Increased Colonic Expression of ACE2 Associates with Poor Prognosis in Crohn's disease.

BACKGROUND AND AIMS: The host receptor for SARS-CoV-2, angiotensin-converting enzyme 2 (ACE2), is highly expressed in small intestine. Our aim was to study colonic ACE2 expression in Crohn's disease (CD) and non-inflammatory bowel disease (non-IBD) controls. We hypothesized that the colonic expression levels of ACE2 impacts CD course. METHODS: We examined the expression of colon ACE2 using RNA-seq and quantitative (q) RT-PCR from 69 adult CD and 14 NIBD control patients. In a subset of this cohort we validated ACE2 protein expression and localization in formalin-fixed, paraffin-embedded matched colon and ileal tissues using immunohistochemistry. The impact of increased ACE2 expression in CD for the risk of surgery was evaluated by a multivariate regression analysis and a Kaplan-Meier estimator. To provide critical support for the generality of our findings, we analyzed previously published RNA-seq data from two large independent cohorts of CD patients. RESULTS: Colonic ACE2 expression was significantly higher in a subset of adult CD patients (ACE2-high CD). IHC in a sampling of ACE2-high CD patients confirmed high ACE2 protein expression in the colon and ileum compared to ACE2-low CD and NIBD patients. Notably, we found that ACE2-high CD patients are significantly more likely to undergo surgery within 5 years of diagnosis, with a Cox regression analysis finding that high ACE2 levels is an independent risk factor (OR 2.18; 95%CI, 1.05-4.55; p=0.037). CONCLUSION: Increased intestinal expression of ACE2 is associated with deteriorated clinical outcomes in CD patients. These data point to the need for molecular stratification that may impact CD disease-related outcomes.

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

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.

Crohn's Disease Differentially Affects Region-Specific Composition and Aerotolerance Profiles of Mucosally Adherent Bacteria.

BACKGROUND: The intestinal microbiota play a key role in the onset, progression, and recurrence of Crohn disease (CD). Most microbiome studies assay fecal material, which does not provide region-specific information on mucosally adherent bacteria that directly interact with host systems. Changes in luminal oxygen have been proposed as a contributor to CD dybiosis. METHODS: The authors generated 16S rRNA data using colonic and ileal mucosal bacteria from patients with CD and without inflammatory bowel disease. We developed profiles reflecting bacterial abundance within defined aerotolerance categories. Bacterial diversity, composition, and aerotolerance profiles were compared across intestinal regions and disease phenotypes. RESULTS: Bacterial diversity decreased in CD in both the ileum and the colon. Aerotolerance profiles significantly differed between intestinal segments in patients without inflammatory bowel disease, although both were dominated by obligate anaerobes, as expected. In CD, high relative levels of obligate anaerobes were maintained in the colon and increased in the ileum. Relative abundances of similar and distinct taxa were altered in colon and ileum. Notably, several obligate anaerobes, such as Bacteroides fragilis, dramatically increased in CD in one or both intestinal segments, although specific increasing taxa varied across patients. Increased abundance of taxa from the Proteobacteria phylum was found only in the ileum. Bacterial diversity was significantly reduced in resected tissues of patients who developed postoperative disease recurrence across 2 independent cohorts, with common lower abundance of bacteria from the Bacteroides, Streptococcus, and Blautia genera. CONCLUSIONS: Mucosally adherent bacteria in the colon and ileum show distinct alterations in CD that provide additional insights not revealed in fecal material.

Colonic epithelial miR-31 associates with the development of Crohn's phenotypes.

BACKGROUND: Crohn'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. METHODS: We 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. RESULTS: Small 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. FUNDING: This 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).

Molecular classification of Crohn's disease reveals two clinically relevant subtypes.

