Introduction

Esophageal achalasia (EA) is a rare functional disorder caused by neuromuscular dysfunction at the esophagogastric junction, characterized by impaired diastole of the lower esophageal sphincter (LES) and loss of distal esophageal peristaltic function. The etiology of the disease is still unknown, and the current etiological hypothesis is that the loss of esophageal neurons due to ganglion inflammation caused by an abnormal immune response to infection in a genetically susceptible background affects the function of the muscles innervated by it. Previous studies have suggested that genetic susceptibility may be strongly associated with the onset of achalasia. Based on this platform, our group is one of the largest EA clinical centers in China, and we used whole-exome sequencing and technology validation to screen out a new genetic mutation locus FAM129C from 22 Trios core families with achalasia. This paper intends to investigate whether Fam129c mutant mice have an EA phenotype, and to investigate the pathogenic mechanism of this mutation with multi-omics. This study will provide a new basis for uncovering the genetic cause of EA, a new mouse model for basic EA research, and potential therapeutic targets for the treatment of EA.

Aims & Methods

The new pathogenic genes inherited from the family line of achalasia in the Han population were screened, and the mutant mice were constructed by CRSPR/Cas9 technology. To verify whether the mutant mice have the cardia apopause phenotype, and to explore the specific mechanism of disease in mutant mice through multi-omics joint analysis and in vivo and in vivo validation experiments.

Results

Fam129c mutant mice were successfully constructed, and it was confirmed that they had EA phenotype from anatomical to functional levels. Immunofluorescence results showed that FAM129C was localized in B cells. Abnormal activation of plasma cells in mutant mice, specific deletion of B cells can partially alleviate the achalasia phenotype of mutant mice. Transcriptome sequencing and single-cell sequencing results revealed a significant enrichment of B cell subsets, and differential genes elevated in mutant mice mainly included immunoglobulins. Single-cell BCR repertoire sequencing analysis showed that mutant mice had increased V-J pairing, increased clone number and clone types, and increased immunoglobulin secretion. Further analysis of proteomics results found that the anti-inhibitory neuron autoantibody GABA-AR was elevated in serum and in situ tissue of mutant mice, and the results of ELISA verification in mice were consistent with it. IvIgG treatment of mutant mice in vitro and in vivo can partially alleviate the EA phenotype.

Conclusion

A new potential mutated genes FAM129C in achalasia were identified, and Fam129c mutant mice conforming to the population mutation pattern were established. Fam129c mutant mice have an EA phenotype, and their anatomical features, pathological changes in the esophageal intermuscular plexus and diastolic function of the lower esophagus are consistent with EA patients. Fam129c mutant mice are pathogenic through a B-cell-dependent pathway. Abnormal activation of plasma cells in mutant mice, increased secretion of immunoglobulins, especially anti-neuronal autoantibody GABA-AR, and neuromuscular dysfunction after specific injury of esophageal inhibitory intermuscular neurons, eventually leading to the onset of achalasia.

Introduction

Endoscopic submucosal dissection (ESD) is one of the effective treatments for superficial non-ampullary duodenal epithelial tumors (SNADETs). To avoid postoperative perforation and bleeding, our hospital had introduced D-LECS with ESD for SNADETs. From 2021, D-LECS was divided into D-LECS with FTR, which is a full-thickness resection, and D-LECS with ESD, which is intended for post-ESD reinforcement (1). D-LECS with ESD is considered to be a therapeutic procedure that not only ensures safety but also contributes to the improvement of endoscopic skills.

Aims & Methods

The aim of this study is to evaluate the impact of D-LECS with ESD on the skill development of endoscopists. The subjects were 17 cases in which D-LECS was performed among 79cases diagnosed with SNADA by biopsy at our hospital between January 2000 and October 2023. 5 cases in which D-LECS with ESD was performed. The ESD assessment score was defined using a ladder chart based on muscle layer exposure, perforation, ESD time, and pathological examination results. The scoring for points each item is as follows. (Exposure of muscle layer none: 0, <3mm: 1pt, 3~10mm: 2pt, 10mm<: 3pt, Perforation of the muscle none: 0, <3mm: 1pt, 3~10mm: 5pt, 10mm<: 10pt, ESD time <60min.: 0pt, 60~120min.: 5pt, 120min.<: 10pt, Pathological examination result pR0: 0pt, cR0+pRX: 5pt, pR1:10pt) Images and videos were used in combination to evaluate the muscle layer.

