Loss of NHE3 alters gut microbiota composition and influencesBacteroides thetaiotaomicrongrowth

Abstract

Changes in the intestinal microbiota have been linked to diabetes, obesity, inflammatory bowel disease, and Clostridium difficile ( C. difficile)-associated disease. Despite this, it remains unclear how the intestinal environment, set by ion transport, affects luminal and mucosa-associated bacterial composition. Na+/H+-exchanger isoform 3 (NHE3), a target of C. difficile toxin B, plays an integral role in intestinal Na+ absorption. Thus the NHE3-deficient mouse model was chosen to examine the effect of pH and ion composition on bacterial growth. We hypothesized that ion transport-induced change in the intestinal environment would lead to alteration of the microbiota. Region-specific changes in ion composition and pH correlated with region-specific alteration of luminal and mucosal-associated bacteria with general decreases in Firmicutes and increases in Bacteroidetes members. Bacteroides thetaiotaomicron ( B. thetaiotaomicron) increased in NHE3−/− terminal ileum and was examined in vitro to determine whether altered Na+ was sufficient to affect growth. Increased in vitro growth of B. thetaiotaomicron occurred in 43 mM Na+ correlating with the NHE3−/− mouse terminal ileum [Na+]. NHE3−/− terminal ileum displayed increased fut2 mRNA and fucosylation correlating with B. thetaiotaomicron growth. Inoculation of B. thetaiotaomicron in wild-type and NHE3−/− terminal ileum organoids displayed increased fut2 and fucosylation, indicating that B. thetaiotaomicron alone is sufficient for the increased fucosylation seen in vivo. These data demonstrate that loss of NHE3 alters the intestinal environment, leading to region-specific changes in bacteria, and shed light on the growth requirements of some gut microbiota members, which is vital for creating better treatments of complex diseases with an altered gut microbiota.

Role of epithelial ion transports in inflammatory bowel disease

“…Surprisingly, the major inflammation trigger in NHE3 Ϫ/Ϫ mice was the increased bacterial adhesion and translocation, since antibiotic treatment reversed many colitis manifestations (83). Later studies on the colonic microbiota of NHE3 Ϫ/Ϫ mice found increased Bacteroidetes but decreased Firmicutes bacteria (40) and overall lower bacterial diversity (81), which goes along with reports of reduced and altered microbiota in patients with IBD (107). Curiously, NHE2 Ϫ/Ϫ mice (which are not a model of IBD) featured radically different microbiota changes (41).…”

Organoids as Model for Infectious Diseases: Culture of Human and Murine Stomach Organoids and Microinjection of Helicobacter Pylori

“…It is practical to use a 3-dimensional manipulator instead of a 2 dimensional manipulator to allow maximal flexibility to reach into the wells. Oil-based microinjectors (such as Celltram, or IM-5B) provide a finer and slower manual control compared to air-based microinjectors (such as FemtoJet 11 , or Nanojet microinjector 10 ). In turn, the latter have the important advantage that a precise injection volume can be programmed.…”

“…They have been used to model infections with viruses, such as Cytomegalovirus 1,2 or Hepatitis C Virus [3][4][5][6][7] , parasites, such as Plasmodium falciparum 8 or Toxoplasma gondii 9 , and bacteria, such as Bacterioides thetaiotaomicron 10 or Salmonella enterica 11 . Most recently, several approaches have been published to model infection with H. pylori with organoids derived from ESC or iPS cells 12 , mouse adult stem cells 21,22 or human adult stem cells [13][14][15] .…”

Modeling infectious diseases and host-microbe interactions in gastrointestinal organoids

“…In addition, many members of the gastrointestinal microbiota are obligate anaerobic bacteria and their anaerobic lifestyle hinders in vitro experimentation with 2D cell lines grown in standard 20% O2. Two anaerobic bacteria, Bacteroides thetaiotaomicron (B. thetaiotaomicron), a member of the healthy microbiota, and the anaerobic pathogen Clostridium difficile (C. difficile), have each been injected into organoids (Engevik et al, 2013;Engevik et al, 2015;Leslie et al, 2015). While the B. thetaiotaomicron study did not address the question of bacterial survival, authors of the C. difficile study re-cultured bacteria from organoids after 12 hours of infection, indicating that anaerobes can survive for 12 hours in organoid culture (Leslie et al, 2015).…”

“…Toxins produced by the bacterium then cause mild to severe gastroenteritis, but can also lead to severe complicated pseudomembranous colitis, characterized by inflammatory lesions and pseudomembranes in the colon (reviewed in Rupnik et al, 2009). During C. difficile infection of organoids, bacterial toxins could also be shown to inhibit barrier function, as assessed by the retention of FITC-dextran (Leslie et al, 2015), and diminish expression of Na+/H+ exchanger 3 (NHE3), which may influence gut microbiota composition (Engevik et al, 2013;Engevik et al, 2015). Salmonella enteritica serovar Typhimurium (S. Typhimurium) causes self-limiting gastroenteritis in humans and systemic disease in susceptible mice.…”