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BACKGROUND: IL-33 is a type 2 inflammatory cytokine that is elevated in the esophageal epithelium of EoE subjects. We previously developed a mouse model of EoE dependent on constitutive overexpression of IL-33 from the esophageal epithelium (EoE33). OBJECTIVE: Our objective was to develop an inducible, IL-33-dependent model of EoE and examine induction of EoE-associated pathology. METHODS: We utilized a tetracycline-inducible system to express IL-33 in the esophagus by generating two transgenic mice. The first (iSophagus) expresses a reverse tetracycline transactivator (rtTA) from the esophageal epithelium. The second (TRE33) features a tetracycline-response element driving expression of IL-33. When crossed, these mice generate an inducible model of EoE (iEoE33). Mice were administered doxycycline-infused chow for up to 2 weeks. Cytokines were assessed by ELISA or bead-based multiplex. T cells were assessed by flow cytometry. Pathology was assessed by histology and immunohistochemistry for IL-33, eosinophil peroxidase, CD4, and Ki-67. iEoE33 was treated with steroids and crossed with IL-13-/- mice. For detailed Methods, please see the Methods section in this article's Online Repository at www.jacionline.org. RESULTS: Doxycycline-treated iEoE33 mice demonstrated expression of IL-33 in the esophageal epithelium, and esophageal pathology including eosinophilia, CD4+ cell infiltrate, basal zone hyperplasia, and dilated intercellular spaces. These findings became pronounced on day 7 of induction, were accompanied by weight loss and esophageal thickening, and were steroid responsive and IL-13 dependent. CONCLUSION: Inducible IL-33 expression in the esophageal epithelium elicited features pathognomonic of EoE. iEoE33 enables investigation of EoE disease mechanisms as well as initiation, progression, and resolution.

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Head and neck squamous cell carcinoma (HNSCC) is the sixth leading cause of cancer worldwide according to GLOBOCAN estimates from 2022. Current therapy options for recurrent or metastatic disease are limited to conventional cytotoxic chemotherapy and immunotherapy, with few targeted therapy options readily available. Recent single-cell transcriptomic analyses identified TGF-ß signaling as an important mediator of functional interplays between cancer-associated fibroblasts and a subset of mesenchymal cancer cells. This signaling was shown to drive invasiveness, treatment resistance, and immune evasion. These data provide renewed interest in the TGF-ß pathway as an alternative therapeutic target, prompting a critical review of previous clinical data which suggest a lack of benefit from TGF-ß inhibitors. While preclinical data have demonstrated the great anti-tumorigenic potential of TGF-ß inhibitors, the underwhelming results of ongoing and completed clinical trials highlight the difficulty actualizing these benefits into clinical practice. This topical review will discuss the relevant preclinical and clinical findings for TGF-ß inhibitors in HNSCC and will explore the potential role of patient stratification in the development of this therapeutic strategy.

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During tumor development, promoter CpG islands that are normally silenced by Polycomb repressive complexes (PRCs) become DNA-hypermethylated. The molecular mechanism by which de novo DNA methyltransferase(s) [DNMT(s)] catalyze CpG methylation at PRC-regulated regions remains unclear. Here, we report a cryo-electron microscopy structure of the DNMT3A long isoform (DNMT3A1) amino-terminal region in complex with a nucleosome carrying PRC1-mediated histone H2A lysine-119 monoubiquitination (H2AK119Ub). We identify regions within the DNMT3A1 amino terminus that bind H2AK119Ub and the nucleosome acidic patch. This bidentate interaction is required for effective DNMT3A1 engagement with H2AK119Ub-modified chromatin in cells. Further, aberrant redistribution of DNMT3A1 to Polycomb target genes recapitulates the cancer-associated DNA hypermethylation signature and inhibits their transcriptional activation during cell differentiation. This effect is rescued by disruption of the DNMT3A1-acidic patch interaction. Together, our analyses reveal a binding interface critical for mediating promoter CpG island DNA hypermethylation, a major molecular hallmark of cancer.

