Grover, Madhusudan

Abstract Background There is increased recognition of duodenal disturbances (inflammation, altered mucosal protein expression, and chemosensitivity) in functional dyspepsia (FD). Besides sensorimotor functions, enteric submucosal neurons also regulate epithelial ion transport. We hypothesized that duodenal mucosal ion transport and expression of associated genes are altered in FD. Methods Duodenal mucosal ion transport (basal and acetylcholine‐ and glucose‐evoked changes in short‐circuit current [Isc]) and expression of associated genes and regulatory miRNAs were evaluated in 40 FD patients and 24 healthy controls. Results Basal Isc (FD: 88.2 [52.6] μA/cm 2 vs healthy: 20.3 [50.2] μA/cm 2 ; P ≤ .0001), acetylcholine‐evoked Isc (FD: Emax 50.4 [35.8] μA/cm 2 vs healthy: 16.6 [15] μA/cm 2 ; P ≤ .001), and glucose‐evoked Isc responses (FD: E max 69.8 [42.1] μA/cm 2 vs healthy: 40.3 [24.6] μA/cm 2 ; P = .02) were greater in FD than in controls. The Emax for glucose was greater in FD patients on selective serotonin reuptake inhibitors. In FD, the mRNA expression of SLC4A7 and SLC4A4 , which transport bicarbonate into cells at the basolateral surface, and the apical anion exchanger SLC26A3 were reduced (false discovery rate <0.05), the serotonin receptor HTR4 was increased, and the serotonin transporter SLC6A4 was decreased. Selected miRNAs (hsa‐miR‐590‐3p, hsa‐miR‐32‐5p) that target genes associated with ionic transport were upregulated in FD. Conclusions Compared to controls, FD patients had greater baseline and agonist‐evoked duodenal mucosal secretory responses. These findings may be explained by reduced gene expression, which would be anticipated to reduce luminal bicarbonate secretion. The upregulated miRNAs may partly explain the downregulation of these genes in FD.

INTRODUCTION: Intestinal pseudo-obstruction is characterized by impaired transit and luminal dilation in the absence of mechanical obstruction. Our study aims to describe the clinical, radiographic, and physiological findings in pseudo-obstruction associated with systemic sclerosis (SSc), amyloidosis, and paraneoplastic syndrome. METHODS: A retrospective cohort of patients evaluated at our institution between January 1, 2008, and August 1, 2018, was assembled. Clinical, imaging, and physiological characteristics were abstracted from electronic medical records. RESULTS: We identified 100 cases of pseudo-obstruction (55 SSc, 27 amyloidosis, and 18 paraneoplastic). Female population predominance was seen in SSc (71%) vs male population in amyloidosis (74%). Most common symptom was abdominal bloating in all 3 groups. Vomiting was more common in SSc than amyloidosis (73% vs 46%, P = 0.02). Diarrhea was more common in amyloidosis and SSc compared with paraneoplastic (81% and 67% vs 28%, P < 0.01). Weight loss (>5%) was more common in SSc compared with amyloidosis and paraneoplastic (78% vs 31% and 17%, P < 0.0001). Only small bowel dilation was seen in 79%, 40%, and 44% and only large bowel dilation in 2%, 44%, and 44% of patients in SSc, amyloidosis, and paraneoplastic, respectively. Five of 8 SSc patients had myopathic and 3 of 5 paraneoplastic had neuropathic involvement on gastroduodenal manometry. DISCUSSION: SSc-associated pseudo-obstruction demonstrates female population predominance and presents with vomiting, diarrhea, and weight loss. Amyloidosis-associated pseudo-obstruction shows male population predominance. Small bowel is more commonly involved than large bowel on both imaging and transit studies in SSc. Myopathic involvement was more common in SSc, contrary to neuropathic in paraneoplastic syndrome.

