Crohn's Disease Phenotypes and Associations With Comorbidities, Surgery Risk, Medications and Nonmedication Approaches: The MAGIC in IMAGINE Study.

BACKGROUND: We aimed to establish a cohort of persons with Crohn's disease (CD) enrolled from 14 Canadian centers to describe the contemporary presentation of CD in Canada. METHODS: All enrollees were at least 18 years old and underwent chart review for phenotype documentation by Montreal Classification at time of enrollment, comorbidities, inflammatory bowel disease (IBD) and other surgeries, and use IBD and other therapies. RESULTS: Of 2112 adults, 59% were female, and the mean age was 44.1 (+/-14.9SD) years. The phenotype distribution was B1 = 50.4%, B2 = 22.4%, B3 = 17.3%, and missing information = 9.9%. Perineal disease was present in 14.2%. Pertaining to disease location, 35.2% of patients had disease in L1, 16.8% in L2, 48% in L3, and 0.4% in L4. There was no difference in phenotype by gender, anxiety score, depression score. Disease duration was significantly different depending on disease behavior type (B1 = 12.2 ±â€…10.1; B2 = 19.4 ±â€…12.9; B3 = 18.9 ±â€…11.8, P < .0001). Isolated colonic disease was much less likely to be fibrostenotic or penetrating than inflammatory disease. Penetrating disease was more likely to be associated with ileocolonic location than other locations. Perineal disease was most commonly seen in persons with B3 disease behavior (24%) than other behaviors (11% B1; 20% B2 disease, P < .0001) and more likely to be seen in ileocolonic disease (L3;19%) vs L2 (17%) and L1 (11%; P < .0001). Surgery related to IBD occurred across each behavior types at the following rates: B1 = 23%, B2 = 64%, and B3 = 74%. Inflammatory bowel disease-related surgery rates by location of disease were L1 = 48%, L2 = 21%, and L3 = 51%. CONCLUSIONS: In exploring this large contemporary CD cohort we have determined that inflammatory disease is the main CD phenotype in Canada and that CD-related surgery remains very common.

Constipated patients with functional defecatory disorder have secondary rectal hyposensitivity due to altered rectal biomechanics.

BACKGROUND AND AIM: Rectal hyposensitivity (RH) is common in constipation and often coexists with functional defecatory disorder (FDD). Rectal sensory thresholds are routinely evaluated with the anorectal manometry probe; however, the gold standard for the assessment of rectal sensitivity is with a barostat, use of which is limited by time constraints and availability. A novel rapid barostat bag (RBB) may facilitate measurements of rectal sensitivity. The aim is to evaluate the relationship between RH (measured by the RBB) and FDD (defined as any minor disorder of rectoanal coordination by the London classification) in constipated patients. METHODS: Consecutive constipated patients referred for anorectal function testing underwent anorectal manometry with the 3D-HDAM probe as well as rectal sensation testing with the RBB pump. RH was defined by volume to first sensation >30%, urge to defecate >80%, or discomfort >100% (normalized to rectal capacity). RESULTS: Fifty-three percent of constipated patients had RH. Patients with FDD had a significantly increased volume to first sensation (134.5 mL vs 102.0, P = 0.02), urge to defecate (187.0 mL vs 149.0, P = 0.04), and rectal capacity (253.5 mL vs 209.0, P = 0.04) compared to constipated patients without FDD. There was no difference in normalized sensory thresholds (percent of rectal capacity) nor the prevalence of hyposensitivity to each sensory threshold nor overall hyposensitivity. CONCLUSION: Patients with FDD, when measured with the RBB, have increased sensory thresholds on volumetric distension, but RH was not observed when sensory threshold volume were normalized to rectal capacity. This may reflect "secondary" RH due to altered rectal biomechanics.

Esophageal motility in systemic sclerosis before and after autologous hematopoietic cell transplantation.

