Oxyresveratrol Ameliorates Dextran Sulfate Sodium-Induced Colitis in Rats by Suppressing Inflammation

Colitis causes destruction of the intestinal mucus layer and increases intestinal inflammation. The use of antioxidants and anti-inflammatory agents derived from natural sources has been recently highlighted as a new approach for the treatment of colitis. Oxyresveratrol (OXY) is an antioxidant known to have various beneficial effects on human health, such as anti-inflammatory, antibacterial activity, and antiviral activity. The aim of this study was to investigate the therapeutic effect of OXY in rats with dextran sulfate sodium (DSS)-induced acute colitis. OXY ameliorated DSS-induced colitis and repaired damaged intestinal mucosa. OXY downregulated the expression of pro-inflammatory cytokine genes (TNF-α, IL-6, and IL-1β) and chemokine gene MCP-1, while promoting the production of anti-inflammatory cytokine IL-10. OXY treatment also suppressed inflammation via inhibiting cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) expression in the colon, as well as the activity of myeloperoxidase (MPO). OXY exhibited anti-apoptotic effects, shifting the Bax/Bcl-2 balance. In conclusion, OXY might improve DSS-induced colitis by restoring the intestinal mucus layer and reducing inflammation within the intestine.

Research on the fate of polymeric nanoparticles in the process of the intestinal absorption based on model nanoparticles with various characteristics: size, surface charge and pro-hydrophobics

Background The use of drug nanocarriers to encapsulate drugs for oral administration may become an important strategy in addressing the challenging oral absorption of some drugs. In this study—with the premise of controlling single variables—we prepared model nanoparticles with different particle sizes, surface charges, and surface hydrophobicity/hydrophilicity. The two key stages of intestinal nanoparticles (NPs) absorption—the intestinal mucus layer penetration stage and the trans-intestinal epithelial cell stage—were decoupled and analyzed. The intestinal absorption of each group of model NPs was then investigated. Results Differences in the behavioral trends of NPs in each stage of intestinal absorption were found to result from differences in particle properties. Small size, low-magnitude negative charge, and moderate hydrophilicity helped NPs pass through the small intestinal mucus layer more easily. Once through the mucus layer, an appropriate size, positive surface charge, and hydrophobic properties helped NPs complete the process of transintestinal epithelial cell transport. Conclusions To achieve high drug bioavailability, the basic properties of the delivery system must be suitable for overcoming the physiological barrier of the gastrointestinal tract.

EXPLORING CO-INFECTIONS IN THE GASTROINTESTINAL TRACT: DISSECTING THE INTERACTION BETWEEN FUSOBACTERIUM NUCLEATUM AND CLOSTRIDIODES DIFFICILE

Background Clostridioides difficile is a common healthcare associated pathogen in U.S. hospitals, incurring billions of dollars in treatment costs each year. Microbiome analysis of C. difficile infected (CDI) patients have revealed alterations of the gut microbiota. It has been speculated that select members of this altered microbiota may influence C. difficile pathogenesis. C. difficile is known to reside in the intestinal mucus layer, but at present the interactions between C. difficile and other mucus-associated bacteria are poorly defined. To address these gaps in knowledge, we have focused on an entirely human-centered approach, employing human-derived MUC2, fecal bioreactors and patient samples. We hypothesized that select mucus-associated bacteria would promote C. difficile colonization and biofilm formation. Methods & Results To create a model of the human intestinal mucus layer and gut microbiota, we developed a bioreactor system with human MUC2-coated coverslips. Bioreactors were inoculated with healthy human feces, treated with clindamycin and infected with C. difficile to mimic CDI. C. difficile was found to colonize and form biofilms on MUC2-coated coverslips and 16S rRNA sequencing revealed a unique biofilm profile with substantial co-colonization with Fusobacterium. Consistent with our bioreactor data, publicly available datasets and patient stool samples revealed that a subset of patients with C. difficile infection harbored high levels of F. nucleatum OTUs. We also isolated microbes from adult patients and pediatric IBD patient stool who were positive for C. difficile and F. nucleatum and identified co-localization between these strains. RNAseq data revealed significant changes in C. difficile chemotaxis and surface adhesion genes following exposure to F. nucleatum metabolites. C. difficile was found to co-aggregate with F. nucleatum; an effect that was inhibited by blocking the Fusobacterial adhesin RadD and C. difficile flagella. Moreover, a ΔradD mutant of F. nucleatum lost the ability to aggregate with C. difficile. Conversely, removal of flagella from C. difficile significantly reduced the interaction between WT F. nucleatum and C. difficile. Addition of F. nucleatum also enhanced C. difficile biofilm formation, increasing the levels extracellular polysaccharide. Conclusions Collectively, these data demonstrate the unique role of mucus-associated bacteria such as F. nucleatum in facilitating colonization of the mucus layer by pathogenic C. difficile.

