Identification of a dihydropyridine scaffold that blocks ryanodine receptors.

Ryanodine receptors (RyRs) are large, intracellular ion channels that control Ca2+ release from the sarco/endoplasmic reticulum. Dysregulation of RyRs in skeletal muscle, heart, and brain has been implicated in various muscle pathologies, arrhythmia, heart failure, and Alzheimer's disease. Therefore, there is considerable interest in therapeutically targeting RyRs to normalize Ca2+ homeostasis in scenarios involving RyR dysfunction. Here, a simple invertebrate screening platform was used to discover new chemotypes targeting RyRs. The approach measured Ca2+ signals evoked by cyclic adenosine 5'-diphosphate ribose, a second messenger that sensitizes RyRs. From a 1,534-compound screen, FLI-06 (currently described as a Notch "inhibitor") was identified as a potent blocker of RyR activity. Two closely related tyrosine kinase inhibitors that stimulate and inhibit Ca2+ release through RyRs were also resolved. Therefore, this simple screen yielded RyR scaffolds tractable for development and revealed an unexpected linkage between RyRs and trafficking events in the early secretory pathway.

H+/K+ATPase Expression in the Larynx of Laryngopharyngeal Reflux and Laryngeal Cancer Patients.

OBJECTIVES: The gastric H+/K+ ATPase proton pump has previously been shown to be expressed in the human larynx, however its contribution to laryngopharyngeal reflux (LPR) signs, symptoms and associated diseases such as laryngeal cancer is unknown. Proton pump expression in the larynx of patients with LPR and laryngeal cancer was investigated herein. A human hypopharyngeal cell line expressing the proton pump was generated to investigate its effects. STUDY DESIGN: In-vitro translational. METHODS: Laryngeal biopsies were obtained from three LPR and eight LSCC patients. ATP4A, ATP4B and HRPT1 were assayed via qPCR. Human hypopharyngeal FaDu cell lines stably expressing proton pump were created using lentiviral transduction and examined via transmission electron microscopy and qPCR for genes associated with inflammation or laryngeal cancer. RESULTS: Expression of ATP4A and ATP4B was detected in 3/3 LPR, 4/8 LSCC-tumor and 3/8 LSCC-adjacent specimens. Expression of ATP4A and ATP4B in FaDu elicited mitochondrial damage and expression of IL1B, PTGS2, and TNFA (P < .0001); expression of ATP4B alone did not. CONCLUSIONS: Gastric proton pump subunits are expressed in the larynx of LPR and LSCC patients. Mitochondrial damage and changes in gene expression observed in cells expressing the full proton pump, absent in those expressing a single subunit, suggest that acid secretion by functional proton pumps expressed in upper airway mucosa may elicit local cell and molecular changes associated with inflammation and cancer. LEVEL OF EVIDENCE: NA Laryngoscope, 131:130-135, 2021.

GATA4 Is Required for Budding Morphogenesis of Posterior Foregut Endoderm in a Model of Human Stomach Development.

Three-dimensional gastrointestinal organoid culture systems provide innovative and tractable models to investigate fundamental developmental biology questions using human cells. The goal of this study was to explore the role of the zinc-finger containing transcription factor GATA4 in gastric development using an organoid-based model of human stomach development. Given GATA4's vital role in the developing mouse gastrointestinal tract, we hypothesized that GATA4 plays an essential role in human stomach development. We generated a human induced pluripotent stem cell (hiPSC) line stably expressing an shRNA targeted against GATA4 (G4KD-hiPSCs) and used an established protocol for the directed differentiation of hiPSCs into stomach organoids. This in vitro model system, informed by studies in multiple non-human model systems, recapitulates the fundamental processes of stomach development, including foregut endoderm patterning, specification, and subsequent tissue morphogenesis and growth, to produce three-dimensional fundic or antral organoids containing functional gastric epithelial cell types. We confirmed that GATA4 depletion did not disrupt hiPSC differentiation to definitive endoderm (DE). However, when G4KD-hiPSC-derived DE cells were directed to differentiate toward budding SOX2+, HNF1B+ posterior foregut spheroids, we observed a striking decrease in the emergence of cell aggregates, with little to no spheroid formation and budding by GATA4-depleted hiPSCs. In contrast, control hiPSC-derived DE cells, expressing GATA4, formed aggregates and budded into spheroids as expected. These data support an essential role for GATA4 during the earliest stages of human stomach development.