INTRODUCTION: Systemic sclerosis (SSc) is associated with esophageal dysmotility. Autologous hematopoietic cell transplantation (HCT) results in improvement of skin tightness and lung function. Whether esophageal motility improves after HCT is unknown. METHODS: Esophageal motility was studied using high-resolution esophageal manometry in 21 SSc patients before and at multiple time points after autologous HCT. Median posttransplant follow-up was 2 years (range, 6 months to 5 years). RESULTS: Prior to HCT, all 21 patients had abnormal motility-10 (48%) had unmeasurable and 11 (52%) had measurable peristalsis. Manometric diagnosis in the former 10 patients was "absent contractility" and in the latter 11 patients "ineffective esophageal motility (IEM)." After HCT, among the 10 patients with absent contractility, 9 continued to have absent contractility and one demonstrated weak measurable peristalsis. Of the 11 patients with IEM, 5 experienced SSc relapse, and 2 out of these 5 patients developed absent contractility. Among the 6 non-relapsed patients, 4 continued to have IEM, and 2 developed normal motility. CONCLUSIONS: HCT appears to have no beneficial effect on motility in patients with unmeasurable peristalsis. In patients with measurable peristalsis, HCT appears to stabilize and in some normalize motility, unless relapse occurs. Key Points • In patients with systemic sclerosis, esophageal dysmotility is a significant contributor to morbidity and so far, there has been no data describing the effects of hematopoietic cell transplantation on esophageal motility. • Our work demonstrated that in patients with systemic sclerosis and unmeasurable esophageal peristalsis prehematopoietic cell transplantation, there was no measurable beneficial effect of transplantation on esophageal motility. • In patients with systemic sclerosis and measurable peristalsis prehematopoietic cell transplantation, esophageal motility stabilized, except in relapsed patients.

Manometric parameters, when measured with the 3-dimensional high-definition anorectal manometry probe, poorly predict prolonged balloon expulsion time.

BACKGROUND: There are little data evaluating the performance of the 3-dimensional high-definition anorectal manometry (3D-HDAM) system in the diagnosis of dyssynergic defecation. Physical properties of the thicker, rigid, 3D-HDAM probe may have implications on the measurements of anorectal pressures. AIM: Our aim was to compare 3D-HDAM to balloon expulsion test and magnetic resonance (MR) defecography. METHODS: Consecutive constipated patients referred for anorectal function testing at the Calgary Gut Motility Centre (Calgary, Canada) between 2014 and 2019 were assessed. All patients underwent anorectal manometry with the 3D-HDAM probe, and a subset underwent BET or MR defecography. Anorectal manometric variables were compared between patients who had normal and abnormal BET. RESULTS: Over the study period, 81 patients underwent both 3D-HDAM and BET for symptoms of constipation. 52 patients expelled the balloon within 3 minutes. Patients with abnormal BET had significantly lower rectoanal pressure differential (RAPD) (-61 vs. -31 mmHg for normal BET, p = 0.03) and defecation index (0.29 vs. 0.56, p = 0.03). On logistic regression analysis, RAPD (OR: 0.99, 95% CI: 0.97-0.99, p = 0.03) remained a negative predictor of abnormal BET. On ROC analysis, RAPD had an AUC of 0.65. There was good agreement between dyssynergic patterns on 3D-HDAM and defecographic evidence of dyssynergia (sensitivity 80%, specificity 90%, PLR 9, NLR 0.22, accuracy 85%). CONCLUSIONS: Manometric parameters, when measured with the 3D-HDAM probe, poorly predict prolonged balloon expulsion time. RAPD remains the best predictor of prolonged balloon expulsion time. The 3D-HDAM probe may not be the ideal tool to diagnose functional defecatory disorders.

Prucalopride in diabetic and connective tissue disease-related gastroparesis: Randomized placebo-controlled crossover pilot trial.

