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Decreased Colonic Activin Receptor-Like Kinase 1 Disrupts Epithelial Barrier Integrity in Patients With Crohn's Disease.

ABSTRACT: BACKGROUND & AIMS:Intestinal epithelial cell (IEC) barrier dysfunction is critical to the development of Crohn's disease (CD). However, the mechanism is understudied. We recently reported increased microRNA-31-5p (miR-31-5p) expression in colonic IECs of CD patients, but downstream targets and functional consequences are unknown. METHODS:microRNA-31-5p target genes were identified by integrative analysis of RNA- and small RNA-sequencing data from colonic mucosa and confirmed by quantitative polymerase chain reaction in colonic IECs. Functional characterization of activin receptor-like kinase 1 (ACVRL1 or ALK1) in IECs was performed ex vivo using 2-dimensional cultured human primary colonic IECs. The impact of altered colonic ALK1 signaling in CD for the risk of surgery and endoscopic relapse was evaluated by a multivariate regression analysis and a Kaplan-Meier estimator. RESULTS:ALK1 was identified as a target of miR-31-5p in colonic IECs of CD patients and confirmed using a 3'-untranslated region reporter assay. Activation of ALK1 restricted the proliferation of colonic IECs in a 5-ethynyl-2-deoxyuridine proliferation assay and down-regulated the expression of stemness-related genes. Activated ALK1 signaling increased colonic IEC differentiation toward colonocytes. Down-regulated ALK1 signaling was associated with increased stemness and decreased colonocyte-specific marker expression in colonic IECs of CD patients compared with healthy controls. Activation of ALK1 enhanced epithelial barrier integrity in a transepithelial electrical resistance permeability assay. Lower colonic ALK1 expression was identified as an independent risk factor for surgery and was associated with a higher risk of endoscopic relapse in CD patients. CONCLUSIONS:Decreased colonic ALK1 disrupted colonic IEC barrier integrity and was associated with poor clinical outcomes in CD patients.

SUBMITTER: Toyonaga T

PROVIDER: S-EPMC7502566 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Decreased Colonic Activin Receptor-Like Kinase 1 Disrupts Epithelial Barrier Integrity in Patients With Crohn's Disease.

Toyonaga Takahiko T Steinbach Erin C EC Keith Benjamin P BP Barrow Jasmine B JB Schaner Matthew R MR Wolber Elisabeth A EA Beasley Caroline C Huling Jennifer J Wang Yuli Y Allbritton Nancy L NL Chaumont Nicole N Sadiq Timothy S TS Koruda Mark J MJ Jain Animesh A Long Millie D MD Barnes Edward L EL Herfarth Hans H HH Isaacs Kim L KL Hansen Jonathan J JJ Shanahan Michael T MT Rahbar Reza R Furey Terrence S TS Sethupathy Praveen P Sheikh Shehzad Z SZ

Cellular and molecular gastroenterology and hepatology 20200616 4

<h4>Background & aims</h4>Intestinal epithelial cell (IEC) barrier dysfunction is critical to the development of Crohn's disease (CD). However, the mechanism is understudied. We recently reported increased microRNA-31-5p (miR-31-5p) expression in colonic IECs of CD patients, but downstream targets and functional consequences are unknown.<h4>Methods</h4>microRNA-31-5p target genes were identified by integrative analysis of RNA- and small RNA-sequencing data from colonic mucosa and confirmed by qu ...[more]

