Aire in Autoimmunity.

The role of the autoimmune regulator (Aire) in central immune tolerance and thymic self-representation was first described more than 20 years ago, but fascinating new insights into its biology continue to emerge, particularly in the era of advanced single-cell genomics. We briefly describe the role of human genetics in the discovery of Aire, as well as insights into its function gained from genotype-phenotype correlations and the spectrum of Aire-associated autoimmunity-including insights from patients with Aire mutations with broad and diverse implications for human health. We then highlight emerging trends in Aire biology, focusing on three topic areas. First, we discuss medullary thymic epithelial diversity and the role of Aire in thymic epithelial development. Second, we highlight recent developments regarding the molecular mechanisms of Aire and its binding partners. Finally, we describe the rapidly evolving biology of the identity and function of extrathymic Aire-expressing cells (eTACs), and a novel eTAC subset called Janus cells, as well as their potential roles in immune homeostasis. Expected final online publication date for the Annual Review of Immunology, Volume 42 is April 2024. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

Cholangioscopy-guided tunneling and coaxial stenting of a large choledocholithiasis: a novel approach to mechanical lithotripsy.

Video 1Cholangioscopy-guided tunneling and coaxial stenting of a large choledocholithiasis: a novel approach to mechanical lithotripsy. Fluoroscopy image of initial ERCP shows large filling defect (blue circle) and guidewire passing toward the common hepatic duct (black arrow).

Aspirin exposure and its association with metal stent patency in malignant distal biliary obstruction: a large international multicenter propensity score-matched study.

BACKGROUND AND AIMS: Stent dysfunction is common after ERCP with self-expandable metal stent (SEMS) insertion for malignant distal biliary obstruction (MDBO). Chronic aspirin (acetylsalicylic acid; ASA) exposure has been previously shown to potentially decrease this risk. We aim to further ascertain the protective effect of ASA and to identify other predictors of stent dysfunction. METHODS: This multicenter retrospective cohort study was conducted at 9 sites in Canada and 1 in the United States. Patients with MDBO who underwent ERCP with SEMS placement between January 2014 and December 2019 were included and divided into 2 cohorts: ASA exposed (ASA-E) and ASA unexposed (ASA-U). Propensity-score matching (PSM) was performed to limit selection bias. Matched variables were age, sex, tumor stage, and type of metal stent. The primary outcome was the hazard rate of stent dysfunction. A multivariable Cox proportional hazards model was used to identify independent predictors of stent dysfunction. RESULTS: Of 1396 patients assessed, after PSM 496 patients were analyzed (248 ASA-E and 248 ASA-U). ERCP with SEMS placement was associated with a high clinical success of 82.2% in ASA-E and 81.2% in ASA-U cohorts (P = .80). One hundred eighty-four patients had stent dysfunction with a mean stent patency time of 229.9 ± 306.2 days and 245.4 ± 241.4 days in ASA-E and ASA-U groups, respectively (P = .52). On multivariable analysis, ASA exposure did not protect against stent dysfunction (hazard ratio [HR], 1.25; 95% confidence interval [CI], .96-1.63). An etiology of pancreatic cancer (HR, 1.36; 95% CI, 1.15-1.61) predicted stent dysfunction, whereas cancer therapy was protective (HR, .73; 95% CI, .55-.96). Chronic ASA use was not associated with an increased risk for adverse events including bleeding, post-ERCP pancreatitis, and perforation. CONCLUSIONS: In this large, multicenter study using PSM, chronic exposure to ASA did not protect against stent dysfunction in MDBO. Instead, the analysis revealed that the etiology of pancreatic cancer was an independent predictor of stent dysfunction and cancer therapy was protective.

Cortical bone adaptation response is region specific, but not peak load dependent: insights from µ CT image analysis and mechanostat simulations of the mouse tibia loading model.