BACKGROUND: Gastroparesis, defined by delayed gastric emptying in the absence of mechanical outlet obstruction, is a frequent neuropathic complication of diabetes mellitus, and effective treatments are lacking. Prucalopride is a pan-gut prokinetic with selective agonist effects on serotonin 5-HT4 receptors in the gut. This study aimed to assess the effect of prucalopride 4 mg daily on Gastroparesis Cardinal Symptom Index (GCSI), meal-related symptom score (MRSS), and gastric emptying rate in diabetic or connective tissue disease (CTD)-related gastroparesis patients. METHODS: This was a double-blind crossover trial of four-week treatment periods with prucalopride or placebo divided by two weeks of washout. GSCI, MRSS, gastric emptying scintigraphy, PAGI-SYM, and PAGI-QoL were assessed at baseline and the end of each treatment period. Daily bowel movement (BM) frequency and gastrointestinal symptoms were recorded in each period. KEY RESULTS: Fifteen gastroparesis patients (13 diabetic, 2 CTD) were enrolled. GCSI scores were lower than baseline but not different between treatment arms. MRSS scores over time or cumulative score were not significantly different between groups. Gastric emptying was more rapid in the prucalopride treatment period, with mean four-hour meal retention of 22 ± 6% in PRU period vs 40 ± 9% in the placebo period (P = 0.05). Weekly BM frequency was significantly higher in prucalopride than placebo periods (10.5 ± 1.8 vs 7.5 ± 0.8, P < 0.0001). Perception of weight loss was higher in patients on prucalopride. Analysis of diabetic gastroparesis (n = 13) population did not change the conclusions. CONCLUSION AND INFERENCE: Prucalopride at 4 mg accelerates gastric emptying and bowel movement frequency but does not appear to ameliorate gastroparesis or meal-related symptoms in this study.

Anticholinergic, anti-depressant and other medication use is associated with clinically relevant oesophageal manometric abnormalities.

BACKGROUND: Medications can affect gastrointestinal tract motility. However, their effects on oesophageal motility in particular are often not as widely known or may be underestimated. AIM: To review the effect of existing medication use on high-resolution oesophageal manometry (HRM) in a 'real-world' setting. METHODS: Adult patients with upper gut symptoms and normal endoscopy or imaging who had HRM over a 22-month period were analysed. Achalasia and major disorders of peristalsis were excluded. All medications taken within 24 hours of the procedure were prospectively recorded and compared with HRM results, controlling for age, gender and proton pump inhibitor use. RESULTS: A total of 502 patients (323 female, mean age 51) were recruited. Of these, 41.2% had normal oesophageal HRM, while 41.4% had ineffective oesophageal motility (IOM) and 7.6% had oesophagogastric junction outflow obstruction (OGJOO). Serotonin/norepinephrine reuptake inhibitors (SNRI) and opioids were associated with significantly higher resting lower oesophageal sphincter pressure. Benzodiazepines and opioids were associated with elevated integrated relaxation pressure. SNRI and inhaled beta-agonists were associated with increased distal contractile index, whereas calcium channel blockers were associated with a lower distal contractile index. Odds ratio of being on anticholinergics was higher in IOM patients vs normal (3.6, CI 1.2-10.8). Odds ratio for anticholinergics, inhaled beta-agonists, anticonvulsants, SNRIs and opioids (trend) were all > 3 for OGJOO patients vs normal. CONCLUSION: Many medication classes are associated with abnormal HRM variables and diagnoses such as OGJOO and IOM; some of these associations are probably causal. These possible links should be taken into consideration during manometry interpretation.

Gastroesophageal Reflux Disease Is Not Associated With Jackhammer Esophagus: A Case-control Study.

BACKGROUND/AIMS: The pathophysiology of jackhammer esophagus (JE) remains unknown but may be related to gastroesophageal reflux disease or medication use. We aim to determine if pathologic acid exposure or the use of specific classes of medications (based on the mechanism of action) is associated with JE. METHODS: High-resolution manometry (HRM) studies from November 2013 to March 2019 with a diagnosis of JE were identified and compared to symptomatic control patients with normal HRM. Esophageal acid exposure and medication use were compared between groups. Multivariate regression analysis was performed to look for predictors of mean distal contractile integral. RESULTS: Forty-two JE and 127 control patients were included in the study. Twenty-two (52%) JE and 82 (65%) control patients underwent both HRM and ambulatory pH monitoring. Two (9%) JE patients and 14 (17%) of controls had evidence of abnormal acid exposure (DeMeester score > 14.7); this difference was not significant (P = 0.290). Thirty-six (86%) JE and 127 (100%) control patients had complete medication lists. Significantly more JE patients were on long-acting beta agonists (LABA) (JE = 5, control = 4; P = 0.026) and calcium channel blockers (CCB) (JE = 5, control = 3; P = 0.014). Regular opioids (ß = 0.298, P = 0.042), CCB (ß = 0.308, P = 0.035), and inhaled anticholinergics (ß = 0.361, P = 0.049) predicted mean distal contractile integral (R2 = 0.082, F = 4.8; P = 0.003). CONCLUSIONS: Pathologic acid exposure does not appear to be associated with JE. JE patients had increased CCB and LABA use. The unexpected finding of increased LABA use warrants more investigation and may provide support for a cholinergic etiology of JE.

