Shear-Enhanced Dynamic Adhesion of Lactobacillus rhamnosus GG on Intestinal Epithelia: Correlative Effect of Protein Expression and Interface Mechanics.

The oral uptake of probiotic microorganisms as food additives is one widely used strategy to sustain and improve the homeostasis of intestinal microbiota that protect the intestinal epithelia from attack by pathogenic bacteria. Once delivered to the ileum and colon, probiotics must adhere and form colonies on mucus that coats the surface of intestinal epithelial cells. Although an increasing amount of knowledge about the genetic and molecular level mechanisms of probiotics-mucus interactions has been accumulated, little is known about the physicochemical aspects of probiotics-mucus interactions under physiological shear in intestines. In this study, we established well-defined models of intestinal epithelial cell monolayers based on two major constituents of gut epithelia, enterocytes and goblet cells. First, the formation of a polarized cell monolayer sealed by tight junctions was monitored by transepithelial electrical resistance over time. The establishment of tight junctions and secretion of mucus proteins (mucin) was confirmed by immunofluorescence staining. In the next step, we measured the elasticity of cell monolayer surfaces by indentation using particle-assisted atomic force microscopy. The effective elastic modulus of goblet cell-like cells was 30 times smaller compared to that of enterocyte-like cells, which can be attributed to the secretion of a 3 µm thick mucin layer. As probiotics, we used Lactobacillus rhamnosus GG (LGG), which is one of the most widely used strains as food additives. To investigate the dynamic adhesion of LGG to the intestine model surface, we transferred the epithelial cell monolayer into a microfluidic chamber. A distinct difference in dynamic adhesion between two cell types was observed, which could be attributed to the difference in the mucin expression amount. Remarkably, we found that the dynamic LGG adhesion is enhanced by the increase in shear stress, showing a maximum binding efficiency at 0.3 Pa. Finally, we examined the persistence of LGG adhesion by a stepwise increase in the shear stress exerted on adherent LGG, demonstrating that LGG could withstand high shear stress even beyond that of physiological stress. The obtained results present a large potential to quantitatively understand the influence of engineered foods and probiotics on the homeostasis of microbiota on the surface of intestinal epithelia.

Biopolymer-Based Minimal Formulations Boost Viability and Metabolic Functionality of Probiotics Lactobacillus rhamnosus GG through Gastrointestinal Passage.

The delivery of probiotic microorganisms as food additives via oral administration is a straightforward strategy to improve the intestinal microbiota. To protect probiotics from the harsh environments in the stomach and small intestine, it is necessary to formulate them in biocompatible carriers, which finally release them in the ileum and colon without losing their viability and functions. Despite major progresses in various polymer-based formulations, many of them are highly heterogeneous and too large in size and hence often "felt" by the tongue. In this study, we established a new formulation for probiotics Lactobacillus rhamnosus GG (LGG) and systematically correlated the physicochemical properties of formulations with the functions of probiotics after the delivery to different gastrointestinal compartments. By reducing the stirring speed by 1 order of magnitude during the emulsification of polyalginate in the presence of xanthan gum, we fabricated microparticles with a size well below the limit of human oral sensory systems. To improve the chemical stability, we deposited chitosan and polyalginate layers on particle surfaces and found that the deposition of a 20 nm-thick layer is already sufficient to perfectly sustain the viability of all LGG. Compared to free LGG, the colony-forming units of LGG in these formulations were by factors of 107 larger in stomach fluid and 104 larger in small intestine fluid. The metabolic functionality of LGG in polymer formulations was assessed by measuring the amount of lactate produced by LGG in a human gastrointestinal simulator, showing 5 orders of magnitude larger values compared to free LGG. The obtained results have demonstrated that the minimal formulation of LGG established here boosts not only the viability but also the metabolic functionality of probiotics throughout oral uptake, passage through the gastrointestinal tract, and delivery to the ileum and colon.

Intestinal iron bio-accessibility changes by Lignin and the subsequent impact on cell metabolism and intestinal microbiome communities.

The detrimental effects of high concentrations of colonic iron have been linked to intestinal inflammation and microbial dysbiosis. Exploiting chelation against this luminal pool of iron may restore intestinal health and have beneficial impacts on microbial communities. This study aimed to explore whether lignin, a heterogenous polyphenolic dietary component, has iron-binding affinity and can sequester iron within the intestine and thus, potentially modulate the microbiome. Within in vitro cell-culture models, the treatment of RKO and Caco-2 cells with lignin almost abolished intracellular iron import (96% and 99% reduction of iron acquisition respectively) with corresponding changes in iron metabolism proteins (ferritin and transferrin receptor-1) and reductions in the labile-iron pool. In a Fe-59 supplemented murine model, intestinal iron absorption was significantly inhibited by 30% when lignin was co-administered compared to the control group with the residual iron lost in the faeces. The supplementation of lignin into a microbial bioreactor colonic model increased the solubilisation and bio-accessibility of iron present by 4.5-fold despite lignin-iron chelation previously restricting intracellular iron absorption in vitro and in vivo. The supplementation of lignin in the model increased the relative abundance of Bacteroides whilst levels of Proteobacteria decreased which could be attributed to the changes in iron bio-accessibility due to iron chelation. In summary, we demonstrate that lignin is an effective luminal iron chelator. Iron chelation leads to the limitation of intracellular iron import whilst, despite increasing iron solubility, favouring the growth of beneficial bacteria.