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Background & Aims: Colonic adenomatous polyps, or adenomas, are frequent precancerous lesions and the origin of most cases of colorectal adenocarcinoma. However, we know from epidemiologic studies that although most colorectal cancers (CRCs) originate from adenomas, only a small fraction of adenomas (3%–5%) ever progress to cancer. At present, there are no molecular markers to guide follow-up surveillance programs. Methods: We profiled, by mass spectrometry–based proteomics combined with machine learning analysis, a selected cohort of formalin-fixed, paraffin-embedded high-grade (HG) adenomas with long clinical follow-up, collected as part of the Danish national screening program. We grouped subjects in the cohort according to their subsequent history of findings: a nonmetachronous advanced neoplasia group (G0), with no new HG adenomas or CRCs up to 10 years after polypectomy, and a metachronous advanced neoplasia group (G1) where individuals developed a new HG adenoma or CRC within 5 years of diagnosis. Results: We generated a proteome dataset from 98 selected HG adenoma samples, including 20 technical replicates, of which 45 samples belonged to the nonmetachronous advanced neoplasia group and 53 to the metachronous advanced neoplasia group. The clear distinction of these 2 groups seen in a uniform manifold approximation and projection plot indicated that the information contained within the abundance of the ∼5000 proteins was sufficient to predict the future occurrence of HG adenomas or development of CRC. Conclusions: We performed an in-depth analysis of quantitative proteomic data from 98 resected adenoma samples using various novel algorithms and statistical packages and found that their proteome can predict development of metachronous advanced lesions and progression several years in advance.

BACKGROUND The laminin gamma 1 chain (LMγ1) is abundant along the crypt-villus axis in the intestinal basement membrane. AIMS We investigated whether a serological biomarker of laminin degradation was associated with disease activity in patients with Crohn's disease (CD) and in rats with dextran sulfate sodium (DSS)-induced colitis. METHODS Serum samples from CD patients (n = 43), healthy subjects (n = 19), and Sprague Dawley rats receiving 5-6% DSS water for five days and regular drinking water for 11 days were included in this study. The LG1M biomarker, a neo-epitope degradation fragment of the LMγ1 chain generated by matrix metalloproteinases-9 (MMP-9), was measured in serum to estimate the level of laminin degradation. RESULTS Serum LG1M was elevated in CD patients with active and inactive disease compared to healthy subjects (p

ObjectiveAlthough clinical guidelines exist, the diagnostic work-up for diagnosing inflammatory bowel disease (IBD) is complex and varies in clinical practice. This study used real-life data to characterise the current diagnostic procedures used to establish IBD diagnoses in a Danish nationwide setting.DesignPerson-level data on patients diagnosed with IBD between 1 January 2014 and 30 June 2018 were linked between Danish health registers. Information on age, sex, registration of other gastrointestinal diseases, and diagnostic procedures (endoscopies, biopsies, and imaging) performed in relation to the first IBD hospital admission was analysed for the total study population and was stratified by IBD type, sex, and age.ResultsThe majority of the 12 871 patients with IBD included underwent endoscopy (84%), had a biopsy taken (84%), and/or underwent imaging procedures (44%). In total, 7.5% of the population (6% for Crohn’s disease and 8% for ulcerative colitis) were diagnosed with IBD despite not undergoing any of these diagnostic procedures. Patients with Crohn’s disease underwent more procedures than patients with ulcerative colitis (94% vs 92%, pConclusionFor 7.5% of patients with IBD, this study did not detect any registrations of the recommended diagnostic procedures for establishing an IBD diagnosis. Further research is needed to examine whether these findings are mainly explained by limitations of the register data or also indicate shortcomings of the general approach to IBD.

PURPOSE: Chemotherapy-induced gastrointestinal toxicity is a common adverse event during chemotherapeutic treatment. No uniformly applicable strategies exist to predict, prevent, or treat gastrointestinal toxicity. Thus, a goal of mucositis research is to identify targets for therapeutic interventions and individualized risk prediction. Fibrinogen C domain containing 1 (FIBCD1) is a transmembrane protein expressed in human intestinal epithelial cells with functions in the innate immune system. Previous observations have shown that FIBCD1 ameliorates dextran sulfate sodium (DSS)-induced intestinal inflammation in vivo. We evaluated the effect of FIBCD1 in a murine model of chemotherapy-induced gastrointestinal toxicity and inflammation.METHODS: Transgenic (Tg) mice overexpressing FIBCD1 in the intestinal epithelium (Fibcd1Tg) and wild-type (WT) littermates (C57BL/6N) were randomized to receive an intraperitoneal injection of doxorubicin 20 mg/kg or saline and were terminated 2 or 7 days after the injection. Gastrointestinal toxicity was evaluated by weight change, intestinal length, villus height/crypt depth, and histological mucositis score. Expression of inflammatory markers (IL-6, IL-1β, and Tnfα) was measured by quantitative real-time PCR in intestinal tissue samples.RESULTS: Following doxorubicin treatment, WT mice exhibited an increased weight loss compared with Tg littermates (p < 0.001). No differences between genotypes were seen in mucositis score, intestinal length, villus height/crypt depth, or IL-6, IL-1β, and Tnfα expression.CONCLUSION: Our findings suggest that FIBCD1 could ameliorate chemotherapy-induced gastrointestinal toxicity by reducing weight loss; however, the mechanism of this possible protective effect remains to be defined warranting additional investigations.