Esophagogastric junction outflow obstruction on manometry: Outcomes and lack of benefit from CT and EUS.

BACKGROUND: Esophagogastric junction outflow obstruction (EGJOO) is a manometric diagnosis based on the Chicago Classification defined by inadequate relaxation of the gastroesophageal junction (GEJ) with swallowing, but with sufficient peristalsis such that the criteria for achalasia are not met. Possible causes include anatomical and functional etiologies. Further investigations, including computed tomography (CT) of the chest and endoscopic ultrasound (EUS), to help elucidate the etiology of EGJOO have been suggested, but the utility of this approach has not been proven. METHODS: All new diagnoses of EGJOO made in the calendar years 2015-2016 were included. A review was performed for each patient to assess clinical outcomes, diagnostic, and therapeutic interventions after the EGJOO diagnosis. KEY RESULTS: 107 EGJOO patients were included. Their primary complaints were dysphagia (68%), chest pain (12%), reflux (8%), pre-operative assessment (6%), regurgitation (3%), and cough (3%). The mean IRP was 21.8 mm Hg. After a mean follow-up period of 463 days, the etiology of EGJOO remained undetermined in 67% of patients. 48% of patients were investigated with cross-sectional imaging (and 10% with EUS to rule out external compression or malignancy as a cause of EGJOO; none of these tests provided any further useful information). In only two cases did the EGJOO progress to achalasia. CONCLUSIONS & INFERENCES: EGJOO is a manometric diagnosis with unclear clinical significance and outcome. CT and EUS of the GEJ were unhelpful at determining the cause of this entity. In this series, very few appear to progress to achalasia, none were due to malignancy, and many resolved spontaneously.

Reversible Jackhammer esophagus in a patient with a gastric band.

We wish to report a case of Jackhammer esophagus in a patient with laparoscopic gastric band, with confirmed resolution of hypertensive peristalsis on deflation of the band. This finding adds to the growing body of evidence that outlet obstruction plays an important role in the pathophysiology of Jackhammer esophagus, which remains incompletely defined.

Real-world clinical, endoscopic and radiographic efficacy of vedolizumab for the treatment of inflammatory bowel disease.

BACKGROUND: Vedolizumab is an α4ß7 integrin antagonist with proven efficacy for inducing and maintaining clinical response and remission in Crohn's disease (CD) and ulcerative colitis (UC). AIM: To evaluate clinical and objective response and remission rates with vedolizumab in a large, real world cohort. METHODS: A retrospective cohort study of adult CD and UC patients receiving vedolizumab between 2012 and 2017 was conducted. PRIMARY OUTCOME: clinical or objective response and remission at 3, 6 and 12 months after induction. Clinical remission was defined by complete, steroid-free absence of symptoms. Objective remission was defined by endoscopic mucosal healing or normalisation of radiographic appearance on contrast-enhanced ultrasound or CT/MR enterography. RESULTS: The study included 222 vedolizumab patients (122 CD, 100 UC). In CD, clinical remission at 3, 6 and 12 months was achieved in 19.8% (22/111), 22.1% (21/95) and 22.1% (15/68) of patients, respectively. Objective remission occurred in 11.5% (6/52), 21.2% (14/66), and 18.9% (7/37) of patients at 3, 6 and 12 months, respectively. In UC, clinical remission at 3, 6, and 12 months was 51.0% (51/100), 61.8% (55/89) and 61.9% (39/63), respectively. Endoscopic remission occurred in 27.5% (11/40), 41.0% (16/39) and 47.8% (22/46) of patients at 3, 6 and 12 months, respectively. In multivariable analysis, patients with UC as compared to CD, and those with milder disease activity were more likely to achieve objectively defined remission at both 6 and 12 months. CONCLUSIONS: Vedolizumab was effective for induction and maintenance of clinical and objective remission, both in Crohn's disease and ulcerative colitis.