The two major aims of the present study were: (i) quantify localised cortical bone adaptation at the surface level using contralateral endpoint imaging data and image analysis techniques, and (ii) investigate whether cortical bone adaptation responses are universal or region specific and dependent on the respective peak load. For this purpose, we re-analyse previously published µ CT data of the mouse tibia loading model that investigated bone adaptation in response to sciatic neurectomy and various peak load magnitudes (F = 0, 2, 4, 6, 8, 10, 12 N). A beam theory-based approach was developed to simulate cortical bone adaptation in different sections of the tibia, using longitudinal strains as the adaptive stimuli. We developed four mechanostat models: universal, surface-based, strain directional-based, and combined surface and strain direction-based. Rates of bone adaptation in these mechanostat models were computed using an optimisation procedure (131,606 total simulations), performed on a single load case (F = 10 N). Subsequently, the models were validated against the remaining six peak loads. Our findings indicate that local bone adaptation responses are quasi-linear and bone region specific. The mechanostat model which accounted for differences in endosteal and periosteal regions and strain directions (i.e. tensile versus compressive) produced the lowest root mean squared error between simulated and experimental data for all loads, with a combined prediction accuracy of 76.6, 55.0 and 80.7% for periosteal, endosteal, and cortical thickness measurements (in the midshaft of the tibia). The largest root mean squared errors were observed in the transitional loads, i.e. F = 2 to 6 N, where inter-animal variability was highest. Finally, while endpoint imaging studies provide great insights into organ level bone adaptation responses, the between animal and loaded versus control limb variability make simulations of local surface-based adaptation responses challenging.

Topical Tacrolimus Compared With Oral Tacrolimus for Postoperative Immunosuppression in Primary Keratolimbal Allograft.

PURPOSE: The aim of this study was to compare outcomes between topical tacrolimus and oral tacrolimus as the primary calcineurin inhibitor for postoperative immunosuppression after primary keratolimbal allograft (KLAL) transplantation for limbal stem cell deficiency (LSCD). METHODS: We performed a retrospective, comparative cohort study at a single tertiary referral center (University of MN) of all patients who underwent primary KLAL between 2014 and 2021. Eyes were grouped into those which received topical tacrolimus as the only calcineurin inhibitor (topical group) and eyes in which patients received oral tacrolimus with or without topical tacrolimus (oral group). Clinical and donor tissue data were obtained and compared between the 2 groups. RESULTS: In total, 27 eyes of 22 patients (median age 42 years, range 20-79 years) were included, of which 18 eyes were in the oral group and 9 eyes were in the topical group. The mean follow-up time was 33.2 ± 22.6 months. The most frequent etiology of LSCD was alkaline burn (33.3%). At 36 months, graft failure occurred in 6 eyes in the oral group (33.3%) and 2 eyes in the topical group (22.1%) ( P = 0.57). The failure rate in the oral group was 9.1 per 1000 person-months versus 8.4 per 1000 person-months in the topical group ( P = 0.96). The median improvement in BCVA was logMAR -0.975 and logMAR -0.45 for the oral and topical group, respectively ( P = 0.50). CONCLUSIONS: With careful patient selection, topical tacrolimus may be a viable alternative to oral tacrolimus in KLAL.

Cortical thickness adaptation to combined mechanical loading and parathyroid hormone treatments is site specific and synergistic in the mouse tibia model.

In this study, we aimed to quantify the localised effects of mechanical loading (ML), low (20 µg/kg/day), moderate (40 µg/kg/day) or high (80 µg/kg/day) dosages of parathyroid hormone (PTH), and combined (PTHML) treatments on cortical bone adaptation in healthy 19-week old female C57BL/6 mice. To this end, we utilise a previously reported image analysis algorithm on µCT data of the mouse tibia published by Sugiyama et al. (2008) to measure changes in cortical area, marrow cavity area and local cortical thickness measures (ΔCt.Ar, ΔMa.Ar, ΔCt.Th respectively), evaluated at two cross-sections within the mouse tibia (proximal-middle (37 %) and middle (50 %)), and are compared to a superposed summation (P + M) of individual treatments to determine the effectiveness of combining treatments in vivo. ΔCt.Ar analysis revealed a non-linear, synergistic interactions between PTH and ML in the 37 % cross-section that saturates at higher PTH dosages, whereas the 50 % cross-section experiences an approximately linear, additive adaptation response. This coincided with an increase in ΔMa.Ar (indicating resorption of the endosteal surface), which was only counteracted by combined high dose PTH with ML in the middle cross-section. Regional analysis of ΔCt.Th changes reveal localised cortical thinning in response to low dose PTH treatment in the posteromedial region of the middle cross-section, signifying that PTH does not provide a homogeneous adaptation response around the cortical perimeter. We observe a synergistic response in the proximal-middle cross-section, with regions of compressive strain experiencing the greatest adaptation response to PTHML treatments, (peak ΔCt.Th of 189.32, 213.78 and 239.30 µm for low, moderate and high PTHML groups respectively). In contrast, PTHML treatments in the middle cross-section show a similar response to the superposed P + M group, with the exception of the combined high dose PTHML treatment which shows a synergistic interaction. These analyses suggest that, in mice, adding mechanical loading to PTH treatments leads to region specific bone responses; synergism of PTHML is only achieved in some regions experiencing high loading, while other regions respond additively to this combined treatment.