OBJECTIVE: The clinical presentation and course of Crohn's disease (CD) is highly variable. We sought to better understand the cellular and molecular mechanisms that guide this heterogeneity, and characterise the cellular processes associated with disease phenotypes. DESIGN: We examined both gene expression and gene regulation (chromatin accessibility) in non-inflamed colon tissue from a cohort of adult patients with CD and control patients. To support the generality of our findings, we analysed previously published expression data from a large cohort of treatment-naïve paediatric CD and control ileum. RESULTS: We found that adult patients with CD clearly segregated into two classes based on colon tissue gene expression-one that largely resembled the normal colon and one where certain genes showed expression patterns normally specific to the ileum. These classes were supported by changes in gene regulatory profiles observed at the level of chromatin accessibility, reflective of a fundamental shift in underlying molecular phenotypes. Furthermore, gene expression from the ilea of a treatment-naïve cohort of paediatric patients with CD could be similarly subdivided into colon-like and ileum-like classes. Finally, expression patterns within these CD subclasses highlight large-scale differences in the immune response and aspects of cellular metabolism, and were associated with multiple clinical phenotypes describing disease behaviour, including rectal disease and need for colectomy. CONCLUSIONS: Our results strongly suggest that these molecular signatures define two clinically relevant forms of CD irrespective of tissue sampling location, patient age or treatment status.

Alterations to chromatin in intestinal macrophages link IL-10 deficiency to inappropriate inflammatory responses.

Intestinal macrophages (IMs) are uniquely programmed to tolerate exposure to bacteria without mounting potent inflammatory responses. The cytokine IL-10 maintains the macrophage anti-inflammatory response such that loss of IL-10 results in chronic intestinal inflammation. To investigate how IL-10-deficiency alters IM programming and bacterial tolerance, we studied changes in chromatin accessibility in response to bacteria in macrophages from two distinct niches, the intestine and bone-marrow, from both wild-type and IL-10-deficient (Il10(-/-) ) mice. We identified chromatin accessibility changes associated with bacterial exposure and IL-10 deficiency in both bone marrow derived macrophages and IMs. Surprisingly, Il10(-/-) IMs adopted chromatin and gene expression patterns characteristic of an inflammatory response, even in the absence of bacteria. Further, when recombinant IL-10 was added to Il10(-/-) cells, it could not revert the chromatin landscape to a normal state. Our results demonstrate that IL-10 deficiency results in stable chromatin alterations in macrophages, even in the absence of bacteria. This supports a model in which IL-10-deficiency leads to chromatin alterations that contribute to a loss of IM tolerance to bacteria, which is a primary initiating event in chronic intestinal inflammation.

Diverting ostomy with pouch procedure: "causes more morbidity than it prevents!".

Temporary diverting ileostomies are routine performed as a "protective" adjunct to ileal pouch procedures. The need for routine diverting ileostomies has been challenged because of the risks associated with their management. A review of the available data and my personal experience demonstrates that diversion results in lower pouch leaks but a higher instance of pouch failure and strictures. The creation of ileal pouches without an ileostomy is preferable under selective conditions with quite acceptable results.

Prevention of postoperative abdominal adhesions by a novel, glycerol/sodium hyaluronate/carboxymethylcellulose-based bioresorbable membrane: a prospective, randomized, evaluator-blinded multicenter study.

INTRODUCTION: Postoperative abdominal adhesions are associated with significant morbidity and mortality, placing a substantial burden on healthcare systems worldwide. Development of a bioresorbable membrane containing up to 23 percent glycerol and chemically modified sodium hyaluronate/carboxymethylcellulose offers ease of handling and has been shown to provide significant postoperative adhesion prevention in animals. This study was designed to assess the safety of glycerol hyaluronate/carboxymethylcellulose and to evaluate its efficacy in reducing the incidence, extent, and severity of postoperative adhesion development in surgical patients. METHODS: Twelve centers enrolled 120 patients with ulcerative colitis or familial polyposis who were scheduled for a restorative proctocolectomy and ileal pouch-anal anastomosis with diverting loop ileostomy. Before surgical closure, patients were randomized to no anti-adhesion treatment (control) or treatment with glycerol hyaluronate/carboxymethylcellulose membrane under the midline incision. At ileostomy closure, laparoscopy was used to evaluate the incidence, extent, and severity of adhesion formation to the midline incision. RESULTS: Data were analyzed using the intent-to-treat population. Treatment with glycerol hyaluronate/carboxymethylcellulose resulted in 19 of 58 patients (33 percent) with no adhesions compared with 6 of 60 adhesion-free patients (10 percent) in the no treatment control group (P = 0.002). The mean extent of postoperative adhesions to the midline incision was significantly lower among patients treated with glycerol hyaluronate/carboxymethylcellulose compared with patients in the control group (P < 0.001). The severity of postoperative adhesions to the midline incision was significantly less with glycerol hyaluronate/carboxymethylcellulose than with control (P < 0.001). Adverse events were similar between treatment and no treatment control groups with the exception of abscess and incisional wound complications were more frequently observed with glycerol hyaluronate/carboxymethylcellulose. CONCLUSIONS: Glycerol hyaluronate/carboxymethylcellulose was shown to effectively reduce adhesions to the midline incision and adhesions between the omentum and small bowel after abdominal surgery. Safety profiles for the treatment and no treatment control groups were similar with the exception of more infection complications associated with glycerol hyaluronate/carboxymethylcellulose use. Animal models did not predict these complications.