P071 MODULATION OF THE MUCUS LAYER BY BIFIDOBACTERIUM DENTIUM PROVIDES PROTECTION IN A MODEL OF COLITIS

Background The intestinal mucus layer serves as a critical interface between the environment and the host. Patients with inflammatory bowel disease (IBD), particularly ulcerative colitis, exhibit reduced synthesis and secretion of the mucus protein MUC2 and decreased mucus thickness. This in turn promotes immune activation and inflammation. The clinical relevance of the mucus layer emphasizes the need to address strategies to modulate this barrier. Although bifidobacteria represent only 3–6% of the healthy adult fecal microbiota, their presence has been associated with numerous health benefits, including bolstering mucus production. However, the molecular mechanisms that underlie these positive effects appear to be strain-specific and are not well defined. We hypothesized that the human-derived Bifidobacterium dentium would increase intestinal mucus synthesis and expulsion via specific metabolites. We also speculated that modulation of goblet cells would be beneficial during colitis. Methods & Results In silico genome analysis revealed that B. dentium lacked the enzymatic repertoire required for degradation of mucin glycans. Consistent with these findings, we found that B. dentium could not use mucin glycans as a primary carbon source in vitro. To examine mucus modulation in vivo, germ-free mice were mono-associated with live or heat-killed B. dentium. Live B. dentium mono-associated mice exhibited increased colonic expression of goblet cell markers Krüppel-Like Factor 4 (Klf4), Relmβ, trefoil factor 3 (Tff3), Muc2, and several mucin glycosyltransferases compared to both heat-killed B. dentium and germ-free counterparts. Likewise, live B. dentium mono-associated colon had increased acidic mucin-filled goblet cells as denoted by MUC2 and PAS-AB staining. In vitro, B. dentium secreted products, including acetate, were able to increase MUC2 levels in T84 cells, mouse colonoids and human colonoids. We also identified that B. dentium secreted products, such as GABA, stimulated autophagy-mediated calcium signaling and MUC2 release. To identify whether B. dentium could enhance MUC2 production in mice harboring a complete microbiota, specific pathogen free mice were treated with live B. dentium by oral gavage. Administration of B. dentium increased the inner mucus layer compared to controls. Moreover, in a TNBS model of colitis, B. dentium treated mice had increased goblet cell numbers and MUC2 mRNA. Mirroring these findings, B. dentium treated mice lost less weight, had improved histology and had decreased levels of TNF, KC (IL-8), and IL-6. Conclusions This work illustrates that B. dentium enhances the intestinal mucus layer and goblet cell function via upregulation of gene expression and autophagy signaling pathways with a net increase in mucin production. Ultimately, these pathways may be targeted for the development of novel therapeutics.

Specific targeting of intestinalPrevotella copriby aListeria monocytogenesbacteriocin

Deciphering the specific function of every microorganism in microbial gut communities is a key issue to interrogate their role during infection. Here, we report the discovery of aListeriabacteriocin, Lmo2776, that specifically targets the abundant gut commensalPrevotella copriand affectsListeriainfection. Oral infection of conventional mice with a Δlmo2776mutant leads to a thinner intestinal mucus layer and higherListerialoads both in the intestinal content and deeper tissues compared to WTListeria, while no difference is observed in germ-free mice. This microbiota-dependent effect is phenocopied by precolonization of germ-free mice beforeListeriainfection, withP. copri, but not with other commensals,. Together, these data unveil a role forPrevotellain controlling intestinal infection, highlighting that pathogens may selectively deplete microbiota to avoid excessive inflammation.

Bifidobacterium dentiumFortifies the Intestinal Mucus Layer via Autophagy and Calcium Signaling Pathways

ABSTRACTMuch remains unknown about how the intestinal microbiome interfaces with the protective intestinal mucus layer.Bifidobacteriumspecies colonize the intestinal mucus layer and can modulate mucus production by goblet cells. However, selectBifidobacteriumstrains can also degrade protective glycans on mucin proteins. We hypothesized that the human-derived speciesBifidobacterium dentiumwould increase intestinal mucus synthesis and expulsion, without extensive degradation of mucin glycans.In silicodata revealed thatB. dentiumlacked the enzymes necessary to extensively degrade mucin glycans. This finding was confirmed by demonstrating thatB. dentiumcould not use naive mucin glycans as primary carbon sourcesin vitro. To examineB. dentiummucus modulationin vivo, Swiss Webster germfree mice were monoassociated with live or heat-killedB. dentium. LiveB. dentium-monoassociated mice exhibited increased colonic expression of goblet cell markersKrüppel-like factor 4(Klf4),Trefoil factor 3(Tff3),Relm-β,Muc2, and several glycosyltransferases compared to both heat-killedB. dentiumand germfree counterparts. Likewise, liveB. dentium-monoassociated colon had increased acidic mucin-filled goblet cells, as denoted by Periodic Acid-Schiff-Alcian Blue (PAS-AB) staining and MUC2 immunostaining.In vitro,B. dentium-secreted products, including acetate, were able to increase MUC2 levels in T84 cells. We also identified thatB. dentium-secreted products, such as γ-aminobutyric acid (GABA), stimulated autophagy-mediated calcium signaling and MUC2 release. This work illustrates thatB. dentiumis capable of enhancing the intestinal mucus layer and goblet cell function via upregulation of gene expression and autophagy signaling pathways, with a net increase in mucin production.IMPORTANCEMicrobe-host interactions in the intestine occur along the mucus-covered epithelium. In the gastrointestinal tract, mucus is composed of glycan-covered proteins, or mucins, which are secreted by goblet cells to form a protective gel-like structure above the epithelium. Low levels of mucin or alterations in mucin glycans are associated with inflammation and colitis in mice and humans. Although current literature links microbes to the modulation of goblet cells and mucins, the molecular pathways involved are not yet fully understood. Using a combination of gnotobiotic mice and mucus-secreting cell lines, we have identified a human-derived microbe,Bifidobacterium dentium, which adheres to intestinal mucus and secretes metabolites that upregulate the major mucin MUC2 and modulate goblet cell function. Unlike otherBifidobacteriumspecies,B. dentiumdoes not extensively degrade mucin glycans and cannot grow on mucin alone. This work points to the potential of usingB. dentiumand similar mucin-friendly microbes as therapeutic agents for intestinal disorders with disruptions in the mucus barrier.