Remote ischemic conditioning (RIC) by repetitive brief periods of limb ischemia and reperfusion renders organs more resistant to ischemic injury. The protection is partly through down-regulation of the inflammatory response. Our aim was to investigate the clinical and anti-inflammatory effects of RIC in patients with active ulcerative colitis (UC). We included 22 patients with active UC in this explorative, randomized, sham-controlled clinical trial. The patients were randomly assigned 1:1 to RIC (induced in the arm through four cycles of 5-min inflation and 5-min deflation of a blood-pressure cuff) or sham (incomplete inflation of the blood-pressure cuff) once daily for 10 days. Outcome variables were measured at baseline and on day 11. When compared with sham, RIC did not affect inflammation in the UC patients measured by fecal calprotectin, plasma C-reactive protein, Mayo Score, Mayo Endoscopic Subscore, Nancy Histological Index or inflammatory cytokines involved in UC and RIC. The mRNA and miRNA expression profiles in the UC patients were measured by RNA sequencing and multiplexed hybridization, respectively, but were not significantly affected by RIC. We used the Langendorff heart model to assess activation of the organ protective mechanism induced by RIC, but could not confirm activation of the organ protective mechanism in the UC patients.

The effect of remote pre- and postconditioning on anastomotic healing has been sparsely studied. The aim of our study was to investigate whether remote ischemic conditioning (RIC) applied before and after the creation of a small bowel anastomosis had an effect on anastomotic healing on postoperative day five evaluated by a tensile strength test and histological analysis.Twenty-two female piglets were randomized into two groups. The intervention group (Mean tensile strength when the first transmural rupture appeared (MATS-2) was significantly lower in the first anastomosis in the intervention group compared to the control group (11.4 N vs 14.7 N,In conclusion, post-anastomotic remote ischemic conditioning had a detrimental effect and pre-anastomotic conditioning seems to have no effect on small intestinal anastomotic strength.

Introduction: Chemotherapy-induced gastrointestinal toxicity (CIGT) is a frequent, severe and dose-limiting side effect. Few treatments have proven effective for CIGT. CIGT is characterized by activation of the nuclear factor kappa B pathway which, leads to upregulation of proinflammatory cytokines. The innate immune protein peptidoglycan recognition peptide 2 (PGLYRP2) binds to and hydrolyzes microbial peptidoglycan. Expression of PGLYRP2 is upregulated in the intestine of chemotherapy-treated piglets. In this experimental study, we investigated the role of Pglyrp2 in the development and severity of murine CIGT.Methods:Pglyrp2 wildtype and Pglyrp2 knockout mice received intraperitoneal injections of chemotherapy (Doxorubicin 20 mg/kg) to induce CIGT. Weight was monitored daily, and animals were euthanized after 2 or 7 days. Expression of proinflammatory cytokines in the jejunum was measured by quantitative real-time polymerase-chain reaction and enzyme-linked immunosorbent assay. Villus height, crypt depth, and histologic inflammation were evaluated on haematoxylin and eosin stained tissue specimens.Results: Chemotherapeutic treatment induced weight loss (p < 0.05), shortening of the small intestine (p < 0.05), elongation of villus height (p < 0.05), increased crypt depth (p < 0.05), and led to elevated mRNA levels of II1β (p < 0.05), II6 (p < 0.05), and Tnf (p < 0.001) at day 2. Protein levels of IL1β, IL6, and TNFα did not change after exposure to chemotherapy. Doxorubicin treated wildtype mice had a more pronounced weight loss compared to knockout mice from day 3 to day 7 (D3-D6: p < 0.05 and D7: p < 0.01). No other phenotypic differences were detected.Conclusion:Pglyrp2 aggravates chemotherapy-induced weight loss but does not induce a specific pattern of inflammation and morphological changes in the small intestine.