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BACKGROUND: Catheter ablation (CA) improves clinical outcomes in patients with atrial fibrillation (AF) and heart failure (HF) with reduced ejection fraction (HFrEF). We aimed to evaluate the impact of CA on clinical and quality-of-life outcomes across HF subtypes. METHODS: All patients undergoing AF ablation at a tertiary center were enrolled in a prospective registry and included in this study (2013-2021). The primary end point was AF recurrence. Secondary end points included AF-related hospitalizations and quality-of-life outcomes. Patients were categorized according to their HF status: no HF, HFrEF, HF with mildly reduced ejection fraction (HFmrEF), and HF with preserved ejection fraction (HFpEF). RESULTS: 7020 patients were included (80% no HF, 8% HFrEF, 7% HFmrEF, and 5% HFpEF). Over 3 years, the cumulative incidence of AF recurrence after ablation was as follows: HFpEF (53%), HFmrEF (41%), HFrEF (41%), and no HF (34%); P<0.01. Multivariable Cox analyses confirmed these findings using no HF group as reference (HFpEF: hazard ratio, 1.47 [95% CI, 1.21-1.78]; HFmrEF: hazard ratio, 1.23 [95% CI, 1.04-1.45]; and HFrEF: hazard ratio, 1.17 [95% CI, 1.01-1.37]; P<0.05 for all). In all groups, CA resulted in a significant reduction of AF-related hospitalization (mean rate per 1 patient-years [before and after CA]; HFpEF [1.8 versus 0.3], HFmrEF [1.1 versus 0.2], HFrEF [1.1 versus 0.2], and no HF [1 versus 0.1]; P<0.01 for each comparison) and significant improvement in quality of life as measured by both the AF symptom severity score and the AF burden score (P<0.01 for the comparison between baseline and follow-up for each score when tested separately). CONCLUSIONS: AF recurrence rates after CA were higher in patients with HF compared with those without HF, with patients with HFpEF being at the highest risk of recurrence. Nonetheless, CA was associated with a significant reduction in AF symptoms, AF-related hospitalization, and HF symptoms in most patients irrespective of HF subtypes.

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Esophageal squamous cell carcinoma (ESCC) remains the most common malignancy of the esophagus worldwide. Environmental and lifestyle exposures such as alcohol and tobacco have been well defined in the pathogenesis of ESCC, acting in concert with cell intrinsic epigenomic, genomic and transcriptomic changes. However, a variety of non-environmental etiologies including Fanconi anemia, lichen planus, chronic mucocutaneous candidiasis, esophageal epidermoid metaplasia, epidermolysis bullosa, tylosis, esophageal atresia, and achalasia receive minimal attention despite a high risk of ESCC in these diseases. The goal of this review is to promote clinical recognition and suggest a diagnostic framework for earlier detection of ESCC in patients with these rare diseases. In all of the discussed conditions, a change in symptoms should trigger a prompt endoscopic evaluation, and endoscopic surveillance programs with advanced imaging techniques and chromoendoscopy should be considered. Moreover, we leverage the convergence of these diseases on ESCC to identify common mechanisms underlying malignant transformation including aberrant proliferation, mucosal barrier dysfunction, increased inflammation, and genome instability. Herein, we summarize the clinical presentation, pathologic findings, potential screening strategies, and common mechanisms of malignant transformation associated with these rare diseases that drive ESCC.

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BACKGROUND: During pulsed field ablation (PFA), relationships between ablation parameters (contact force [CF], number of burst pulses, impedance decrease, and electrode temperature) and lesion size in beating hearts have not been well validated. METHODS: A 7.5F-catheter with a 3.5-mm ablation electrode and CF sensor (ThermoCool SmartTouch SF-Dual-Energy, Biosense Webster, Inc, Irwindale, CA) was connected to a PFA system (TRUPULSE2, Biosense Webster, Inc). In 11 closed-chest swine, biphasic PFA current was delivered between the ablation electrode and the skin patch at 219 sites in left ventricle and right ventricle using 12, 18, and 24 burst pulses with 4 different levels of CF: (1) low (n=57; CF, 4-15g; median, 10g); (2) moderate (n=60; CF, 16-30g; median, 22.5g); (3) high (n=68; CF, 32-65g; median, 40g); and (4) no electrode contact (n=34), 2 mm away from the endocardium. Swine were euthanized 2 hours after ablation, and lesion size was measured using triphenyl tetrazolium chloride staining. RESULTS: All PFA lesions with electrode-myocardium contact were well demarcated with triphenyl tetrazolium chloride staining, demonstrating (1) pale central zone (contraction band necrosis with minimal coagulation necrosis), (2) dark brown zone (contraction band necrosis with hemorrhage), and (3) hyperstained red zone by triphenyl tetrazolium chloride (unaffected normal myocardium with preserved mitochondrial activity, consistent with reversible zone). Lesion depth increased significantly with increasing CF and the number of PFA burst pulses. An exponential/logarithmic formula combined with CF and the number of PFA burst pulses correlated lesion depth with high accuracy: R=0.809, P<0.0001, ±1.0-mm accuracy in 128 of 163 (79%) lesions, and ±1.5-mm accuracy in 153 of 163 (94%) lesions. Impedance decrease and electrode temperature were poor predictors of lesion size. There were no detectable lesions resulting from ablation without electrode contact. CONCLUSIONS: Acute PFA ventricular lesions demonstrate irreversible and reversible lesion boundaries. Electrode-tissue contact is required for effective lesion formation. Lesion depth increases significantly with increasing CF and PFA burst pulses. A new exponential/logarithmic formula combined with CF and the number of PFA burst pulses correlates lesion depth with high accuracy.