BACKGROUND Mechanical stimulation of the gastrointestinal (GI) epithelium causes changes in epithelial ion transport by an unclear mechanism. We recently discovered the mechanosensitive ion channel Piezo2 is specifically expressed by the enteroendocrine cells (EECs) and is necessary for EEC mechanosensitivity. Further, in mouse small bowel, the pharmacologic inhibition or genetic knockout of the epithelial Piezo2 led to an inhibition of the pressure‐induced short‐circuit current (I sc ) in the Ussing chamber. These results suggested that EEC Piezo2 plays a role in pressure‐induced epithelial ion transport, but the mechanism and whether these responses are region‐specific remains uknown. AIM Determine whether epithelial Piezo2‐mediated pressure‐induced I sc is due to Clsecretion in the mouse colon. METHODS Full‐thickness mid‐colon preparations of Vil Cre/+ ::Piezo2 f/f ( Piezo2 CKO ) or Vil +/+ ::Piezo2 f/f ( Piezo2 WT ) mice at 0.3 cm 2 were mounted in 4mL Ussing chambers with normal Krebs solution. The epithelial side was intermittently pressure‐clamped at a range of pressures (5, 10, 20, 30, 40, 60 mmHg) for 10s each. Forskolin (FSK, 10μM) was applied at the end of the experiments, and FSK response was used to normalize the pressure‐induced responses (ΔI sc /ΔI sc,FSK ). RESULTS Pressure applied to the mucosa of Piezo2 CKO and Piezo2 WT colons caused a pressure‐dependent increase in I sc . Responses in Piezo2 CKO colons were smaller than Piezo2 WT at 40mmHg (0.41±0.1 vs. 0.76±0.1) and 60mmHg (0.51±0.1 vs. 0.87±0.1) (n=9–11, P <0.05, two‐way ANOVA). To determine the nature of the pressure‐induced I sc we removed Cl − from the Krebs solutions. Cl‐removal resulted in XX. We found a nearloss in the mechanically induced ΔI sc in both Piezo2 WT tissues and Piezo2 CKO : 5mmHg (− 88.85±0.1% and −87.44±0.1% Piezo2 WT ), 10mmHg (−86.46±0.1% Piezo2 CKO and − 90.30±0.1% Piezo2 WT ), 20mmHg (−88.50±0.01% Piezo2 CKO and −89.86±0.1% Piezo2 WT ), 40mmHg (−91.78±0.1% Piezo2 CKO and −91.35±0.04% Piezo2 WT ), and 60mmHg (−93.93±0.04% Piezo2 CKO and −92.08±0.04% Piezo2 WT (n= 5–10, P <0.05, unpaired t ‐test). Replacement of Cl − led to full recovery of the mechanically induced ΔI sc in both Piezo2 WT and Piezo2 CKO tissues when tested at 30 mmHg ( Piezo2 WT : 2.51±1.4 vs. 73.52±33.8, n=6 and 3, P <0.05, unpaired t ‐test), ( Piezo2 CKO : 1.80±1.0 vs. 45.14±12.4, n=5 and 4, P <0.05, unpaired t ‐test). CONCLUSIONS Hydrostatic pressure applied to the colon epithelium leads to an increase in epithelial secretion that depends chloride and Piezo2 contributes to this mechanosensitive chloride secretion. Support or Funding Information Supported by AGA RSA, NIH DK106456, NIH DK119683, NIH DK123549

Background Campylobacter is a leading cause of bacterial enterocolitis in the U.S. and around the world. In addition to the acute morbidity, it is associated with delayed sequelae such as environmental enteropathy and post‐infection irritable bowel syndrome. Host responses to Campylobacter infection are poorly studied due to colonization resistance in mice thereby restricting virulence studies to immortalized cell lines. These models lack the complexity of the host environment. Recent studies have highlighted the use of organoids for studying enteric pathogens. We propose that human organoids provide a unique platform for studying pathogenic mechanisms incorporating unique host (human) and infection (strain) characteristics. Objective To demonstrate human colonic organoids (colonoids) as a model for studying C. jejuni infection. Methods Sigmoid colonic biopsies were collected from healthy volunteers between 18–35 years of age (n=3). Intestinal crypts were isolated, washed and plated in matrigel domes for colonoid formation. Mature colonoids were harvested after 7 days, and dissociated using a 0.05% trypsin solution. These were plated on collagen IV coated plates, generating 2D‐monolayers for infection experiments. Three different C. jejuni strains (Sequence Types 45, 22, 8) were grown to an OD 600 of 1 and used to infect the colonoid monolayers. To assess adhesion and internalization, colonoids were infected for a total of 3 hours after which monolayers were either washed and subsequently lysed (to determine attached plus invaded bacteria) or treated with gentamycin and then lysed (to determine invaded bacteria). Bacterial quantification was done by plating on Mueller Hinton agar. Additional colonoid monolayers were grown on permeable transwell inserts. Baseline transepithelial resistance (TER) of confluent monolayers was recorded and then continuously assessed every hour after the addition of the C. jejuni strains (using impedance spectroscopy in cellZscope) to evaluate changes in barrier integrity. Results All C. jejuni strains attached and invaded colonoid monolayers derived from all three volunteers. Two strains (ST 45 and 22) caused greater adhesion and invasion in all three colonoids compared to the third strain (Adhesion mean (SEM): 4.32 x 10 −2 (2.33 x 10 −3 ), 9.36 x 10 −3 (4.82 x 10 −4 ) vs 7.79 x 10 −3 (3.51 x 10 −3 ); Invasion mean (SEM) 3.47 x 10 −4 (2.13 x 10 −5 ), 2.33 x 10 −4 (1.36 x 10 −4 ) vs 4.57 x 10 −5 (1.73 x 10 −5 ) (ANOVA, p<0.05, n=3/group for both analyses). Additionally, greater drop in TER was seen in colonoid monolayers exposed to the two strains with higher adhesion and invasion (Mean % of baseline TER at 30 hr post‐infection 2.58% (1.21), 14.14% (3.39) vs 42.29% (18.49), p<0.01). These results correlated with virulence of these strains in conventional T84 monolayers. Transcriptomic and proteomic characterization of the C. jejuni infection in these colonoids is in progress. Conclusions Human colonoids are useful to assess the role of the host in infectivity and epithelial barrier responses to C. jejuni infection. Future work involves determining responses in colonoids derived from volunteers with specific characteristics to create unique host‐bacterial infection models. Support or Funding Information Supported by funding from Department of Defense W81XWH‐18‐1‐0074

