Gastroenterology
Abstract
Crohn's disease (CD) is the chronic inflammation of the terminal ileum and colon triggered by a dysregulated immune response to bacteria, but insights into specific molecular perturbations at the critical bacteria-epithelium interface are limited. Here we report that the membrane mucin MUC17 protected small intestinal enterocytes against commensal and pathogenic bacteria. In non-inflamed CD ileum, reduced MUC17 levels and a compromised glycocalyx barrier allowed recurrent bacterial contact with enterocytes. Muc17 deletion in mice rendered the small intestine particularly prone to atypical bacterial infection while maintaining resistance to colitis. The loss of Muc17 resulted in spontaneous deterioration of epithelial homeostasis and in the extra-intestinal translocation of bacteria. Finally, Muc17-deficient mice harbored specific small intestinal bacterial taxa observed in CD patients. Our findings highlight MUC17 as an essential regiospecific line of defense in the small intestine with relevance for early epithelial defects in CD.
Authors
Elena Layunta, Sofia Jäverfelt, Fleur C. van de Koolwijk, Molly Sivertsson, Brendan Dolan, Liisa Arike, Sara I.M. Thulin, Bruce A. Vallance, Thaher Pelaseyed
Abstract
Authors
Huafeng Fu, Qinbo Cai, Zhijun Zhou, Yulong He, Min Li, DongJie Yang
Abstract
The pathogenesis of the murine model of autoimmune pancreatitis associated with IgG4-related disease (AIP/IgG4-RD) induced by administration of polyinosinic-polycytidylic acid, is incompletely understood. While it is known that murine and human AIP/IgG4-RD is driven by plasmacytoid dendritic cells (pDCs) producing IFN-α, the origin of these cells and their relation to effector T cells is not known. Here we show that murine AIP was initiated by TLR3-bearing conventional DCs in the uninflamed pancreas whose activation by TLR3 ligand (polyinosinic-polycytidylic acid) caused IFN-α, CXCL9, and CXCL10 secretion. This, in turn, induced pancreatic recruitment of CXCR3+ T cells and these T cells, via their secretion of CCL25, facilitated migration of pDCs bearing CCR9 into the pancreas. This established a feedback loop anchored by the now dominant pDC production of IFN-α and the continued CXCR3+ T cell facilitation of pDC migration. Remarkably, the interaction between CXCR3+ T cells and pDCs also existed at the functional levels since this interaction enhanced the production of CCL25 and IFN-α by CXCR3+ T cells and pDCs, respectively. Evidence presented here that a similar disease mechanism was present in human AIP/IgG4-RD creates new avenues of disease treatment.
Authors
Akane Hara, Tomohiro Watanabe, Kosuke Minaga, Tomoe Yoshikawa, Masayuki Kurimoto, Ikue Sekai, Yasuhiro Masuta, Ryutaro Takada, Yasuo Otsuka, Ken Kamata, Shiki Takamura, Masatoshi Kudo, Warren Strober
Abstract
Thrombospondin-1 (TSP1) is a matricellular protein associated with the regulation of cell migration through direct binding interactions with integrin proteins and by associating with other receptors known to regulate integrin function, including CD47 and CD36. We previously demonstrated that deletion of an epithelial TSP1 receptor CD47 attenuates epithelial wound repair following intestinal mucosal injury. However, the mechanisms by which TSP1 contributes to intestinal mucosal repair remains poorly understood. Our results show upregulated TSP1 expression in colonic mucosal wounds and impaired intestinal mucosal wound healing in vivo upon intestinal epithelial specific loss of TSP1 (VillinCre/+Thbs1f/f or Thbs1ΔIEC). We report that exposure to exogenous TSP1 enhanced migration of IECs in a CD47– and TGFβ1-dependent manner, and that deficiency of TSP1 in primary murine colonic epithelial cells resulted in impaired wound healing. Mechanistically, TSP1 modulated epithelial actin cytoskeletal dynamics by suppression of RhoA activity, activation of Rac1, and changes in F-actin bundling. Overall, TSP1 was found to regulate intestinal mucosal wound healing via CD47 and TGFβ1, coordinate integrin-containing cell-matrix adhesion dynamics and remodel the actin cytoskeleton in migrating epithelial cells to enhance cell motility and promote wound repair.