Results

ESD assessment scores of cases undergoing D-LECS with ESD were presented. The state of exposure of the muscle layer had a median of 5 (1-7), a mean of 4.8, and a standard deviation (SD) of 2.3. The state of perforation of the muscle layer had a median of 0 (0-10), a mean of 2.6, and a SD of 4.3. The median ESD time was 170 minutes, the mean was 146 minutes (41-198). The state of pathological examination had a median of 0 (0-10), a mean of 2.0, and a SD of 4.5. The ESD assessment scores are presented as follows: (Exposure of muscle layer, Perforation of muscles layer, ESD, Pathological examination result). The results of each score are as shown on the right, Case1: (7, 0,10, 0), Case2: (5,10, 10, 0), Case3: (5, 0, 10, 0), Case4: (6, 3, 10, 10), Case5: (1, 0, 1, 0). Although the number of cases was small, the time required for ESD was shortened and the degree of perforation tended to improve with experience. 2 examples revealed some important points. Case 1: This case had many blood vessels, and the ST hood was needed because it is difficult to perform ESD. When the endoscope was extended, perforation occurred. Once the perforation was made, it was difficult to move the patient to other bed, so the muscle layer was sutured with OTSC, and a PGA sheet. Case 2: The endoscopic strategy and device were improved, and the lesion could be dissected within 1 hour. ESD was performed with Dual Knife J 1.5. Pulling the lesion with an S-O clip. The RDI function was used during hemostasis to avoid the burn effect on the muscle layer.

Conclusion

D-LECS with ESD will lead to improvements of duodenal ESD techniques. But to ensure safety, laparoscopic closure is preferable until reliable suturing via endoscope becomes possible.

References

(1) Souya Nunobe, Motonari Ri, Naoki Hiki et al: Safety and Feasibility of laparoscopic and endoscopic cooperative surgery for duodenal neoplasm: a retrospective multicenter study. Endoscopy 53: 1065-1068, 2021

Introduction

Liver fibrosis, characterized by diffuse fibrous deposition and distortion of hepatic vascular architecture in histology, is an intermediate step in the development of cirrhosis resulting from chronic liver disease. Liver sinosoidal endothelial cells (LSECs) are critical gatekeepers of sinusoidal homeostasis. Following injury, LSECs are the first to be affected and initiate a cascade of reactions within the sinusoidal, which contribute to the development and progression of liver fibrosis and portal hypertension. Our preliminary studies have shown that hepatic cyclooxygenase-2 (COX-2) expression was correlated with the severity of inflammation and fibrosis, and its inhibitor could ameliorate intrahepatic fibrosis.

Aims & Methods

The present study aimed to investigate the role of COX-2 on LSECs function and its underlying molecular mechanism.

Results

Ptgs2 deficiency attenuates liver fibrosis
In both inducible and non-inducible mouse models, Ptgs2 knockdown resulted in a reduction in collagen deposition. TAA- or DDC-induced fibrosis-related protein expression (collagen I and α-SMA) was significantly reduced by Ptgs2 deletion.
Ptgs2 deficiency alleviates hepatic inflammatory injury
RNA-seq analysis indicated that down-regulated DEGs by Ptgs2 knockdown were enriched in leukocyte chemotaxis, leukocyte migration, and leukocyte cell-cell adhesion. Consistently, significantly decreased numbers of infiltrating neutrophils and macrophages were observed in Ptgs2-deficient mice. In addition to inflammatory cell infiltration, liver function was also improved suggested by significant decreases in serum levels of ALT and AST.
COX-2 expression is elevated in LSECs in reponse to liver injury
D-IF showed that COX-2 expression was elevated in injury liver tissues. The immunofluorescence for COX-2 in TAA-A and TAA-C groups was also co-stained with LYVE1, a LSEC-specific maker. RNA-seq data from the GEO database further confimed the COX-2 overexpression in murine LSECs isolated from various fibrotic models. In vivo study, COX-2-mPGES-1-EP2 pathway could be up-regulated by pro-inflammatory factor TNFα.
COX-2-PGE2-EP2 promotes leukocyte recruitment through NF-κB pathway
RNA-seq analysis found that inhibition of COX-2 supressed TNF-induced gene transcription which were enriched in leukocyte recruitment, TNFα pathway, and NF-κB pathway. In HHSEC, TNFα activated NF-κB signaling pathway and increased expression levels of chemokines (CXCL1 and CCL2) and adhesion molecules (ICAM1 and VCAM1). This TNFα-induced effect could be potently suppressed by COX-2 inhibitor (Celecoxib), mPGES-1 inhibitor (MF63) or EP2 receptor antagonist (TG4-155). PGE2 also activated NF-κB pathway, which was abolished after EP2 blockage.
COX-2 inhibitors attenuate fibrosis and liver injury
Both Celecoxib and Etoricoxib (both were selective COX-2 inhibitors) were able to decrease collagen deposition, fibrosis-related protein expression, inflammatory cell infiltration and serum transaminase levels in TAA or DDC models.