EUS-guided gastroenterostomy using a novel through-the-scope exchangeable dual-balloon enteroclysis catheter: a potentially secure and scalable approach.

Video 1Clinical case of dual-balloon through-the-scope exchangeable enteroclysis catheter-assisted EUS-guided gastroenterostomy.

Autoantibodies against type I IFNs in humans with alternative NF-κB pathway deficiency.

Patients with autoimmune polyendocrinopathy syndrome type 1 (APS-1) caused by autosomal recessive AIRE deficiency produce autoantibodies that neutralize type I interferons (IFNs)1,2, conferring a predisposition to life-threatening COVID-19 pneumonia3. Here we report that patients with autosomal recessive NIK or RELB deficiency, or a specific type of autosomal-dominant NF-κB2 deficiency, also have neutralizing autoantibodies against type I IFNs and are at higher risk of getting life-threatening COVID-19 pneumonia. In patients with autosomal-dominant NF-κB2 deficiency, these autoantibodies are found only in individuals who are heterozygous for variants associated with both transcription (p52 activity) loss of function (LOF) due to impaired p100 processing to generate p52, and regulatory (IκBδ activity) gain of function (GOF) due to the accumulation of unprocessed p100, therefore increasing the inhibitory activity of IκBδ (hereafter, p52LOF/IκBδGOF). By contrast, neutralizing autoantibodies against type I IFNs are not found in individuals who are heterozygous for NFKB2 variants causing haploinsufficiency of p100 and p52 (hereafter, p52LOF/IκBδLOF) or gain-of-function of p52 (hereafter, p52GOF/IκBδLOF). In contrast to patients with APS-1, patients with disorders of NIK, RELB or NF-κB2 have very few tissue-specific autoantibodies. However, their thymuses have an abnormal structure, with few AIRE-expressing medullary thymic epithelial cells. Human inborn errors of the alternative NF-κB pathway impair the development of AIRE-expressing medullary thymic epithelial cells, thereby underlying the production of autoantibodies against type I IFNs and predisposition to viral diseases.

Ikaros is a principal regulator of Aire+ mTEC homeostasis, thymic mimetic cell diversity, and central tolerance.

Mutations in the gene encoding the zinc-finger transcription factor Ikaros (IKZF1) are found in patients with immunodeficiency, leukemia, and autoimmunity. Although Ikaros has a well-established function in modulating gene expression programs important for hematopoietic development, its role in other cell types is less well defined. Here, we uncover functions for Ikaros in thymic epithelial lineage development in mice and show that Ikzf1 expression in medullary thymic epithelial cells (mTECs) is required for both autoimmune regulator-positive (Aire+) mTEC development and tissue-specific antigen (TSA) gene expression. Accordingly, TEC-specific deletion of Ikzf1 in mice results in a profound decrease in Aire+ mTECs, a global loss of TSA gene expression, and the development of autoimmunity. Moreover, Ikaros shapes thymic mimetic cell diversity, and its deletion results in a marked expansion of thymic tuft cells and muscle-like mTECs and a loss of other Aire-dependent mimetic populations. Single-cell analysis reveals that Ikaros modulates core transcriptional programs in TECs that correlate with the observed cellular changes. Our findings highlight a previously undescribed role for Ikaros in regulating epithelial lineage development and function and suggest that failed thymic central tolerance could contribute to the autoimmunity seen in humans with IKZF1 mutations.

EUS-guided gastroenterostomy vs. surgical gastrojejunostomy and enteral stenting for malignant gastric outlet obstruction: a meta-analysis.