Authors
Zachary S. Wilson, Arturo Raya-Sandino, Jael Miranda, Shuling Fan, Jennifer C. Brazil, Miguel Quiros, Vicky Garcia-Hernandez, Qingyang Liu, Chang H. Kim, Kurt D. Hankenson, Asma Nusrat, Charles A. Parkos
Abstract
The goal of this study was to determine if transplantation of enteric neural stem cells (ENSCs) can rescue the enteric nervous system (ENS), restore gut motility, reduce colonic inflammation, and improve survival in the Ednrb knock-out (KO) mouse model of Hirschsprung disease (HSCR). ENSCs were isolated from mouse intestine, expanded to form neurospheres, and microinjected into the colon of recipient Ednrb KO mice. Transplanted ENSCs were identified in recipient colons as cell clusters in “neo-ganglia”. Immunohistochemical evaluation demonstrated extensive cell migration away from the sites of cell delivery and across the muscle layers. Electrical field stimulation and optogenetics showed significantly enhanced contractile activity of aganglionic colonic smooth muscle following ENSC transplantation and confirmed functional neuromuscular integration of the transplanted ENSC-derived neurons. ENSC injection also partially restored the colonic migrating motor complex. Histological examination revealed a significant reduction in inflammation in ENSC-transplanted aganglionic recipient colon compared to sham-operated mice. Interestingly, mice that received cell transplant also had prolonged survival compared with controls. This study demonstrates that ENSC transplantation can improve outcomes in HSCR by restoring gut motility and reducing the severity of Hirschsprung-associated enterocolitis, the leading cause of death in human HSCR.
Authors
Ahmed A. Rahman, Takahiro Ohkura, Sukhada Bhave, Weikang Pan, Kensuke Ohishi, Leah Ott, Christopher Han, Abigail Leavitt, Rhian Stavely, Alan J. Burns, Allan M. Goldstein, Ryo Hotta
Abstract
Duodenal bicarbonate secretion is critical to epithelial protection, nutrient digestion/absorption and is impaired in cystic fibrosis (CF). We examined if linaclotide, typically used to treat constipation, may also stimulate duodenal bicarbonate secretion. Bicarbonate secretion was measured in vivo and in vitro using mouse and human duodenum (biopsies and enteroids). Ion transporter localization was identified with confocal microscopy and de novo analysis of human duodenal single cell RNA sequencing (sc-RNAseq) datasets was performed. Linaclotide increased bicarbonate secretion in mouse and human duodenum in the absence of CFTR expression (Cftr knockout mice) or function (CFTRinh-172). NHE3 inhibition contributed to a portion of this response. Linaclotide-stimulated bicarbonate secretion was eliminated by down-regulated in adenoma (DRA, SLC26A3) inhibition during loss of CFTR activity. Sc-RNAseq identified that 70% of villus cells expressed SLC26A3, but not CFTR, mRNA. Loss of CFTR activity and linaclotide increased apical brush border expression of DRA in non-CF and CF differentiated enteroids. These data provide further insights into the action of linaclotide and how DRA may compensate for loss of CFTR in regulating luminal pH. Linaclotide may be a useful therapy for CF individuals with impaired bicarbonate secretion.
Authors
Jessica B. Sarthi, Annie M. Trumbull, Shayda M. Abazari, Vincent van Unen, Joshua E. Chan, Yanfen Jiang, Jesse Gammons, Marc O. Anderson, Onur Cil, Calvin J. Kuo, Zachary M. Sellers
Abstract
Immunosuppression is a common feature of esophageal adenocarcinoma (EAC) and has been linked to poor overall survival (OS). We hypothesized that upstream factors might negatively influence CD3 levels and T-cell activity, thus promoting immunosuppression and worse survival. We used clinical data and patient samples of those who progressed from Barrett’s (BE) to dysplasia to EAC, investigated gene (RNAseq), protein (tissue microarray) expression and performed cell biology studies to delineate a pathway impacting CD3 protein stability that might influence EAC outcome. We show that the loss of both CD3-ε expression and CD3+ T-cell number are correlated with worse OS in EAC. The GRAIL (gene related to anergy in lymphocytes) isoform 1 (GRAIL1), which is the prominent isoform in EACs, degrades (ε, γ, δ) CD3s and inactivates T-cells. In contrast, isoform 2 (GRAIL2), which is reduced in EACs, stabilizes CD3s. Further, GRAIL1 mediated CD3 degradation is facilitated by interferon stimulated gene 15 (ISG15), a ubiquitin-like protein. Consequently, either the overexpression of a ligase-dead GRAIL1, ISG15 knockdown, or the overexpression of a conjugation-defective ISG15-LRAA mutant can increase CD3 levels. Together, we identified that an ISG15→GRAIL1→mutant p53 amplification loop negatively influencing CD3 levels and T-cell activity, thus promoting immunosuppression in EAC.
Authors
Dyke P. McEwen, Paramita Ray, Derek J. Nancarrow, Zhuwen Wang, Srimathi Kasturirangan, Saeed Abdullah, Ayushi Balan, Rishi Hoskeri, Dafydd Thomas, Theodore S. Lawrence, David G. Beer, Kiran H. Lagisetty, Dipankar Ray
Abstract
Fibroblast growth factor 15/19 (FGF15/19, mouse/human ortholog) is expressed in the ileal enterocytes of the small intestine and released postprandially in response to bile acid absorption. Previous reports of FGF15–/– mice have limited our understanding of gut-specific FGF15’s role in metabolism. Therefore, we studied the role of endogenous gut-derived FGF15 in bile acid, cholesterol, glucose, and energy balance. We found that circulating levels of FGF19 were reduced in individuals with obesity and comorbidities, such as type 2 diabetes and metabolic dysfunction–associated fatty liver disease. Gene expression analysis of ileal FGF15-positive cells revealed differential expression during the obesogenic state. We fed standard chow or a high-fat metabolic dysfunction-associated steatohepatitis–inducing diet to control and intestine-derived FGF15-knockout (FGF15INT-KO) mice. Control and FGF15INT-KO mice gained similar body weight and adiposity and did not show genotype-specific differences in glucose, mixed meal, pyruvate, and glycerol tolerance. FGF15INT-KO mice had increased systemic bile acid levels but decreased cholesterol levels, pointing to a primary role for gut-derived FGF15 in regulating bile acid and cholesterol metabolism when exposed to obesogenic diet. These studies show that intestinal FGF15 plays a specific role in bile acid and cholesterol metabolism regulation but is not essential for energy and glucose balance.