Conclusion

COX-2 expression in LSECs is up-regulated in liver fibrosis. Upon inflammatory stimulation, COX-2-PGE2-EP2 axis increases chemokines and adhesion molecules through the synergistic activation of NF-κB signaling pathway, leading to leukocyte recruitment and hepatic inflammation. Inhibition of COX-2 by Celecoxib or Etoricoxib attenuate fibrosis and liver injury. The present study sheds light on the regulation of LSECs on leukocyte recruitment through a cyclooxygenase-2-dependent mechanism. Taken together, our results suggest COX-2 as a potential therapeutic target in the treatment of liver fibrosis.

Disclosure

The authors disclose no conflicts.

Introduction

Perforation is one of the most serious complications in endoscopic retrograde cholangiopancreatography (ERCP). Conventional non-surgical treatment, including intravenous antibiotics with nothing per oral(NPO) and traditional bile drainage (plastic ERBD, PTGBD, PTBD), is generally approved treatment for the ERCP-related Stapfer’s type ll perforation (perivaterian type). The biliary covered metal stent placement has recently been reported to have favorable outcomes in ERCP-related type ll perforation.

Aims & Methods

We aimed to compare the outcomes of conventional treatment and biliary covered metal stent insertion in patients with Stapfer’s type II perforation. ​The medical records of patients who underwent ERCP at Kyungpook National University Hospital in Daegu from 2011 through 2022 were retrospectively reviewed.

Results

8,402 ERCP procedures were conducted in our hospital. 66 ERCP-related perforations (0.78%) were identified with 47 patients (71.2%) had type II perforations. 34 patients received conventional treatments, 13 patients received biliary covered metal stent insertions. There were also no significant differences in clinical success rate (97.1% vs 92.3%, p=0.481), hospital stay (15.53±11.35 vs. 9.46±5.97 days, p=0.227), and NPO times (3.79±3.00 vs. 5.42±3.40 days, p=0.127). The conventional bile drainage group took shorter time for ERCP than SEMS group (12.62±6.315 vs. 18.46±11.16 minutes, p=0.028).

Conclusion

The biliary covered metal stent insertion did not improve patient’ outcomes when compared with the conventional treatment method in ERCP-related type II perforations. Conventional treatment is sufficient for management of ERCP-related type II small sized perforations.

Introduction

Inflammatory bowel disease (IBD), encompassing Crohn’s disease (CD) and ulcerative colitis (UC), often manifests around the fertile age. It is known that disease activity affects pregnancy outcomes. However, little is known about how IBD affects pregnancy outcomes and vice versa. Both IBD and pregnancy complications, particularly those in which placental insufficiency is implicated, are accompanied by oxidative stress. Oxidative stress is defined as an imbalance between reactive oxygen species (ROS) production and antioxidant availability. Free thiols (FT) are circulating biomarkers of oxidative stress since they readily scavenge ROS. Previous research has shown reduced levels of FT in both IBD¹ and pregnancy complications². Classical biomarkers of placental insufficiency include soluble FMS-like tyrosine kinase-1 (sFlt-1) and placental growth factor (PLGF), and their ratio is reflective of maternal pregnancy complications². However, the behavior of these biomarkers in pregnant women with IBD remains unclear.

Aims & Methods

In this study, we aimed to investigate the relationship between FT, sFlt-1, and PLGF and the occurrence of pregnancy complications in patients with IBD, and to assess their dynamics before, during, and shortly after pregnancy.
In this retrospective clinical pilot study, we investigated a cohort of pregnant women with IBD and healthy pregnant controls who were pregnant between 2000 and 2023. Serum samples taken before (IBD n=41), during (IBD n=33, controls n=14), and after (IBD n=25) pregnancy were analyzed for PLGF, sFlt-1 and FT levels. FTs were measured using a colorimetric detection method. sFlt-1 and PLGF were measured using an immunofluorescent assay. Extensive clinical data were collected, including data about pregnancy complications such as pre-eclampsia, fetal growth restriction, and gestational diabetes, and IBD disease parameters, including fecal calprotectin and clinical disease scores.