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OBJECTIVE: The development of acquired middle ear cholesteatoma is associated with a single nucleotide polymorphism, 538G>A, in the human adenosine triphosphate-binding cassette transporter C11 (ABCC11) gene, which is a determinant of the earwax morphotype, such as wet- and dry-type earwax; however, the mechanism underlying this association is unclear. We focused on the earwax pH and aimed to elucidate the mechanism between ABCC11 genotypes and acquired middle ear cholesteatoma. STUDY DESIGN: Prospective observational study. SETTING: Single-center, academic hospital. METHODS: We recruited 40 patients with acquired middle ear cholesteatoma who underwent surgery and 115 controls with no history of middle ear cholesteatoma. We assessed the earwax pH and ABCC11 genotypes in all participants. Clinical information was collected from the patients with cholesteatoma. RESULTS: The earwax pH was significantly less acidic in patients with cholesteatoma and those carrying wet earwax genotypes (ABCC11 538G/G or 538G/A) than in the controls and those carrying the dry earwax genotype (ABCC11 538A/A), respectively. Furthermore, earwax pH was significantly positively correlated with high preoperative cholesteatoma stages in the patients with cholesteatoma. CONCLUSION: Our results show that the less acidic earwax pH was significantly related to the development and progression of acquired middle ear cholesteatoma. The less acidic earwax pH may play an important role in the mechanism underlying the association between acquired middle ear cholesteatoma and the ABCC11 gene at site 538.

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Arsenic is widely present in the environment in trivalent and pentavalent forms; long-term arsenic exposure due to environmental pollution has become a problem. Previous reports have shown that 24-h exposure to arsenate (as pentavalent arsenic) potentiates erythropoietin (EPO) production via reactive oxygen species (ROS) in EPO-producing HepG2 cells. However, the effects of long-term arsenate exposure on EPO production remain unclear. In HepG2 cells subcultured for 3 weeks in the presence of arsenate, EPO mRNA levels were lower than those in untreated cells. Levels of ARSENITE METHYLTRANSFERASE mRNA, as well as those of Nuclear factor erythroid 2-related factor 2, glutathione, and superoxide dismutase proteins, were increased compared to untreated cells, but levels of malondialdehyde were not significantly altered. Thus, long-term exposure to arsenate enhances ROS scavenging, suggesting that the ROS-induced accumulation of EPO mRNA is attenuated by arsenate exposure. The induction of EPO accumulation by hypoxia also was attenuated by long-term arsenate exposure, suggesting an impairment in responsivity of EPO production. Furthermore, mRNA levels of SIRTUIN-1, which affects EPO transcription, were potentiated by long-term arsenate exposure. These results suggest that long-term arsenate exposure has multiple, distinct effects on EPO production in vitro.