Hydrogen and Methane-Based Breath Testing in Gastrointestinal Disorders: The North American Consensus.

OBJECTIVES: Breath tests (BTs) are important for the diagnosis of carbohydrate maldigestion syndromes and small intestinal bacterial overgrowth (SIBO). However, standardization is lacking regarding indications for testing, test methodology and interpretation of results. A consensus meeting of experts was convened to develop guidelines for clinicians and research. METHODS: Pre-meeting survey questions encompassing five domains; indications, preparation, performance, interpretation of results, and knowledge gaps, were sent to 17 clinician-scientists, and 10 attended a live meeting. Using an evidence-based approach, 28 statements were finalized and voted on anonymously by a working group of specialists. RESULTS: Consensus was reached on 26 statements encompassing all five domains. Consensus doses for lactulose, glucose, fructose and lactose BT were 10, 75, 25 and 25 g, respectively. Glucose and lactulose BTs remain the least invasive alternatives to diagnose SIBO. BT is useful in the diagnosis of carbohydrate maldigestion, methane-associated constipation, and evaluation of bloating/gas but not in the assessment of oro-cecal transit. A rise in hydrogen of ≥20 p.p.m. by 90 min during glucose or lactulose BT for SIBO was considered positive. Methane levels ≥10 p.p.m. was considered methane-positive. SIBO should be excluded prior to BT for carbohydrate malabsorption to avoid false positives. A rise in hydrogen of ≥20 p.p.m. from baseline during BT was considered positive for maldigestion. CONCLUSIONS: BT is a useful, inexpensive, simple and safe diagnostic test in the evaluation of common gastroenterology problems. These consensus statements should help to standardize the indications, preparation, performance and interpretation of BT in clinical practice and research.

Prospective evaluation of gastric neurostimulation for diabetic gastroparesis in Canada.

BACKGROUND: The efficacy of gastric neurostimulation therapy for diabetic gastroparesis (GP) in a 'real-life' Canadian setting has not been assessed. AIMS: To assess changes in health-related quality of life (QoL), weekly vomiting frequency (WVF), total symptom score (TSS) and health care utilization 12 months before and after gastric neurostimulator implantation in a diabetic GP cohort. METHODS: Medication-refractory diabetic GP patients (n=7, four female, mean age 42 years) were prospectively recruited from 2008 to 2012. QoL scores were self-administered and obtained at baseline, 24 and 48 weeks postimplantion. WVF and TSS were assessed similarly. Health care usage, measured as hospitalization frequency and medication cost, was obtained six and 12 months before and after implant. Changes from baseline to six and 12 months for all outcomes were compared. RESULTS: The mean ( ± SD) QoL according to EuroQol was significantly better at 24 weeks after the baseline measurement (baseline 29 ± 5, 24 weeks 52 ± 7; P = 0.03). The mean improvement in TSS was significantly better at one year postintervention (baseline score 35 ± 5 versus 12 months 27 ± 3; P = 0.03). Changes in Short-Form 36 Health Survey and WVF were not significant. Days of GP-related hospitalization were highly variable but decreased from a median of 71 days (range 0 to 227 days) to 29 days (range two to 334 days) one year before and after surgery, respectively (P = 0.735). Outpatient medication costs did not decrease to a significant extent. CONCLUSION: Gastric neurostimulation for diabetic GP appeared to show some beneficial palliative effects overall in the present small open-label series, but the effect is highly variable among patients, and placebo effect cannot be ruled out.

Venous thromboembolism in cirrhosis: a review of the literature.