Acute vagotomy does not augment the systemic inflammatory response in patients with peptic ulcer disease.

Recent accumulating evidence suggests that the vagus nerve modulates the response to peripheral immunologic stimuli and that intact vagal mediation decreases the systemic inflammatory response. We hypothesized that patients who had vagotomy for complicated peptic ulcer disease would be at increased risk of an enhanced systemic inflammatory response compared to patients that did not have a vagotomy as part of their operative treatment. Ninety-six patients were identified from 1985 to 2000 and their medical records were reviewed. Patients were assigned to three groups based on the performance of a truncal vagotomy: truncal vagotomy (TV; N = 62 patients), nontruncal vagotomy (NTV; N = 34 patients), or a subgroup of the TV group, acute truncal vagotomy (ATV; N = 40 patients). Operative indications in the NTV and ATV groups were perforation (94% vs 47%) and bleeding (6% vs 53%). Systemic or organ-specific complications did not differ between groups (NTV vs ATV), and the sepsis (24% vs 23%) and mortality rates (29% vs 20%) were similar. The ICU and hospital length of stay did not differ substantially among the groups. This clinical study demonstrated that acute truncal vagotomy does not increase the risk of the systemic inflammatory response in surgical patients with complicated peptic ulcer disease.

Psychological well-being of surgery residents before the 80-hour work week: a multiinstitutional study.

BACKGROUND: Accreditation Council on Graduate Medical Education work-hour restrictions are aimed at improving patient safety and resident well-being. Although surgical trainees will be dramatically affected by these changes, no comprehensive assessment of their well-being has been recently attempted. STUDY DESIGN: A multicenter study of psychological well-being of surgical residents (n = 108) across four US training programs before implementation of the 80-hour work week was performed using two validated surveys (Symptom Checklist-90-R [SCL-90-R] and Perceived Stress Scale [PSS]) during academic year 2002-03. Societal normative populations served as controls. Primary outcomes measures were psychologic distress (SCL-90-R) and perceived stress (PSS). Secondary outcomes measures (SCL-90-R) were somatization, depression, anxiety, interpersonal sensitivity, hostility, obsessive-compulsive behavior, phobic anxiety, paranoid ideation, and psychoticism. The impact of personal variables (age, gender, marital status) and programmatic variables (level of training, laboratory experience, institution) was assessed. RESULTS: Mean psychologic distress was significantly higher in general surgery residents than in the normative population (p < 0.0001), with 38% scoring above the 90th percentile and 72% above the 50th percentile. Mean perceived stress among surgery residents was higher than historic controls (p < 0.0001), with 21% scoring above the 90th percentile and 68% above the 50th percentile. Among secondary outcomes, eight of nine symptom dimensions were significantly higher in surgical residents than in societal controls. In subgroup analyses, male gender was associated with phobic anxiety (p < 0.001) and anxiety (p < 0.05), and junior level of training (PGY 1 to 3) with anxiety (p < 0.05), obsessive-compulsive behavior (p < 0.05), and interpersonal sensitivity (p < 0.05). CONCLUSIONS: More than one-third of general surgery residents meet criteria for clinical psychologic distress. Surgery residents perceive significantly more stress than societal controls. Both personal and programmatic variables likely affect resident well-being and should be considered in assessing the full impact of Accreditation Council on Graduate Medical Education directives and in guiding future restructuring efforts.

Fundoplication improves disordered esophageal motility.