Results

In total, 40 patients (n=32 CD, n=7 UC, and n=1 IBD-U) with 47 pregnancies and 14 non-IBD control pregnancies were included. Comparing patients with IBD to controls during pregnancy, serum FT levels were significantly reduced in patients, even after adjustment for gestational age (p=0.043). FT levels significantly decreased during pregnancy compared to preconception (p=0.001), suggesting increased systemic oxidative stress during pregnancy. The sFlt-1/PLGF ratio was significantly higher in IBD compared to controls (p=0.011), which was partially due to a difference in gestational age (median (IQR): controls 32.6 weeks (29.4-33.1) vs. 15.4 weeks (7.0-22.7) in IBD). In total, 32 patients (56%) and 3 controls (21%) developed pregnancy complications. The sFlt-1/PLGF ratio was higher in those developing maternal complications, almost reaching significance (p=0.053), while FT showed a trend towards decreased levels in those developing maternal complications (mean (SD): 368.9 µM (38.7) without vs. 349.1 µM (21.9) with maternal pregnancy complications, p=0.082).

Conclusion

This study provides evidence for increased systemic oxidative stress (reflected by lower systemic FT levels) during pregnancy in IBD, when compared to both healthy controls and their own pre-conceptional levels. These results suggest a difference in redox status between pregnant women with IBD compared to those without. Further research is necessary to substantiate these findings and evaluate the utility of these biomarkers in predicting pregnancy complications and guiding clinical decision making in IBD pregnancy.

References

1. Bourgonje AR, Gabriëls RY, de Borst MH, Bulthuis MLC, Faber KN, van Goor H, Dijkstra G. Serum Free Thiols Are Superior to Fecal Calprotectin in Reflecting Endoscopic Disease Activity in Inflammatory Bowel Disease. Antioxidants (Basel). 2019 Sep 1;8(9):351. doi: 10.3390/antiox8090351.
2. Schoots MH, Bourgonje MF, Bourgonje AR, Prins JR, van Hoorn EGM, Abdulle AE, Muller Kobold AC, van der Heide M, Hillebrands JL, van Goor H, Gordijn SJ. Oxidative stress biomarkers in fetal growth restriction with and without preeclampsia. Placenta. 2021 Nov;115:87-96. doi: 10.1016/j.placenta.2021.09.013.

Introduction

Pfizer's Institute for Translational Equitable Medicine and our Chief Medical Officer, Aida Habtezion, MD, hosted an IBD Health Equity Summit at DDW 2023 in Chicago, Illinois on May 5, 2023 with a group of 22 key opinion leaders in the gastroenterology field. The purpose of the Summit was to uncover health equity research gaps and quality of care gaps that will enable Pfizer and our partners to determine future directions in IBD research and contribute to achieving healthcare equity by providing opportunities to improve patient outcomes, as well as address practice-based disparities.

Aims & Methods

A full day Summit was designed and developed, bringing together thought leaders and experts (representing research, HCPs and patients) to discuss health disparity gaps and possible solutions. The day was split into two parts: a morning session focused on research, with IBD researchers presenting different facets of the research construct with follow-on discussion with the audience. The afternoon was dedicated to clinical care and also included IBD experts addressing Health Equity concerns and opportunities. A follow-on summary report was developed, distributed and serves as the basis for current and future strategies aimed at closing disparity gaps among IBD patients.

Results

Several overarching themes emerged, highlighting IBD as a global affliction, and challenges that transcend geographical borders. Addressing these challenges will require solutions tailored to local and regional contexts that consider factors such as race, ethnicity, socio-economic status, culture, diets, and sexual orientations. It was also recognized that many of the challenges posed by IBD are not unique to this disease, and that valuable insights can be gained by examining other complex disease states to improve the quality of care and address health disparities in patients with IBD.

Conclusion

IBD Research
Key gaps include low clinical trial participation of patients from diverse communities due to lack of healthcare literacy, low historical representation, bias in recruitment, lack of diversity in providers.
Strategies to address these gaps include improving health literacy, workforce development and educating providers, adequate compensation for time, flexibility for participating in trials, and incorporation of digital health tools for follow-ups.
Clinical Care
There are groups of patients in several regions that lack any care, and addressing healthcare disparities in IBD care for these patients cannot be done online but will need to be done on a local and regional level on the ground.
When addressing health inequities, the heterogeneities within underserved populations like Black and Hispanic patients need to be considered.
Gaps in data on the effectiveness of therapies in specific patient populations (ie, response to anti-TNFs in black patients and patients with comorbidities).
Sponsorship is lacking to establish efforts in underserved communities on a global level; There is an opportunity to provide connection for minority physicians to resources.
By identifying gaps and potential strategies to close them, the participants have provided Pfizer and our partners a roadmap to address health disparities among IBD patients worldwide. These data and information have served as the basis for a go-forward plan to support and invest in change initiatives to close gaps. Our desire is to enlist additional partners and collaborators through a Call to Action to identify solutions at the local level that can accelerate improved IBD care and outcomes for some of the most vulnerable populations in the world.

Disclosure

Employee of Pfizer Inc