Background and study aims Malignant gastric outlet obstruction (MGOO) is traditionally treated with surgical gastrojejunostomy (SGJ), which is effective but associated with high rates of morbidity, or endoscopic stenting (ES), which is less invasive but associated with significant risk of stent dysfunction and need for reintervention. Endoscopic ultrasound-guided gastroenterostomy (EUS-GE) provides a robust bypass without the invasiveness of surgery. Methods We performed a systematic review and meta-analysis comparing EUS-GE to SGJ and ES for MGOO. Electronic databases were searched from inception through February 2022. A meta-analysis was performed with results reported as odds ratios (ORs) with 95% confidence intervals (CIs) using random effects models. Primary outcomes included clinical success without recurrent GOO and adverse events (AEs). Results Sixteen studies involving 1541 patients were included. EUS-GE was associated with higher clinical success without recurrent GOO compared to ES or SGJ [OR 2.60, 95% CI1.58-4.28] and compared to ES alone [OR 5.08, 95% CI 3.42-7.55], but yielded no significant difference compared to SGJ alone [OR 1.94, 95% CI 0.97-3.88]. AE rates were significantly lower for EUS-GE compared to ES or SGJ grouped together [OR 0.34, 95% CI 0.20-0.58], or SGJ alone [OR 0.17, 95% CI 0.10-0.30] but were not significant different versus ES alone [OR 0.57, 95% CI 0.29-1.14]. Conclusions EUS-GE is the most successful approach to treating MGOO, exhibiting a lower risk of recurrent obstruction compared to ES, and fewer AEs compared to SGJ.

Endoscopic Ultrasound-Guided Biliary Drainage of First Intent With a Lumen-Apposing Metal Stent vs Endoscopic Retrograde Cholangiopancreatography in Malignant Distal Biliary Obstruction: A Multicenter Randomized Controlled Study (ELEMENT Trial).

BACKGROUND & AIMS: Endoscopic ultrasound-guided choledochoduodenostomy with a lumen-apposing metal stent (EUS-CDS) is a promising modality for management of malignant distal biliary obstruction (MDBO) with potential for better stent patency. We compared its outcomes with endoscopic retrograde cholangiopancreatography with metal stenting (ERCP-M). METHODS: In this multicenter randomized controlled trial, we recruited patients with MDBO secondary to borderline resectable, locally advanced, or unresectable peri-ampullary cancers across 10 Canadian institutions and 1 French institution. This was a superiority trial with a noninferiority assessment of technical success. Patients were randomized to EUS-CDS or ERCP-M. The primary end point was the rate of stent dysfunction at 1 year, considering competing risks of death, clinical failure, and surgical resection. Analyses were performed according to intention-to-treat principles. RESULTS: From February 2019 to February 2022, 144 patients were recruited; 73 were randomized to EUS-CDS and 71 were randomized to ERCP-M. The mean (SD) procedure time was 14.0 (11.4) minutes for EUS-CDS and 23.1 (15.6) minutes for ERCP-M (P < .01); 40% of the former was performed without fluoroscopy. Technical success was achieved in 90.4% (95% CI, 81.5% to 95.3%) of EUS-CDS and 83.1% (95% CI, 72.7% to 90.1%) of ERCP-M with a risk difference of 7.3% (95% CI, -4.0% to 18.8%) indicating noninferiority. Stent dysfunction occurred in 9.6% vs 9.9% of EUS-CDS and ERCP-M cases, respectively (P = .96). No differences in adverse events, pancreaticoduodenectomy and oncologic outcomes, or quality of life were noted. CONCLUSIONS: Although not superior in stent function, EUS-CDS is an efficient and safe alternative to ERCP-M in patients with MDBO. These findings provide evidence for greater adoption of EUS-CDS in clinical practice as a complementary and exchangeable first-line modality to ERCP in patients with MDBO. CLINICALTRIALS: gov, Number: NCT03870386.

SIV clearance from neonatal macaques following transient CCR5 depletion.