Formalin fixation and paraffin-embedding (FFPE) is the most common method to preserve human tissue for clinical diagnosis, and FFPE archives represent an invaluable resource for biomedical research. Proteins in FFPE material are stable over decades but their efficient extraction and streamlined analysis by mass spectrometry (MS)-based proteomics has so far proven challenging. Herein we describe a MS-based proteomic workflow for quantitative profiling of large FFPE tissue cohorts directly from histopathology glass slides. We demonstrate broad applicability of the workflow to clinical pathology specimens and variable sample amounts, including low-input cancer tissue isolated by laser microdissection. Using state-of-the-art data dependent acquisition (DDA) and data independent acquisition (DIA) MS workflows, we consistently quantify a large part of the proteome in 100 min single-run analyses. In an adenoma cohort comprising more than 100 samples, total workup took less than a day. We observed a moderate trend towards lower protein identification in long-term stored samples (>15 years), but clustering into distinct proteomic subtypes was independent of archival time. Our results underscore the great promise of FFPE tissues for patient phenotyping using unbiased proteomics and they prove the feasibility of analyzing large tissue cohorts in a robust, timely, and streamlined manner. (c) 2020 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Background: Chronic intestinal inflammation results in tissue damage partly caused by an increase in matrix metalloproteinases (MMP) activity causing degradation of extracellular matrix (ECM) proteins. We studied intestinal tissue remodeling by quantifying ECM protein fragments in serum in dextran sulfate sodium (DSS)-induced colitis, to investigate ECM protein fragments as serological biomarkers of intestinal tissue remodeling and disease activity. Methods: Male Sprague–Dawley rats received 5% DSS in drinking water for 5 days followed by 11 days with regular water. Disease activity index (DAI) was scored daily. Serum was collected on day 0, 6, 7, and 16. ELISAs were used to quantify MMP-derived remodeling fragments of basement membrane type IV collagen (C4M and PRO-C4) and interstitial matrix type III collagen (C3M and rPRO-C3). Results: In DSS rats, serum levels relative to baseline of C4M, PRO-C4, and C3M were elevated (P < 0.01; P < 0.001; P < 0.001) at day 7, which declined at day 16. Levels of rPRO-C3 were lower in DSS rats at day 7 and increased to normal levels at day 16. The ratio between C3M and rPRO-C3 showed an overall degradation (P < 0.0001) of collagen type III in DSS rats at day 7, which correlated to the DAI (r 2 = 0.5588, P < 0.0001). Conclusion: Our data suggest that remodeling of the basement membrane (C4M and PRO-C4) and the interstitial matrix (C3M and rPRO-C3) increased during DSS-induced colitis and declined with reversal of the disease. Thus, serological biochemical biomarkers of the ECM reflect tissue remodeling and could be studied as markers of disease activity in IBD.

Formalin fixation and paraffin-embedding (FFPE) is the most common method to preserve human tissue for clinical diagnosis and FFPE archives represent an invaluable resource for biomedical research. Proteins in FFPE material are stable over decades but their efficient extraction and streamlined analysis by mass spectrometry (MS)-based proteomics has so far proven challenging. Here, we describe an MS-based proteomic workflow for quantitative profiling of large FFPE tissue cohorts directly from pathology glass slides. We demonstrate broad applicability of the workflow to clinical pathology specimens and variable sample amounts, including less than 10,000 cancer cells isolated by laser-capture microdissection. Using state-of-the-art data dependent acquisition (DDA) and data independent (DIA) MS workflows, we consistently quantify a large part of the proteome in 100 min single-run analyses. In an adenoma cohort comprising more than 100 samples, total work up took less than a day. We observed a moderate trend towards lower protein identifications in long-term stored samples (>15 years) but clustering into distinct proteomic subtypes was independent of archival time. Our results underline the great promise of FFPE tissues for patient phenotyping using unbiased proteomics and prove the feasibility of analyzing large tissue cohorts in a robust, timely and streamlined manner.

Purpose: The objective of the present study was to investigate the effect of segmental reversal of the distal 20 cm of the small intestine in piglets with induced SBS compared to controls with SBS alone. Primary endpoint was postoperative weight change over a period of 1 month. Secondary endpoints were the influence on cell proliferation and mucosal architecture shown by histological analysis. Methods: Sixteen piglets underwent a 60% resection of the distal small intestine and were randomized into two groups. Group 1 short bowel syndrome alone (SBS) (n = 8) and group 2 with reversal of a distal small intestinal segment (SBS-RS) (n = 8). Body weight was measured daily and the pigs were euthanized after 1 month. Crypt depths, villus heights and muscle layers thicknesses were measured. For the evaluation of microvilli of the brush border of the epithelium and cell proliferation, immunohistochemical staining with Villin and Ki-67 was performed. Results: No statistically significant differences were observed concerning weight gain. Mean ± SD weight gain was 2.31 ± 2.85 kg for SBS-RS and 2.03 ± 1.27 kg for SBS (p = 0.8). In the proximal jejunal segment a significant increase in villus heights was found in the SBS group and increase in the thickness of the circular and longitudinal muscle layers in the SBS-RS group. In the distal ileal segment the longitudinal muscle layer thicknesses were increased in the SBS group. Otherwise, no significant changes were found. Conclusion: Reversal of a 20-cm distal segment showed no effect on weight gain, but there were some significant histological changes of unknown clinical significance.