Patients with gastroesophageal reflux disease (GERD) and disordered esophageal motility are at risk for postoperative dysphagia, and are often treated with partial (270-degree) fundoplication as a strategy to minimize postoperative swallowing difficulties. Complete (360-degree) fundoplication, however, may provide more effective and durable reflux protection over time. Recently we reported that postfundoplication dysphagia is uncommon, regardless of preoperative manometric status and type of fundoplication. To determine whether esophageal function improves after fundoplication, we measured postoperative motility in patients in whom disordered esophageal motility had been documented before fundoplication. Forty-eight of 262 patients who underwent laparoscopic fundoplication between 1995 and 2000 satisfied preoperative manometric criteria for disordered esophageal motility (distal esophageal peristaltic amplitude < or =30 mm Hg and/or peristaltic frequency < or =80%). Of these, 19 had preoperative manometric assessment at our facility and consented to repeat study. Fifteen (79%) of these patients had a complete fundoplication and four (21%) had a partial fundoplication. Each patient underwent repeat four-channel esophageal manometry 29.5 +/- 18.4 months (mean +/- SD) after fundoplication. Distal esophageal peristaltic amplitude and peristaltic frequency were compared to preoperative data by paired t test. After fundoplication, mean peristaltic amplitude in the distal esophagus increased by 47% (56.8 +/- 30.9 mm Hg to 83.5 +/- 36.5 mm Hg; P < 0.001) and peristaltic frequency improved by 33% (66.4 +/- 28.7% to 87.6 +/- 16.3%; P < 0.01). Normal esophageal motor function was present in 14 patients (74%) after fundoplication, whereas in five patients the esophageal motor function remained abnormal (2 improved, 1 worsened, and 2 remained unchanged). Three patients with preoperative peristaltic frequencies of 0%, 10%, and 20% improved to 84%, 88%, and 50%, respectively, after fundoplication. In most GERD patients with esophageal dysmotility, fundoplication improves the amplitude and frequency of esophageal peristalsis, suggesting refluxate has an etiologic role in motor dysfunction. These data, along with prior data showing that postoperative dysphagia is not common, imply that surgeons should apply complete fundoplication liberally in patients with disordered preoperative esophageal motility.

Gallbladder function before and after fundoplication.

No study has reported an association between gastroesophageal reflux disease (GERD) or its therapies and gallbladder function. We compared pre- and postoperative gallbladder function in patients undergoing fundoplication to determine the following: (1) whether patients with chronic GERD have preexisting gallbladder motor dysfunction; (2) whether medical or surgical therapy alters gallbladder function; and (3) whether division of the hepatic branch of the anterior vagus nerve is detrimental to gallbladder motility. Nineteen patients with documented GERD consented to a preoperative cholecystokinin-stimulated technetium hepatobiliary (CCK-HIDA) scan to quantify the gallbladder ejection fraction (GBEF). All patients underwent laparoscopic Nissen fundoplication. One month after fundoplication, 12 patients completed a repeat CCK-HIDA scan for determination of GBEF, with comparison to the preoperative GBEF. Among patients with preoperative GERD, 11 (58%) of 19 met the scintigraphic criteria for gallbladder dysfunction (GBEF <35%), which is a ratio comparable to that in patients undergoing a CCK-HIDA scan for presumed biliary dyskinesia during the same time period (31 [60%] of 53; P = NS, chi-square test) and exceeds the rate of abnormal GBEF reported in healthy volunteers (3%). Six of seven patients with a low preoperative GBEF who underwent repeat evaluation postoperatively had normalization of the GBEF (P < 0.05, paired t-test). In the 12 patients who underwent postoperative CCK-HIDA scanning, there was no association between preservation or division of the hepatic branch of the anterior vagus nerve and postoperative gallbladder dysfunction (P = NS, chi-square test). Unexpectedly, 58% of patients with GERD demonstrated gallbladder motor dysfunction prior to fundoplication, with improvement to normal occurring in most of those studied postoperatively. These data support controlled trials to determine the effect of chronic GERD and antisecretory therapy on gallbladder and global gastrointestinal smooth muscle function. Preservation of the hepatic branch of the anterior vagus nerve during fundoplication offered no clear benefit with regard to early postoperative gallbladder function.