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The design and synthesis of N-desmethyl and N-methyl destruxin E analogs have been demonstrated. The X-ray single crystal structure of destruxin E (1a) revealed a stable three-dimensional (3D) structure, including a s-cis amide bond at the MeVal-MeAla moiety and two intramolecular hydrogen bonds between NH(ß-Ala) and OC(Ile) and between NH(Ile) and OC(ß-Ala). N-Desmethyl analogs 2a (MeAla â†’ Ala) and 2b (MeVal â†’ Val) were synthesized through macrolactonization similar to our previously reported synthesis of 1a. Conversely, for the synthesis of N-methyl analogs 2c (Ile â†’ MeIle) and 2d (ß-Ala â†’ Meß-Ala), macrolactonization did not proceed; therefore, cyclization precursors 10c and 10d were designed to maintain the intramolecular hydrogen bonds described above during their cyclization. The macrolactamization proceeded despite the presence of a less reactive N-methylamino group at the N-terminus in both cases. Analog 2a, which exhibits multiple conformers in solutions, was inactive at 50 µM, whereas analog 2b, which exhibits a conformation similar to that of 1a in solutions, exhibited morphological changes against osteoclast-like multinuclear cells at 1.6 µM. The activity of the MeIle analog 2c, which cannot take the intramolecular hydrogen bond (Ile)NH•••OC(ß-Ala) in 1a, was markedly diminished compared with that of 1a, and that of the Meß-Ala analog 2d, which cannot take the intramolecular hydrogen bond (ß-Ala)NH•••OC(Ile) in 1a, was further reduced to one-fourth of that of 2c. The overall results indicate that both the s-cis amide bond at the MeVal-MeAla moiety and two intramolecular hydrogen bonds (ß-Ala)NH•••OC(Ile) and (Ile)NH•••OC(ß-Ala) are important for constraining the conformation of the macrocyclic peptide backbone in destruxin E, thereby exhibiting its potent biological activity.

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BACKGROUND: Comparative efficacy and safety data on radiofrequency ablation (RFA) versus pulsed field ablation (PFA) for common idiopathic left ventricular arrhythmia (LV-VAs) locations are lacking. OBJECTIVES: This study sough to compare RFA with PFA of common idiopathic LV-VAs locations. METHODS: Ten swine were randomized to PFA or RFA of LV interventricular septum, papillary muscle, LV summit via distal coronary sinus, and LV epicardium via subxiphoid approach. Ablations were delivered using an investigational dual-energy (RFA/PFA) contact force (CF) and local impedance-sensing catheter. After 1-week survival, animals were euthanized for lesion assessment. RESULTS: A total of 55 PFA (4 applications/site of 2.0 KV, target CF ≥10 g) and 36 RFA (CF ≥10 g, 25-50 W targeting ≥50 Ω local impedance drop, 60-second duration) were performed. LV interventricular septum: average PFA depth 7.8 mm vs RFA 7.9 mm (P = 0.78) and no adverse events. Papillary muscle: average PFA depth 8.1 mm vs RFA 4.5 mm (P < 0.01). Left ventricular summit: average PFA depth 5.6 mm vs RFA 2.7 mm (P < 0.01). Steam-pop and/or ventricular fibrillation in 4 of 12 RFA vs 0 of 12 PFA (P < 0.01), no ST-segment changes observed. Epicardium: average PFA depth 6.4 mm vs RFA 3.3 mm (P < 0.01). Transient ST-segment elevations/depressions occurred in 4 of 5 swine in the PFA arm vs 0 of 5 in the RFA arm (P < 0.01). Angiography acutely and at 7 days showed normal coronaries in all cases. CONCLUSIONS: In this swine study, compared with RFA, PFA of common idiopathic LV-VAs locations produced deeper lesions with fewer steam pops. However, PFA was associated with higher rates of transient ST-segment elevations and depressions with direct epicardium ablation.

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Background: Esophageal organoids from a variety of pathologies including cancer are grown in Advanced Dulbecco's Modified Eagle Medium-Nutrient Mixture F12 (hereafter ADF). However, the currently available ADF-based formulations are suboptimal for normal human esophageal organoids, limiting the ability to compare normal esophageal organoids with those representing a given disease state. Methods: We have utilized immortalized normal human esophageal epithelial cell (keratinocyte) lines EPC1 and EPC2 and endoscopic normal esophageal biopsies to generate three-dimensional (3D) organoids. To optimize ADF-based medium, we evaluated the requirement of exogenous epidermal growth factor (EGF) and inhibition of transforming growth factor-(TGF)-ß receptor-mediated signaling, both key regulators of proliferation of human esophageal keratinocytes. We have modeled human esophageal epithelial pathology by stimulating esophageal 3D organoids with interleukin (IL)-13, an inflammatory cytokine, or UAB30, a novel pharmacological activator of retinoic acid signaling. Results: The formation of normal human esophageal 3D organoids was limited by excessive EGF and intrinsic TGFß receptor-mediated signaling. In optimized HOME0, normal human esophageal organoid formation was improved, whereas IL-13 and UAB30 induced epithelial changes reminiscent of basal cell hyperplasia, a common histopathologic feature in broad esophageal disease conditions including eosinophilic esophagitis. Conclusions: HOME0 allows modeling of the homeostatic differentiation gradient and perturbation of the human esophageal epithelium while permitting a comparison of organoids from mice and other organs grown in ADF-based media.