Treatment of people with HIV (PWH) with antiretroviral therapy (ART) results in sustained suppression of viremia, but HIV persists indefinitely as integrated provirus in CD4-expressing cells. Intact persistent provirus, the "rebound competent viral reservoir" (RCVR), is the primary obstacle to achieving a cure. Most variants of HIV enter CD4 + T cells by binding to the chemokine receptor, CCR5. The RCVR has been successfully depleted only in a handful of PWH following cytotoxic chemotherapy and bone marrow transplantation from donors with a mutation in CCR5 . Here we show that long-term SIV remission and apparent cure can be achieved for infant macaques via targeted depletion of potential reservoir cells that express CCR5. Neonatal rhesus macaques were infected with virulent SIVmac251, then treated with ART beginning one week after infection, followed by treatment with either a CCR5/CD3-bispecific or a CD4-specific antibody, both of which depleted target cells and increased the rate of plasma viremia decrease. Upon subsequent cessation of ART, three of seven animals treated with CCR5/CD3-bispecific antibody rebounded quickly and two rebounded 3 or 6 months later. Remarkably, the other two animals remained aviremic and efforts to detect replication-competent virus were unsuccessful. Our results show that bispecific antibody treatment can achieve meaningful SIV reservoir depletion and suggest that functional HIV cure might be achievable for recently infected individuals having a restricted reservoir.

Ehf and Fezf2 regulate late medullary thymic epithelial cell and thymic tuft cell development.

Thymic epithelial cells are indispensable for T cell maturation and selection and the induction of central immune tolerance. The self-peptide repertoire expressed by medullary thymic epithelial cells is in part regulated by the transcriptional regulator Aire (Autoimmune regulator) and the transcription factor Fezf2. Due to the high complexity of mTEC maturation stages (i.e., post-Aire, Krt10+ mTECs, and Dclk1+ Tuft mTECs) and the heterogeneity in their gene expression profiles (i.e., mosaic expression patterns), it has been challenging to identify the additional factors complementing the transcriptional regulation. We aimed to identify the transcriptional regulators involved in the regulation of mTEC development and self-peptide expression in an unbiased and genome-wide manner. We used ATAC footprinting analysis as an indirect approach to identify transcription factors involved in the gene expression regulation in mTECs, which we validated by ChIP sequencing. This study identifies Fezf2 as a regulator of the recently described thymic Tuft cells (i.e., Tuft mTECs). Furthermore, we identify that transcriptional regulators of the ELF, ESE, ERF, and PEA3 subfamily of the ETS transcription factor family and members of the Krüppel-like family of transcription factors play a role in the transcriptional regulation of genes involved in late mTEC development and promiscuous gene expression.

Pan-Esophageal Submucosal Dissection following Transesophageal Echocardiography.

A 73-year-old female underwent open mitral valve replacement with transesophageal echocardiography (TEE) guidance. She developed upper gastrointestinal bleeding postoperatively and was found on upper endoscopy to have a bleeding site at the gastric cardia with the appearance of focal trauma and a possible puncture site. A submucosal bluish protrusion was seen throughout the esophagus with a mucosal flap at the proximal esophagus. As a unifying diagnosis, it was suspected that the intraoperative TEE probe caused a submucosal dissection with point of entry at the proximal esophagus, running the entire length of the esophagus and exiting at the gastric cardia, giving rise to a clinical upper gastrointestinal bleed. Closure of the esophageal defect was achieved using an endoclip. A CT scan showed focal pneumomediastinum along the proximal esophagus, confirming the hypothesis. We report the first case to our knowledge of iatrogenic pan-esophageal submucosal dissection, which, in this case, presented as a clinical bleed from the exit point trauma to the gastric cardia mucosa caused by a TEE probe. Endoscopic management of the gastric injury as well as the esophageal defect led to resolution of the bleeding and avoidance of mediastinitis, respectively, allowing for an excellent recovery.

Preoperative Endoscopic Ultrasound Fine Needle Aspiration Versus Upfront Surgery in Resectable Pancreatic Cancer: A Systematic Review and Meta-analysis of Clinical Outcomes Including Survival and Risk of Tumor Recurrence.

Background and Aim: Endoscopic ultrasound-guided fine needle aspiration (EUS-FNA) is the standard of care in advanced pancreatic cancer. Its role in resectable disease, however, is controversial. This meta-analysis aims to ascertain the clinical outcomes of patients with resectable pancreatic cancer undergoing preoperative EUS-FNA compared to those going directly to surgery. Methods: A literature search was performed from 1996 to April 2019 using MEDLINE, EMBASE, and ISI Web of Knowledge for studies comparing preoperative EUS-FNA to EUS without FNA in resectable pancreatic cancer for clinical outcomes. The primary outcome is overall survival (OS). Secondary outcomes include cancer-free survival, tumor recurrence and peritoneal carcinomatosis, and post-FNA-pancreatitis rate. Results: Six retrospective studies were included. Preoperative EUS-FNA had better OS than the non-FNA group (WMD, 4.40 months [0.02 to 8.78]). Cancer-free survival did not differ significantly between the two groups (WMD, 2.08 months [-2.22 to 6.38]). EUS with FNA was not associated with increased rates of tumor recurrence or peritoneal carcinomatosis. Conclusion: Preoperative EUS-FNA in resectable pancreatic cancer may be associated with significantly greater OS when compared to the non-FNA group, with no significant difference in the rates of tumor recurrence or peritoneal seeding. Important limitations of our meta-analysis include the lack of prospective controlled data, which are unlikely to emerge given feasible constraints.