PMID: 32561494

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Project description:BackgroundEosinophilic chronic rhinitis with nasal polyps (eos-CRSwNP) is a subtype of nasal polyps (NPs) characterized by severe type-2 inflammation and defective epithelial barrier function. The epithelial barrier plays important roles in the pathogenesis of NPs and type-2 inflammation. Particular matter 2.5 (PM2.5) are fine particles with a diameter less than 2.5 μm, containing a mixture of different components. Here, we investigated the impact of PM2.5 on the barrier function of the eos-CRSwNP epithelium and explored the reparative function of budesonide.MethodsSamples from noninflammatory nasal mucosa and eos-CRSwNP were collected to establish an in vitro air-liquid interface cultured model. The cells were exposed to PM2.5 at 50 or 100 µg/ml intermittently for 72 h, with or without budesonide pretreatment. Barrier function and tight junction (TJ) expression were reflected by measuring transepithelial resistance (TER), paracellular flux permeability of fluorescein isothiocyanate-labeled 4-kDa dextran, quantitative real-time polymerase chain reaction (qPCR), and immunofluorescence staining of TJ proteins. Cytokine expression was measured by qPCR and enzyme-linked immunosorbent assay or Luminex.ResultsPM2.5 increased paracellular flux and downregulated TJ protein expression (zona occuldens-1, occludin, and claudin-1), but did not change TER. These changes could be partially restored by budesonide treatment. Interleukin (IL)-8, IL-10, IL-1α, and tissue inhibitor of metalloproteinase (TIMP)-1 concentrations were significantly increased in the culture medium of cells exposed to PM2.5, and budesonide significantly reduced the changes in IL-8, IL-1α, and TIMP-1.ConclusionPM2.5 impaired the barrier function of eos-CRSwNP epithelial cells and increased the permeability of large molecules. PM2.5 also increased the secretion of pro-inflammatory cytokines by nasal epithelial cells. Budesonide could partially repair the damage, suggesting potential applications in clinical practice.

Project description:BackgroundCrohn's disease (CD) is highly heterogeneous, due in large part to variability in cellular processes that underlie the natural history of CD, thereby confounding effective therapy. There is a critical need to advance understanding of the cellular mechanisms that drive CD heterogeneity.MethodsWe performed small RNA sequencing of adult colon tissue from CD and NIBD controls. Colonic epithelial cells and immune cells were isolated from colonic tissues, and microRNA-31 (miR-31) expression was measured. miR-31 expression was measured in colonoid cultures generated from controls and patients with CD. We performed small RNA-sequencing of formalin-fixed paraffin-embedded colon and ileum biopsies from treatment-naive pediatric patients with CD and controls and collected data on disease features and outcomes.ResultsSmall RNA-sequencing and microRNA profiling in the colon revealed 2 distinct molecular subtypes, each with different clinical associations. Notably, we found that miR-31 expression was a driver of these 2 subtypes and, further, that miR-31 expression was particularly pronounced in epithelial cells. Colonoids revealed that miR-31 expression differences are preserved in this ex vivo system. In adult patients, low colonic miR-31 expression levels at the time of surgery were associated with worse disease outcome as measured by need for an end ileostomy and recurrence of disease in the neoterminal ileum. In pediatric patients, lower miR-31 expression at the time of diagnosis was associated with future development of fibrostenotic ileal CD requiring surgeryCONCLUSIONS. These findings represent an important step forward in designing more effective clinical trials and developing personalized CD therapies.FundingThis work was supported by CCF Career Development Award (SZS), R01-ES024983 from NIEHS (SZS and TSF), 1R01DK104828-01A1 from NIDDK (SZS and TSF), P01-DK094779-01A1 from NIDDK (SZS), P30-DK034987 from NIDDK (SZS), 1-16-ACE-47 ADA Pathway Award (PS), UNC Nutrition Obesity Research Center Pilot & Feasibility Grant P30DK056350 (PS), CCF PRO-KIIDS NETWORK (SZS and PS), UNC CGIBD T32 Training Grant from NIDDK (JBB), T32 Training Grant (5T32GM007092-42) from NIGMS (MH), and SHARE from the Helmsley Trust (SZS). The UNC Translational Pathology Laboratory is supported, in part, by grants from the National Cancer Institute (3P30CA016086) and the UNC University Cancer Research Fund (UCRF) (PS).

Dataset Information

Decreased Colonic Activin Receptor-Like Kinase 1 Disrupts Epithelial Barrier Integrity in Patients With Crohn's Disease.

Publications

Decreased Colonic Activin Receptor-Like Kinase 1 Disrupts Epithelial Barrier Integrity in Patients With Crohn's Disease.

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