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Cancer cells have higher heat sensitivity compared to normal cells; therefore, hyperthermia is a promising approach for cancer therapy because of its ability to selectively kill cancer cells by heating them. However, the specific and rapid heating of tumor tissues remains challenging. This study investigated the potential of magnetic nanoparticles (MNPs) modified with tumor-homing peptides (THPs), specifically PL1 and PL3, for tumor-specific magnetic hyperthermia therapy. The synthesis of THP-modified MNPs involved the attachment of PL1 and PL3 peptides to the surface of the MNPs, which facilitated enhanced tumor cell binding and internalization. Cell specificity studies revealed an increased uptake of PL1- and PL3-MNPs by tumor cells compared to unmodified MNPs, indicating their potential for targeted delivery. In vitro hyperthermia experiments demonstrated the efficacy of PL3-MNPs in inducing tumor cell death when exposed to an alternating magnetic field (AMF). Even without exposure to an AMF, an additional ferroptotic pathway was suggested to be mediated by the nanoparticles. Thus, this study suggests that THP-modified MNPs, particularly PL3-MNPs, hold promise as a targeted approach for tumor-specific magnetic hyperthermia therapy.

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Despite the promising outcomes of immune checkpoint inhibitors (ICIs), resistance to ICI presents a new challenge. Therefore, selecting patients for specific ICI applications is crucial for maximizing therapeutic efficacy. Herein, we curated 69 human esophageal squamous cell cancer (ESCC) patients' tumor microenvironment (TME) single-cell transcriptomic datasets to subtype ESCC. Integrative analyses of the cellular network and transcriptional signatures of T cells and myeloid cells define distinct ESCC subtypes characterized by T cell exhaustion, and interleukin (IL) and interferon (IFN) signaling. Furthermore, this approach classifies ESCC patients into ICI responders and non-responders, as validated by whole tumor transcriptomes and liquid biopsy-based single-cell transcriptomes of anti-PD-1 ICI responders and non-responders. Our study stratifies ESCC patients based on TME transcriptional network, providing novel insights into tumor niche remodeling and potentially predicting ICI responses in ESCC patients.

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BACKGROUND: Conventional focal radiofrequency catheters may be modified to enable multiple energy modalities (radiofrequency or pulsed field [PF]) with the benefit of contact force (CF) feedback, providing greater flexibility in the treatment of arrhythmias. Information on the impact of CF on lesion formation in PF ablations remains limited. METHODS: An in vivo study was performed with 8 swine using an investigational dual-energy CF focal catheter with local impedance. Experiment I: To evaluate atrial lesion formation, contiguity, and width, a point-by-point approach was used to create an intercaval line. The distance between the points was prespecified at 4±1 mm. Half of the line was created with radiofrequency energy, whereas the other half utilized PF (single 2.0 kV application with a proprietary waveform). Experiment II: To evaluate single application lesion dimensions with a proprietary waveform, discrete ventricular lesions were performed with PFA (single 2.0 kV application) with targeted levels of CF: low, 5 to 15 g; medium, 20 to 30 g; and high, 35 to 45 g. Following 1 week of survival, animals underwent endocardial/epicardial remapping, and euthanasia to enable histopathologic examination. RESULTS: Experiment I: Both energy modalities resulted in a complete intercaval line of transmural ablation. PF resulted in significantly wider lines than radiofrequency: minimum width, 14.9±2.3 versus 5.0±1.6 mm; maximum width, 21.8±3.4 versus 7.3±2.1 mm, respectively; P<0.01 for each. Histology confirmed transmural lesions with both modalities. Experiment II: With PF, lesion depth, width, and volume were larger with higher degrees of CF (depth: r=0.82, P<0.001; width: r=0.26, P=0.052; and volume: r=0.55, P<0.001), with depth increasing at a faster rate than width. The mean depths were as follows: low (n=17), 4.3±1.0 mm; medium (n=26), 6.4±1.2 mm; and high (n=14), 9.1±1.4 mm. CONCLUSIONS: Using the same focal point CF-sensing catheter, a novel PF ablation waveform with a single application resulted in transmural atrial lesions that were significantly wider than radiofrequency. Lesion depth showed a significant positive correlation with CF with depths of 6.4 mm at moderate CF.