AbstractBackgroundAutonomic dysfunction can be present in patients with idiopathic and diabetic gastroparesis. The role of autonomic dysfunction relating to gastric emptying and upper gastrointestinal symptoms in patients with gastroparesis and chronic unexplained nausea and vomiting (CUNV) remains unclear. The aim of our study is to evaluate autonomic function in patients with gastroparesis and CUNV with respect to etiology, gastric emptying and symptom severity.MethodsWe studied 242 patients with chronic gastroparetic symptoms recruited at eight centers. All patients had a gastric emptying scintigraphy within 6 months of the study. Symptom severity was assessed using the gastroparesis cardinal symptom index. Autonomic function testing was performed at baseline enrollment using the ANX 3.0 autonomic monitoring system which measures heart rate variability and respiratory activity measurements.Key ResultsLow sympathetic response to challenge (Valsalva or standing) was the most common abnormality seen impacting 89% diabetic and 74% idiopathic patients. Diabetics compared to idiopathics, exhibited greater global hypofunction with sympathetic (OR = 4.7, 95% CI 2.2‐10.3; P < .001) and parasympathetic (OR = 7.2, 95% CI 3.4‐15.0; P < .001) dysfunction. Patients with delayed gastric emptying were more likely to have paradoxic parasympathetic excessive during sympathetic challenge [(Valsalva or standing) 40% vs. 26%, P = .05]. Patients with more severe symptoms exhibited greater parasympathetic dysfunction compared to those with mild‐moderate symptoms: resting sympathovagal balance [LFa/RFa 1.8 (1.0‐3.1) vs. 1.2 (0.6‐2.3), P = .006)] and standing parasympathetic activity [0.4 (0.1‐0.8) vs. 0.6 (0.2‐1.7); P = .03].ConclusionsAutonomic dysfunction was common in patients with gastroparesis and CUNV. Parasympathetic dysfunction was associated with delayed gastric emptying and more severe upper gastrointestinal symptoms. Conversely, sympathetic hypofunction was associated with milder symptoms.InferencesGastroparesis and CUNV may be a manifestation of GI autonomic dysfunction or imbalance, such that sympathetic dysfunction occurs early on in the manifestation of chronic upper GI symptoms, while parasympathetic dysfunction results in more severe symptoms and delayed gastric emptying.

OBJECTIVES: Diabetic gastroparesis (Gp) occurs more often in type 1 diabetes mellitus (T1DM) than in type 2 diabetes mellitus (T2DM). Other diabetic end-organ complications include peripheral neuropathy, nephropathy, and retinopathy (together termed triopathy ). This study determines the prevalence of diabetic complications (retinopathy, nephropathy, and peripheral neuropathy) in diabetic patients with symptoms of Gp, assessing the differences between T1DM and T2DM and delayed and normal gastric emptying (GE). METHODS: Diabetic patients with symptoms of Gp underwent history and physical examination, GE scintigraphy, electrogastrography with water load, autonomic function testing, and questionnaires assessing symptoms and peripheral neuropathy. RESULTS: One hundred thirty-three diabetic patients with symptoms of Gp were studied: 59 with T1DM and 74 with T2DM and 103 with delayed GE and 30 without delayed GE. The presence of retinopathy (37% vs 24%; P = 0.13), nephropathy (19% vs 11%; P = 0.22), and peripheral neuropathy (53% vs 39%; P = 0.16) was not significantly higher in T1DM than in T2DM; however, triopathies (all 3 complications together) were seen in 10% of T1DM and 3% of T2DM ( P = 0.04). Diabetic patients with delayed GE had increased prevalence of retinopathy (36% vs 10%; P = 0.006) and number of diabetic complications (1.0 vs 0.5; P = 0.009); however, 39% of diabetic patients with delayed GE did not have any diabetic complications. DISCUSSION: In diabetic patients with symptoms of Gp, delayed GE was associated with the presence of retinopathy and the total number of diabetic complications. Only 10% of patients with T1DM and 3% of those with T2DM had triopathy of complications, and 39% of diabetic patients with Gp did not have any diabetic complications. Thus, the presence of diabetic complications should raise awareness for Gp in either T1DM or T2DM; however, diabetic Gp frequently occurs without other diabetic complications.