Authors
Nadejda Bozadjieva-Kramer, Jae Hoon Shin, Ziru Li, Alan C. Rupp, Nicole Miller, Stace Kernodle, Nicolas Lanthier, Paulina Henry, Nikhil Seshadri, Andriy Myronovych, Ormond A. MacDougald, Robert W. O’Rourke, Rohit Kohli, Charles F. Burant, Amy E. Rothberg, Randy J. Seeley
Abstract
Crohn’s disease (CD) is a chronic inflammatory gut disorder. Molecular mechanisms underlying the clinical heterogeneity of CD remain poorly understood. MicroRNAs (miRNAs) are important regulators of gut physiology, and several have been implicated in the pathogenesis of adult CD. However, there is a dearth of large-scale miRNA studies for pediatric CD. We hypothesized that specific miRNAs uniquely mark pediatric CD. We performed small RNA-Seq of patient-matched colon and ileum biopsies from treatment-naive pediatric patients with CD (n = 169) and a control cohort (n = 108). Comprehensive miRNA analysis revealed 58 miRNAs altered in pediatric CD. Notably, multinomial logistic regression analysis revealed that index levels of ileal miR-29 are strongly predictive of severe inflammation and stricturing. Transcriptomic analyses of transgenic mice overexpressing miR-29 show a significant reduction of the tight junction protein gene Pmp22 and classic Paneth cell markers. The dramatic loss of Paneth cells was confirmed by histologic assays. Moreover, we found that pediatric patients with CD with elevated miR-29 exhibit significantly lower Paneth cell counts, increased inflammation scores, and reduced levels of PMP22. These findings strongly indicate that miR-29 upregulation is a distinguishing feature of pediatric CD, highly predictive of severe phenotypes, and associated with inflammation and Paneth cell loss.
Authors
Alexandria J. Shumway, Michael T. Shanahan, Emilie Hollville, Kevin Chen, Caroline Beasley, Jonathan W. Villanueva, Sara Albert, Grace Lian, Moises R. Cure, Matthew Schaner, Lee-Ching Zhu, Surekha Bantumilli, Mohanish Deshmukh, Terrence S. Furey, Shehzad Z. Sheikh, Praveen Sethupathy
Abstract
The role of long noncoding RNAs (lncRNAs) in disease is incompletely understood, but their regulation of inflammation is increasingly appreciated. We addressed the extent of lncRNA involvement in inflammatory bowel disease (IBD) using biopsy-derived RNA-sequencing data from a large cohort of deeply phenotyped patients with IBD. Weighted gene correlation network analysis revealed gene modules of lncRNAs coexpressed with protein-coding genes enriched for biological pathways, correlated with epithelial and immune cell signatures, or correlated with distal colon expression. Correlation of modules with clinical features uncovered a module correlated with disease severity, with an enriched interferon response signature containing the hub lncRNA IRF1-AS1. Connecting genes to IBD-associated single nucleotide polymorphisms (SNPs) revealed an enrichment of SNP-adjacent lncRNAs in biologically relevant modules. Ulcerative colitis–specific SNPs were enriched in distal colon–related modules, suggesting that disease-specific mechanisms may result from altered lncRNA expression. The function of the IBD-associated SNP-adjacent lncRNA IRF1-AS1 was explored in human myeloid cells, and our results suggested IRF1-AS1 promoted optimal production of TNF-α, IL-6, and IL-23. A CRISPR/Cas9-mediated activation screen in THP-1 cells revealed several lncRNAs that modulated LPS-induced TNF-α responses. Overall, this study uncovered the expression patterns of lncRNAs in IBD that identify functional, disease-relevant lncRNAs.
Authors
John L. Johnson, Davit Sargsyan, Eric M. Neiman, Amy Hart, Aleksandar Stojmirovic, Roman Kosoy, Haritz Irizar, Mayte Suárez-Fariñas, Won-Min Song, Carmen Argmann, Stefan Avey, Liraz Shmuel-Galia, Tim Vierbuchen, Gerold Bongers, Yu Sun, Leonard Edelstein, Jacqueline Perrigoue, Jennifer E. Towne, Aisling O’Hara Hall, Katherine A. Fitzgerald, Kasper Hoebe
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