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Esophageal squamous cell carcinoma (ESCC) is a deadly consequence of radiation exposure to the esophagus. ESCC arises from esophageal epithelial cells that undergo malignant transformation and features a perturbed squamous cell differentiation program. Understanding the dose- and radiation quality-dependence of the esophageal epithelium response to radiation may provide insights into the ability of radiation to promote ESCC. We have explored factors that may play a role in esophageal epithelial radiosensitivity and their potential relationship to ESCC risk. We have utilized a murine three-dimensional (3D) organoid model that recapitulates the morphology and functions of the stratified squamous epithelium of the esophagus to study persistent dose- and radiation quality-dependent changes. Interestingly, although high-linear energy transfer (LET) Fe ion exposure induced a more intense and persistent alteration of squamous differentiation and 53BP1 DNA damage foci levels as compared to Cs, the MAPK/SAPK stress pathway signaling showed similar altered levels for most phospho-proteins with both radiation qualities. In addition, the lower dose of high-LET exposure also revealed nearly the same degree of morphological changes, even though only ~36% of the cells were predicted to be hit at the lower 0.1 Gy dose, suggesting that a bystander effect may be induced. Although p38 and ERK/MAPK revealed the highest levels following high-LET exposure, the findings reveal that even a low dose (0.1 Gy) of both radiation qualities can elicit a persistent stress signaling response that may critically impact the differentiation gradient of the esophageal epithelium, providing novel insights into the pathogenesis of radiation-induced esophageal injury and early stage esophageal carcinogenesis.

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During tumor development, promoter CpG islands (CGIs) that are normally silenced by Polycomb repressive complexes (PRCs) become DNA hypermethylated. The molecular mechanism by which de novo DNA methyltransferase(s) catalyze CpG methylation at PRC-regulated regions remains unclear. Here we report a cryo-EM structure of the DNMT3A long isoform (DNMT3A1) N-terminal region in complex with a nucleosome carrying PRC1-mediated histone H2A lysine 119 monoubiquitination (H2AK119Ub). We identify regions within the DNMT3A1 N-terminus that bind H2AK119Ub and the nucleosome acidic patch. This bidentate interaction is required for effective DNMT3A1 engagement with H2AK119Ub-modified chromatin in cells. Furthermore, aberrant redistribution of DNMT3A1 to Polycomb target genes inhibits their transcriptional activation during cell differentiation and recapitulates the cancer-associated DNA hypermethylation signature. This effect is rescued by disruption of the DNMT3A1-acidic patch interaction. Together, our analyses reveal a binding interface critical for countering promoter CGI DNA hypermethylation, a major molecular hallmark of cancer.

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BACKGROUND: Data are lacking on patient-reported outcomes (PRO) following cryoballoon ablation (CBA) versus radiofrequency ablation (RFA). We sought to evaluate QoL and clinical outcomes of cryoballoon pulmonary vein isolation only (CRYO-PVI-ONLY) versus RFA with PVI and posterior wall isolation (RF-PVI+PWI) in a large prospective PRO registry. METHODS: Patients who underwent AF ablation (2013-2016) at our institution were enrolled in an automated, prospectively maintained PRO registry. CRYO-PVI-ONLY patients were matched (1:1) with RF-PVI+PWI patients based on age, gender, and type of AF (paroxysmal vs. persistent). QoL and clinical outcomes were assessed using PRO surveys at baseline and at 1-year. The atrial fibrillation symptom severity scale (AFSSS) was the measure for QoL. Additionally, we assessed patient-reported clinical improvement, arrhythmia recurrence, and AF burden (as indicated by AF frequency and duration scores). RESULTS: A total of 296 patients were included (148 in each group, 72% paroxysmal). By PRO, a significant improvement in QoL was observed in the overall study population and was comparable between CRYO-PVI-ONLY and RF-PVI+PWI (baseline median AFSSS of 11.5 and 11; reduced to 2 and 4 at 1 year, respectively; p = 0.44). Similarly, the proportion of patients who reported improvement in their overall QoL and AF related symptoms was high and similar between the study groups [92% (CRYO-PVI-ONLY) vs. 92.8% (RF-PVI+PWI); p = 0.88]. Arrhythmia recurrence was significantly more common in the CRYO-PVI-ONLY group (39.7%) compared to RF-PVI+PWI (27.7 %); p = 0.03. Comparable results were observed in paroxysmal and persistent AF. CONCLUSION: CRYO-PVI-ONLY and RF-PVI+PWI resulted in comparable improvements in patient reported outcomes including QoL and AF burden; with RF-PVI+PWI being more effective at reducing recurrences.

