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Critical design aspects involved in the study of Paneth cells and the intestinal microbiota.

ABSTRACT: Paneth cells are long-lived secretory cells that reside in the base of the crypts of Lieberkühn of the small intestine. They produce an arsenal of molecules that are involved in numerous biological processes, ranging from the control of gut microbial populations to supporting the intestinal stem cell niche. Because of these important functions, Paneth cell abnormalities are becoming implicated in a variety of disease processes. As such, it is necessary to establish parameters that will allow for the comprehensive study of Paneth cells in health and disease. In this addendum, we highlight critical design aspects involved in the study of Paneth cells and their downstream effects on the intestinal microbiota. The importance of this approach is demonstrated by our recent findings that Nod2 does not regulate mouse Paneth cell antimicrobial function, in contrast to previous reports. This work defines key issues to consider when studying Paneth cells in mouse systems.

SUBMITTER: Shanahan MT

PROVIDER: S-EPMC4063846 | biostudies-literature | 2014 Mar-Apr

REPOSITORIES: biostudies-literature

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Critical design aspects involved in the study of Paneth cells and the intestinal microbiota.

Shanahan Michael T MT Carroll Ian M IM Gulati Ajay S AS

Gut microbes 20131220 2

Paneth cells are long-lived secretory cells that reside in the base of the crypts of Lieberkühn of the small intestine. They produce an arsenal of molecules that are involved in numerous biological processes, ranging from the control of gut microbial populations to supporting the intestinal stem cell niche. Because of these important functions, Paneth cell abnormalities are becoming implicated in a variety of disease processes. As such, it is necessary to establish parameters that will allow for ...[more]

PMID: 24637592

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Project description:The recognition of autophagy related 16-like 1 (ATG16L1) as a genetic risk factor has exposed the critical role of autophagy in Crohn's disease. Homozygosity for the highly prevalent ATG16L1 risk allele, or murine hypomorphic (HM) activity, causes Paneth cell dysfunction. As Atg16l1(HM) mice do not develop spontaneous intestinal inflammation, the mechanism(s) by which ATG16L1 contributes to disease remains obscure. Deletion of the unfolded protein response (UPR) transcription factor X-box binding protein-1 (Xbp1) in intestinal epithelial cells, the human orthologue of which harbours rare inflammatory bowel disease risk variants, results in endoplasmic reticulum (ER) stress, Paneth cell impairment and spontaneous enteritis. Unresolved ER stress is a common feature of inflammatory bowel disease epithelium, and several genetic risk factors of Crohn's disease affect Paneth cells. Here we show that impairment in either UPR (Xbp1(?IEC)) or autophagy function (Atg16l1(?IEC) or Atg7(?IEC)) in intestinal epithelial cells results in each other's compensatory engagement, and severe spontaneous Crohn's-disease-like transmural ileitis if both mechanisms are compromised. Xbp1(?IEC) mice show autophagosome formation in hypomorphic Paneth cells, which is linked to ER stress via protein kinase RNA-like endoplasmic reticulum kinase (PERK), elongation initiation factor 2? (eIF2?) and activating transcription factor 4 (ATF4). Ileitis is dependent on commensal microbiota and derives from increased intestinal epithelial cell death, inositol requiring enzyme 1? (IRE1?)-regulated NF-?B activation and tumour-necrosis factor signalling, which are synergistically increased when autophagy is deficient. ATG16L1 restrains IRE1? activity, and augmentation of autophagy in intestinal epithelial cells ameliorates ER stress-induced intestinal inflammation and eases NF-?B overactivation and intestinal epithelial cell death. ER stress, autophagy induction and spontaneous ileitis emerge from Paneth-cell-specific deletion of Xbp1. Genetically and environmentally controlled UPR function within Paneth cells may therefore set the threshold for the development of intestinal inflammation upon hypomorphic ATG16L1 function and implicate ileal Crohn's disease as a specific disorder of Paneth cells.

Project description:Hypertriglyceridemia (HTG) aggravates the course of acute pancreatitis (AP). Intestinal barrier dysfunction is implicated in the pathogenesis of AP during which dysbiosis of intestinal microbiota contributes to the dysfunction in intestinal barrier. However, few studies focus on the changes in intestine during HTG-related acute necrotizing pancreatitis (ANP). Here, we investigated the changes in intestinal microbiota and Paneth cell antimicrobial peptides (AMPs) in HTG-related ANP (HANP) in rats. Rats fed a high-fat diet to induce HTG and ANP was induced by retrograde injection of 3.5% sodium taurocholate into biliopancreatic duct. Rats were sacrificed at 24 and 48 h, respectively. Pancreatic and ileal injuries were evaluated by histological scores. Intestinal barrier function was assessed by plasma diamine oxidase activity and D-lactate level. Systemic and intestinal inflammation was evaluated by tumor necrosis factor alpha (TNFα), interleukin (IL)-1β, and IL-17A expression. 16S rRNA high throughput sequencing was used to investigate changes in intestinal microbiota diversity and structure. AMPs (α-defensin5 and lysozyme) expression was measured by real-time polymerase chain reaction (PCR) and immunofluorescence. The results showed that compared with those of normal-lipid ANP (NANP) groups, the HANP groups had more severe histopathological injuries in pancreas and distal ileum, aggravated intestinal barrier dysfunction and increased TNFα, IL-1β, and IL-17A expression in plasma and distal ileum. Principal component analysis showed structural segregation between the HANP and NANP group. α-Diversity estimators in the HANP group revealed decreased microbiota diversity compared with that in NANP group. Taxonomic analysis showed dysbiosis of intestinal microbiota structure. In the HANP group, at phyla level, Candidatus_Saccharibacteria and Tenericutes decreased significantly, whereas Actinobacteria increased. At genus level, Allobaculum, Bifidobacterium, and Parasutterella increased significantly, while Alloprevotella, Anaerotruncus, Candidatus_Saccharimonas, Christensenellaceae_R-7_group, Rikenellaceae_RC9_gut_group, Ruminiclostridium_5, Ruminococcaceae_UCG-005, and Ruminococcaceae_UCG-014 decreased. Compared with those in the NANP rats, mRNA expression of lysozyme and α-defensin5 and protein expression of lysozyme decreased significantly in the HANP rats. Moreover, in the NANP rats and the HANP rats, Allobaculum abundance was inversely correlated with lysozyme expression, while Anaerotruncus abundance was positively correlated with it by Spearman test. In conclusion, intestinal microbiota dysbiosis and decreased AMPs of Paneth cells might participate in the pathogenesis of intestinal barrier dysfunction in HANP.

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Critical design aspects involved in the study of Paneth cells and the intestinal microbiota.

Publications

Critical design aspects involved in the study of Paneth cells and the intestinal microbiota.

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