Modified endoscopic ultrasound-guided double-balloon-occluded gastroenterostomy bypass (M-EPASS): a pilot study.

INTRODUCTION: We recently developed a double-balloon device, using widely available existing technology, to facilitate endoscopic ultrasound-guided gastroenterostomy (EUS-GE). Our aim is to assess the feasibility of this modified approach to EUS-guided double-balloon-occluded gastroenterostomy bypass (M-EPASS). METHODS: This was a single-center retrospective study of consecutive patients undergoing M-EPASS from January 2019 to August 2020. The double-balloon device consists of two vascular balloons that optimize the distension of a targeted small-bowel segment for EUS-guided stent insertion. The primary end point was the rate of technical success. RESULTS: 11 patients (45 % women; mean [standard deviation (SD)] age 64.9 [8.6]) with malignant gastric outlet obstruction were included. Technical and clinical success (ability to tolerate an oral diet) were achieved in 91 % (10/11) and 80 % (8/10) of patients, respectively. There was one adverse event (9 %) due to stent migration. Two patients (18 %) required re-intervention for stent obstruction secondary to food impaction. The mean (SD) time to a low residue diet was 3.5 (2.4) days. CONCLUSION: M-EPASS appears to facilitate the technique of EUS-GE, potentially enhancing its safety and clinical adoption. Larger studies are needed to validate this innovative approach to gastric outlet obstruction.

Endoscopic ultrasound (EUS)-guided fine needle biopsy alone vs. EUS-guided fine needle aspiration with rapid onsite evaluation in pancreatic lesions: a multicenter randomized trial.

BACKGROUND: Endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) is the standard in the diagnosis of solid pancreatic lesions, in particular when combined with rapid onsite evaluation of cytopathology (ROSE). More recently, a fork-tip needle for core biopsy (FNB) has been shown to be associated with excellent diagnostic yield. EUS-FNB alone has however not been compared with EUS-FNA + ROSE in a large clinical trial. Our aim was to compare EUS-FNB alone to EUS-FNA + ROSE in solid pancreatic lesions. METHODS: A multicenter, non-inferiority, randomized controlled trial involving seven centers was performed. Solid pancreatic lesions referred for EUS were considered for inclusion. The primary end point was diagnostic accuracy. Secondary end points included sensitivity/specificity, mean number of needle passes, and cost. RESULTS: 235 patients were randomized: 115 EUS-FNB alone and 120 EUS-FNA + ROSE. Overall, 217 patients had malignant histology. The diagnostic accuracy for malignancy of EUS-FNB alone was non-inferior to EUS-FNA + ROSE at 92.2 % (95 %CI 86.6 %-96.9 %) and 93.3 % (95 %CI 88.8 %-97.9 %), respectively (P = 0.72). Diagnostic sensitivity for malignancy was 92.5 % (95 %CI 85.7 %-96.7 %) for EUS-FNB alone vs. 96.5 % (93.0 %-98.6 %) for EUS-FNA + ROSE (P = 0.46), while specificity was 100 % in both. Adequate histological yield was obtained in 87.5 % of the EUS-FNB samples. The mean (SD) number of needle passes and procedure time favored EUS-FNB alone (2.3 [0.6] passes vs. 3.0 [1.1] passes [P < 0.001]; and 19.3 [8.0] vs. 22.7 [10.8] minutes [P = 0.008]). EUS-FNB alone cost on average 45 US dollars more than EUS-FNA + ROSE. CONCLUSION: EUS-FNB alone is non-inferior to EUS-FNA + ROSE and is associated with fewer needle passes, shorter procedure time, and excellent histological yield at comparable cost.