Background & Aims Intestinal epithelial homeostasis is maintained by complex interactions among epithelial cells, commensal gut microorganisms, and immune cells. Disruption of this homeostasis is associated with disorders such as inflammatory bowel disease, but the mechanisms of this process are not clear. We investigated how Sirtuin 1 (SIRT1), a conserved mammalian NAD+-dependent protein deacetylase, senses environmental stress to alter intestinal integrity. Methods We performed studies of mice with disruption of Sirt1 specifically in the intestinal epithelium (SIRT1 iKO, villin-Cre+, Sirt1flox/flox mice) and control mice (villinCre-, Sirt1flox/flox) on a C57BL/6 background. Acute colitis was induced in some mice by addition of 2.5% dextran sodium sulfate to drinking water for 5–9 consecutive days. Some mice were given antibiotics via their drinking water for 4 weeks to deplete their microbiota. Some mice were fed with a cholestyramine containing diet for 7 days to sequester their bile acids. Feces were collected and proportions of microbiota were analyzed by 16S rRNA amplicon sequencing and quantitative PCR. Intestines were collected from mice and gene expression profiles were compared by microarray and quantitative PCR analyses. We compared levels of specific mRNAs between colon tissues from age-matched patients with ulcerative colitis (n=10) vs without inflammatory bowel disease (n=8, controls). Results Mice with intestinal deletion of SIRT1 (SIRT1 iKO) had abnormal activation of Paneth cells starting at the age of 5–8 months, with increased activation of NF-κB, stress pathways, and spontaneous inflammation at 22–24 months of age, compared with control mice. SIRT1 iKO mice also had altered fecal microbiota starting at 4–6 months of age compared with control mice, in part due to altered bile acid metabolism. Moreover, SIRT1 iKO mice with defective gut microbiota developed more severe colitis than control mice. Intestinal tissues from patients with ulcerative colitis expressed significantly lower levels of SIRT1 mRNA than controls. Intestinal tissues from SIRT1 iKO mice given antibiotics, however, did not have signs of inflammation at 22–24 months of age, and did not develop more severe colitis than control mice at 4–6 months. Conclusions In analyses of intestinal tissues, colitis induction, and gut microbiota in mice with intestinal disruption of SIRT1, we found this protein to prevent intestinal inflammation by regulating the gut microbiota. SIRT1 might therefore be an important mediator of host–microbiome interactions. Agents designed to activate SIRT1 might be developed as treatments for inflammatory bowel diseases.

Objective Although polymorphisms of the NOD2 gene predispose to the development of ileal Crohn's disease, the precise mechanisms of this increased susceptibility remain unclear. Previous work has shown that transcript expression of the Paneth cell (PC) antimicrobial peptides (AMPs) α-defensin 4 and α-defensin-related sequence 10 are selectively decreased in Nod2−/− mice. However, the specific mouse background used in this previous study is unclear. In light of recent evidence suggesting that mouse strain strongly influences PC antimicrobial activity, we sought to characterise PC AMP function in commercially available Nod2−/− mice on a C57BL/6 (B6) background. Specifically, we hypothesised that Nod2−/− B6 mice would display reduced AMP expression and activity. Design Wild-type (WT) and Nod2−/− B6 ileal AMP expression was assessed via real-time PCR, acid urea polyacrylamide gel electrophoresis and mass spectrometry. PCs were enumerated using flow cytometry. Functionally, α-defensin bactericidal activity was evaluated using a gel-overlay antimicrobial assay. Faecal microbial composition was determined using 454-sequencing of the bacterial 16S gene in cohoused WT and Nod2−/− littermates. Results WT and Nod2−/− B6 mice displayed similar PC AMP expression patterns, equivalent α-defensin profiles, and identical antimicrobial activity against commensal and pathogenic bacterial strains. Furthermore, minimal differences in gut microbial composition were detected between the two cohoused, littermate mouse groups. Conclusions Our data reveal that Nod2 does not directly regulate PC antimicrobial activity in B6 mice. Moreover, we demonstrate that previously reported Nod2-dependent influences on gut microbial composition may be overcome by environmental factors, such as cohousing with WT littermates.

Background Increasing evidence supports the central role of Paneth cells in maintaining intestinal host-microbial homeostasis. However, the direct impact of host genotype on Paneth cell function remains unclear. Here, we characterize key differences in Paneth cell function and intestinal microbial composition in two widely utilized, genetically distinct mouse strains (C57BL/6 and 129/SvEv). In doing so, we demonstrate critical influences of host genotype on Paneth cell activity and the enteric microbiota. Methodology and Principal Findings Paneth cell numbers were determined by flow cytometry. Antimicrobial peptide (AMP) expression was evaluated using quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR), acid urea-polyacrylamide gel electrophoresis, and mass spectrometry. Effects of mouse background on microbial composition were assessed by reciprocal colonization of germ-free mice from both background strains, followed by compositional analysis of resultant gut bacterial communities using terminal restriction fragment length polymorphism analysis and 16 S qPCR. Our results revealed that 129/SvEv mice possessed fewer Paneth cells and a divergent AMP profile relative to C57BL/6 counterparts. Novel 129/SvEv á-defensin peptides were identified, including Defa2/18v, Defa11, Defa16, and Defa18. Host genotype profoundly affected the global profile of the intestinal microbiota, while both source and host factors were found to influence specific bacterial groups. Interestingly, ileal α-defensins from 129/SvEv mice displayed attenuated antimicrobial activity against pro-inflammatory E. coli strains, a bacterial species found to be expanded in these animals. Conclusions and Significance This work establishes the important impact of host genotype on Paneth cell function and the composition of the intestinal microbiota. It further identifies specific AMP and microbial alterations in two commonly used inbred mouse strains that have varying susceptibilities to a variety of disorders, ranging from obesity to intestinal inflammation. This will be critical for future studies utilizing these murine backgrounds to study the effects of Paneth cells and the intestinal microbiota on host health and disease.

Paneth cells at the base of small intestinal crypts secrete microbicidal ␣-defensins, termed cryptdins (Crps) in mice, as mediators of innate immunity. Proteomic studies show that five abundant Paneth cell ␣-defensins in C57BL/6 mice are strain specific in that they have not been identified in other inbred strains of mice. Two C57BL/6-specific peptides are coded for by the Defcr20 and -21 genes evident in the NIH C57BL/6 genome but absent from the Celera mixed-strain assembly, which excludes C57BL/6 data and differs from the NIH build with respect to the organization of the ␣-defensin gene locus. Conversely, C57BL/6 mice lack the Crp1, -2, -4, and -6 peptides and their corresponding Defcr1, -2, -4, and -6 genes, which are common to several mouse strains, including those of the Celera assembly. In C57BL/6 mice, ␣-defensin gene diversification appears to have occurred by tandem duplication of a multigene cassette that was not found in the mixed-strain assembly. Both mouse genome assemblies contain conserved ␣-defensin pseudogenes that are closely related to functional myeloid ␣-defensin genes in the rat, suggesting that the neutrophil ␣-defensin defect in mice resulted from progressive gene loss. Given the role of ␣-defensins in shaping the composition of the enteric microflora, such polymorphisms may influence outcomes in mouse models of disease or infection.

Light and moleculesThe Perrin-Jablonski diagram (figure 1) is convenient for visualizing the different processes involved in the