanisms that mediate BB enhancement of the intestinal TJ barrier by examining the interaction of BB with the membrane receptor TLR-2 and the potent TLR modulator TOLLIP in the intestinal epithelial cells. Methods: An in-vitro intestinal epithelial model system consisting of filter-grown Caco-2 monolayers was used, and various biochemical methods and confocal microscopy were applied. Results: 1) A specific strain of BB (1x108 CFU/ml) referred to as BB1 caused a marked increase in Caco-2 transepithelial resistance (TER) (80% increase) and a decrease in trans-epithelial flux of paracellular marker. Other BB strains examined (five total strains including BB4) caused a modest increase to (25%-1%) no changes in Caco-2 TER, suggesting that BB enhancement of Caco-2 TJ barrier function was strain-specific. Strain BB4 had no effect on Caco-2 TJ barrier function. 2) BB1 but not BB4 caused an increase in TLR-2 expression. 3) BB1 induced increase of TLR-2 expression was independent of MYD88, and MYD88 was not required for the BB1-induced enhancement of intestinal TJ barrier. 4) BB1, not BB4, caused an activation of TOLLIP expression 4) The siRNA-induced knockdown of TLR-2 prevented the BB1-induced increase in TOLLIP expression and recruitments to apical membrane junctions, and also prevented the BB1 induced increase in Caco-2 TER and drop in inulin flux. 5) The siRNA-induced knockdown of TOLLIP prevented the BB1-induced increase in Caco-2 TER and drop in inulin flux, and inhibition of IRAK-1 kinase activity. Conclusion: These studies provide a novel insight into the mechanisms that mediate BB1 enhancement and protection of intestinal TJ barrier; our data show for the first time that TLR-2/TOLLIP plays an integral role in BB1 enhancement of TJ barrier function. The BB-induced enhancement of the intestinal epithelial TJ barrier was mediated by TLR-2 dependent signaling recruitment of TOLLIP, and BB enhancement of the TJ barrier was blocked by targeted knockdown of TLR-2 and TOLLIP. TOLLIP represent a potential novel therapeutic target for IBD. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.