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The co-assembly of polyelectrolytes (PE) with proteins offers a promising approach for designing complex structures with customizable morphologies, charge distribution, and stability for targeted cargo delivery. However, the complexity of protein structure limits our ability to predict the properties of the formed nanoparticles, and our goal is to identify the key triggers of the morphological transition in protein/PE complexes and evaluate their ability to encapsulate multivalent ionic drugs. A positively charged PE can assemble with a protein at pH above isoelectric point due to the electrostatic attraction and disassemble at pH below isoelectric point due to the repulsion. The additional hydrophilic block of the polymer should stabilize the particles in solution and enable them to encapsulate a negatively charged drug in the presence of PE excess. We demonstrated that diblock copolymers, poly(ethylene oxide)-block-poly(N,N-dimethylaminoethyl methacrylate) and poly(ethylene oxide)-block-poly(N,N,N-trimethylammonioethyl methacrylate), consisting of a polycation block and a neutral hydrophilic block, reversibly co-assemble with insulin in pH range between 5 and 8. Using small-angle neutron and X-ray scattering (SANS, SAXS), we showed that insulin arrangement within formed particles is controlled by intermolecular electrostatic forces between protein molecules, and can be tuned by varying ionic strength. For the first time, we observed by fluorescence that formed protein/PE complexes with excess of positive charges exhibited potential for encapsulating and controlled release of negatively charged bivalent drugs, protoporphyrin-IX and zinc(II) protoporphyrin-IX, enabling the development of nanocarriers for combination therapies with adjustable charge, stability, internal structure, and size.

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BACKGROUND: Eosinophilic esophagitis (EoE) is an increasingly common inflammatory condition of the esophagus; however, the underlying immunologic mechanisms remain poorly understood. The epithelium-derived cytokine IL-33 is associated with type 2 immune responses and elevated in esophageal biopsy specimens from patients with EoE. OBJECTIVE: We hypothesized that overexpression of IL-33 by the esophageal epithelium would promote the immunopathology of EoE. METHODS: We evaluated the functional consequences of esophageal epithelial overexpression of a secreted and active form of IL-33 in a novel transgenic mouse, EoE33. EoE33 mice were analyzed for clinical and immunologic phenotypes. Esophageal contractility was assessed. Epithelial cytokine responses were analyzed in three-dimensional organoids. EoE33 phenotypes were further characterized in ST2-/-, eosinophil-deficient, and IL-13-/- mice. Finally, EoE33 mice were treated with dexamethasone. RESULTS: EoE33 mice displayed ST2-dependent, EoE-like pathology and failed to thrive. Esophageal tissue remodeling and inflammation included basal zone hyperplasia, eosinophilia, mast cells, and TH2 cells. Marked increases in levels of type 2 cytokines, including IL-13, and molecules associated with immune responses and tissue remodeling were observed. Esophageal organoids suggested reactive epithelial changes. Genetic deletion of IL-13 in EoE33 mice abrogated pathologic changes in vivo. EoE33 mice were responsive to steroids. CONCLUSIONS: IL-33 overexpression by the esophageal epithelium generated immunopathology and clinical phenotypes resembling human EoE. IL-33 may play a pivotal role in the etiology of EoE by activating the IL-13 pathway. EoE33 mice are a robust experimental platform for mechanistic investigation and translational discovery.

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