Cumulative incidence of second intestinal resection in Crohn's disease: a systematic review and meta-analysis of population-based studies.

OBJECTIVES: Approximately 50% of Crohn's disease patients undergo an intestinal resection within 10 years of diagnosis. The risk of second surgery in Crohn's disease and the influence of time are not well characterized. We performed a systematic review and meta-analysis to establish the risk of second abdominal surgery in patients with Crohn's disease among patients who had a previous surgery. METHODS: We searched Medline, EMBASE, PubMed (March 2014), and conference proceedings for terms related to Crohn's disease and intestinal surgery. We included population-based articles (n=11) and an abstract (n=1) reporting surgical risk for the overall study period and for 5 and 10 years after the first surgery for Crohn's disease. We stratified studies by year (start year before vs. after 1980) to explore the role of time. RESULTS: For all population-based studies, the overall risk of second surgery was 28.7% (95% confidence interval (CI): 22.6-36.6%). The 5-year risk of second surgery was 24.2% (95% CI: 22.3-26.4%). The 10-year risk of second surgery was 35.0% (95% CI: 31.8-38.6%). A significant difference in the 10-year risk of second surgery was observed over time such that studies conducted after 1980 had a lower risk of second surgery (33.2%; 95% CI: 31.2-35.4%) compared with those that started before 1980 (44.6%; 95% CI: 37.7-52.7%). CONCLUSIONS: Approximately one-quarter of Crohn's disease patients who have a first surgery also have a second, and the majority of these surgeries occur within 5 years of the first surgery. The 10-year risk of second surgery is significantly decreasing over time.

Risk of surgery for inflammatory bowel diseases has decreased over time: a systematic review and meta-analysis of population-based studies.

BACKGROUND & AIMS: The inflammatory bowel diseases (IBDs) are chronic diseases that often require surgery. However, the risk of requirement of surgery over time has not been well characterized. We performed a systematic review and meta-analysis to establish the cumulative risk of surgery among patients with IBD and evaluated how this risk has changed over time. METHODS: We searched Medline, EMBASE, PubMed, and conference proceedings (2009-2012) on May 8, 2013, for terms related to IBD and intestinal surgery. Two reviewers screened 8338 unique citations to identify 486 for full-text review. The analysis included population-based studies published as articles (n = 26) and abstracts (n = 4) that reported risks of surgery at 1, 5, or 10 years after a diagnosis of Crohn's disease and/or ulcerative colitis. The trend in risk of surgery over time was analyzed by meta-regression using mixed-effect models. RESULTS: Based on all population-based studies, the risk of surgery 1, 5, and 10 years after diagnosis of Crohn's disease was 16.3% (95% confidence interval [CI], 11.4%-23.2%), 33.3% (95% CI, 26.3%-42.1%), and 46.6% (95% CI, 37.7%-57.7%), respectively. The risk of surgery 1, 5, and 10 years after diagnosis of ulcerative colitis was 4.9% (95% CI, 3.8%-6.3%), 11.6% (95% CI, 9.3%-14.4%), and 15.6% (95% CI, 12.5%-19.6%), respectively. The risk of surgery 1, 5, and 10 years after diagnosis of Crohn's disease and 1 and 10 years after diagnosis of ulcerative colitis has decreased significantly over the past 6 decades (P < .05). CONCLUSIONS: Based on systematic review and meta-analysis of population-based studies, the risk of intestinal surgery among patients with IBD has decreased over the past 6 decades.