Although hemorrhage has traditionally been regarded as the most significant hemostatic complication of liver disease, there is increasing recognition that hypercoagulability is a prominent aspect of cirrhosis. Identifying markers of coagulability and monitoring anticoagulation therapy in the setting of cirrhosis is problematic. The bleeding risk of venous thromboembolism (VTE) prophylaxis and treatment in patients with chronic liver disease is unclear and there are currently no recommendations to guide practice in this regard. In the present report, the mechanism of coagulation disturbance in chronic liver disease is reviewed with an examination of the evidence for an increased VTE risk in cirrhosis. Finally, the available evidence is assessed for prophylaxis and therapy of VTE in chronic liver disease, and the role it may play in decreasing clinical decompensation and improving survival.

The prevalence of gestational diabetes mellitus and glucose abnormalities in pregnant women with hepatitis C virus infection in British Columbia.

OBJECTIVE: There is evidence to support an association between diabetes mellitus (DM) and hepatitis C virus (HCV) infection. The insulin resistant state of pregnancy suggests a predisposition to developing gestational diabetes mellitus (GDM) in women infected with HCV. The aim of this study was to compare the prevalence of GDM and impaired glucose tolerance (IGT) of pregnancy between women infected with HCV and the general population of British Columbia screened for GDM. METHODS: The HCV cohort was drawn from a population-based prospective cohort of 148 pregnant women infected with HCV in British Columbia. GDM screening tests were completed in 84 women. The prevalence of GDM and IGT of pregnancy in the general population of British Columbia was estimated by acquiring 24 321 GDM screening tests performed by the two major laboratories in the province. RESULTS: Non-compliance was the primary reason for incomplete screening. The prevalence of GDM was 9.5% in the HCV cohort and 6.8% in the screened general population (chi-square test P = 0.33). Similarly, there was no difference in IGT of pregnancy between the two cohorts (2.4% vs. 3.5%; chi-square test P = 0.57). CONCLUSION: A difference in the prevalence of either GDM or IGT of pregnancy was not detected between HCV-infected patients who were screened for GDM and those screened in the general population. Further studies are required to assess whether HCV infection is an independent risk factor for GDM.

The use of endoscopic ultrasonography and other imaging modalities in the preoperative staging of rectal villous tumours: a case of overstaging by magnetic resonance imaging.

A case of a 60-year-old man with recurrent rectal villous adenoma is described. Preoperative staging with endoscopic ultrasound (EUS) and magnetic resonance imaging (MRI) revealed very discordant results. EUS showed a tumour present in the mucosa with no submucosal invasion, while MRI revealed invasion of the muscularis propria consistent with an invasive stage T2 carcinoma. Based on the MRI findings, the patient underwent a low anterior resection of the tumour. The surgical pathology specimen revealed a villous adenoma with lowgrade dysplasia but no carcinoma and no extension into the muscularis propria. The present case highlights the uncertainty that currently exists as to which imaging modality provides the greatest accuracy in the staging of rectal cancer and in guiding the type of surgical procedure performed. Two recent meta-analyses and a systematic review of the literature point to EUS as the imaging modality of choice for determining muscularis propria and perirectal tissue invasion, as well as nodal involvement.

Thrombin receptor: An endogenous inhibitor of inflammatory pain, activating opioid pathways.

Serine proteases such as thrombin, trypsin and mast cell tryptase can act on different cell types through protease-activated receptors (PARs). These receptors have been shown to be implicated in several phenomena such as inflammation, platelet activation, immune response and atherosclerosis. Several studies recently reported PARs expression on neurons and some of them demonstrated that these receptors could interfere with nociception. The contribution of PAR(1) to inflammatory pain and the mechanism involved in this phenomenon were investigated. Intraplantar injection of PAR(1) agonist increased withdrawal latency and reduced response frequency to von Frey filaments, thus inhibiting nociceptive response to both mechanical and thermal stimuli in mice. PAR(1) agonist also reduced carrageenan-induced inflammatory hyperalgesia. The anti-nociceptive effects of PAR(1) agonist were mediated by endogenous opioids, as this effect was inhibited by local injection of naloxone methiodide, and because intraplantar injection of PAR(1) agonist increased mRNA expression of the endogenous opioid precursor proenkephalin. However, PAR(1) agonist was not able to inhibit calcium signals in isolated sensory neurons exposed to pro-nociceptive agents. Finally, despite similar inflammatory parameters, PAR(1)-deficient mice showed a strong potentiation of inflammatory hyperalgesia induced by the intraplantar injection of either formalin or carrageenan, or in the chronic model of collagen-induced arthritis, compared to wild-type mice. This study highlights a previously unknown endogenous mechanism of analgesia, showing a central role for the thrombin receptor PAR(1) in the regulation of inflammatory pain and as an activator of opioid pathways.

