An esophagus cell atlas reveals dynamic rewiring during active eosinophilic esophagitis and remission.

Coordinated cell interactions within the esophagus maintain homeostasis, and disruption can lead to eosinophilic esophagitis (EoE), a chronic inflammatory disease with poorly understood pathogenesis. We profile 421,312 individual cells from the esophageal mucosa of 7 healthy and 15 EoE participants, revealing 60 cell subsets and functional alterations in cell states, compositions, and interactions that highlight previously unclear features of EoE. Active disease displays enrichment of ALOX15+ macrophages, PRDM16+ dendritic cells expressing the EoE risk gene ATP10A, and cycling mast cells, with concomitant reduction of TH17 cells. Ligand-receptor expression uncovers eosinophil recruitment programs, increased fibroblast interactions in disease, and IL-9+IL-4+IL-13+ TH2 and endothelial cells as potential mast cell interactors. Resolution of inflammation-associated signatures includes mast and CD4+ TRM cell contraction and cell type-specific downregulation of eosinophil chemoattractant, growth, and survival factors. These cellular alterations in EoE and remission advance our understanding of eosinophilic inflammation and opportunities for therapeutic intervention.

CYP2C19 Loss-of-Function is Associated with Increased Risk of Ischemic Stroke after Transient Ischemic Attack in Intracranial Atherosclerotic Disease.

OBJECTIVES: Intracranial atherosclerotic disease (ICAD) is responsible for 8-10% of acute ischemic strokes, and resistance to antiplatelet therapy is prevalent. CYP2C19 gene loss-of-function (up to 45% of patients) causes clopidogrel resistance. For patients with asymptomatic ICAD and ICAD characterized by transient ischemic attack (TIA), this study measures the effect of CYP2C19 loss-of-function on ischemic stroke risk during clopidogrel therapy. MATERIALS AND METHODS: From a deidentified database of medical records, patients were selected with ICD-9/10 code for ICAD, availability of CYP2C19 genotype, clopidogrel exposure, and established patient care. Dual-antiplatelet therapy patients were included. Patients with prior ischemic stroke, other neurovascular condition, intracranial angioplasty/stenting, or observation time <1 month were excluded. Time-to-event analysis using Cox regression was conducted to model first-time ischemic stroke events based on CYP2C19 loss-of-function allele and adjusted for age, gender, race, length of aspirin, length of concurrent antiplatelet/anticoagulant treatment, diabetes, coagulopathy, hypertension, heart disease, atrial fibrillation, and lipid disorder. Subset analyses were performed for asymptomatic and post-TIA subtypes of ICAD. RESULTS: A total of 337 patients were included (median age 68, 58% male, 88% Caucasian, 26% CYP2C19 loss-of-function). A total of 161 (47.8%) patients had TIA at time of ICAD diagnosis, while 176 (52.2%) were asymptomatic. First-time ischemic stroke was observed among 20 (12.4%) post-TIA ICAD patients and 17 (9.7%) asymptomatic ICAD patients. Median observation time was 2.82 [IQR 1.13-5.17] years. CYP2C19 loss-of-function allele was associated with ischemic stroke event (HR 2.2, 95% CI 1.1-4.3, p=0.020) after adjustment. Post-TIA ICAD patients had a higher risk of ischemic stroke from CYP2C19 loss-of-function (HR 3.4, 95% CI 1.4-8.2, p=0.006). CONCLUSIONS: CYP2C19 loss-of-function was associated with 3-fold increased risk of first-time ischemic stroke for ICAD patients treated with clopidogrel after TIA. This effect was not observed for asymptomatic ICAD. CYP2C19-guided antiplatelet selection may improve stroke prevention in ICAD after TIA.

High-throughput in vivo mapping of RNA accessible interfaces to identify functional sRNA binding sites.

Herein we introduce a high-throughput method, INTERFACE, to reveal the capacity of contiguous RNA nucleotides to establish in vivo intermolecular RNA interactions for the purpose of functional characterization of intracellular RNA. INTERFACE enables simultaneous accessibility interrogation of an unlimited number of regions by coupling regional hybridization detection to transcription elongation outputs measurable by RNA-seq. We profile over 900 RNA interfaces in 71 validated, but largely mechanistically under-characterized, Escherichia coli sRNAs in the presence and absence of a global regulator, Hfq, and find that two-thirds of tested sRNAs feature Hfq-dependent regions. Further, we identify in vivo hybridization patterns that hallmark functional regions to uncover mRNA targets. In this way, we biochemically validate 25 mRNA targets, many of which are not captured by typically tested, top-ranked computational predictions. We additionally discover direct mRNA binding activity within the GlmY terminator, highlighting the information value of high-throughput RNA accessibility data.

Integrin αM activation and upregulation on esophageal eosinophils and periostin-mediated eosinophil survival in eosinophilic esophagitis.

Eosinophilic esophagitis (EoE) is an increasingly recognized allergic disease associated with dysphagia and esophageal fibrosis. We aimed to determine expression patterns of specific eosinophil integrins that promote eosinophilic infiltration of the esophageal epithelium, and to determine how key EoE-related cytokines influence eosinophil activation and survival. Esophageal and peripheral eosinophils were isolated from 20 adult subjects with EoE for immunophenotyping and integrin profiling using multicolor flow cytometry and immunohistochemistry. Expression signatures of eosinophil integrins were further assessed by immunohistochemistry using serial sections of esophageal biopsy specimens. Purified eosinophils were used to assess the effect of EoE-relevant cytokines and recombinant periostin on expression of known eosinophil integrins and eosinophil survival and activation. We found that resting eosinophils express high levels of the ß2-pairing integrins αL and αM, and lower levels of α4, α6 and α4ß7. The migration of peripheral eosinophils to the esophagus is characterized by the specific induction of αM, and a significant increase in the proportion of αM in high-activity conformation. Periostin, a secreted extracellular matrix protein that is significantly overexpressed in EoE, enhances eosinophil survival, and this effect is mediated by αM interaction. Integrin αM is a specific marker of activated tissue eosinophils in EoE, and promotes eosinophil survival through interactions with periostin. The ability of αMß2 to mediate eosinophil tissue residency via periostin represents a key mechanism for disease development and a potential therapeutic target in EoE.