Activation of proteinase-activated receptor-1 inhibits neurally evoked chloride secretion in the mouse colon in vitro.

The proteinase-activated thrombin receptor-1 (PAR-1) belongs to a unique family of G protein-coupled receptors activated by proteolytic cleavage. We studied the effect of PAR-1 activation in the regulation of ion transport in mouse colon in vitro. Expression of PAR-1 in mouse colon was assessed by RT-PCR and immunohistochemistry. To study the role of PAR-1 activation in chloride secretion, mouse colon was mounted in Ussing chambers. Changes in short-circuit current (Isc) were measured in tissues exposed to either thrombin, saline, the PAR-1-activating peptide TFLLR-NH2, or the inactive reverse peptide RLLFT-NH2, before electrical field stimulation (EFS). Experiments were repeated in the presence of either a PAR-1 antagonist or in PAR-1-deficient mice to assess receptor specificity. In addition, studies were conducted in the presence of chloride-free buffer or the muscarinic antagonist atropine to assess chloride dependency and the role of cholinergic neurons in the PAR-1-induced effect. PAR-1 mRNA was expressed in full-thickness specimens and mucosal scrapings of mouse colon. PAR-1 immunoreactivity was found on epithelial cells and on neurons in submucosal ganglia where it was colocalized with both VIP and neuropeptide Y. After PAR-1 activation by thrombin or TFLLR-NH2, secretory responses to EFS but not those to forskolin or carbachol were significantly reduced. The reduction in the response to EFS was not observed in the presence of the PAR-1 antagonist, in PAR-1-deficient mice, when chloride was excluded from the bathing medium, or when atropine was present. PAR-1 is expressed in submucosal ganglia in the mouse colon and its activation leads to a decrease in neurally evoked epithelial chloride secretion.

Activation of proteinase-activated receptor 1 stimulates epithelial chloride secretion through a unique MAP kinase- and cyclo-oxygenase-dependent pathway.

Proteinase-activated receptor 1 (PAR-1) is activated by thrombin and induces chloride secretion by intestinal epithelial cells. To elucidate further the mechanisms whereby PAR-1 stimulates secretion, monolayers of SCBN intestinal epithelial cells were studied in modified Ussing chambers. Short circuit current responses were determined after basolateral application of thrombin and the PAR-1-activating peptide, Ala-parafluoro-Phe-Arg-cyclohexyl-Ala-Citrulline-Tyr (Cit-NH2) in the presence or absence of a variety of signal transduction and cyclo-oxygenase (COX) pathway inhibitors. Increased kinase activity was monitored by immunoprecipitation and Western blot analysis of target phosphoproteins. The PAR-1-induced chloride secretory response was significantly attenuated by inhibitors of the EGF receptor tyrosine kinase, Src-kinase, MEK1/2, as well as by inhibitors of cytosolic phospholipase (cPL) A2, COX-1 and COX-2. PAR-1-induced activation of cPLA2, as shown by Western blot of phosphoserine residues, was blocked in cells treated with the MEK inhibitor U0126, indicating that the MEK-ERK1/2 MAP kinase pathway mediated PAR-1-induced cPLA2 phosphorylation. Our data show that PAR-1-induced chloride secretion in SCBN cells involves Src, EGF receptor trans-activation, activation of a MAPK pathway, phosphorylation of cPLA2, COX activity, but not PGF2alpha or PGE2. These findings may be of clinical importance in inflammatory diseases of the intestine where secretory dysfunction is evident and thrombin levels are elevated.