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Gut fungal dysbiosis has been implicated in inflammatory bowel disease (IBD), yet strategies for targeting the gut mycobiota in IBD remain unexplored. Here we leveraged the observation that Candida albicans strains are shared between the oral cavity and gut in patients with IBD with mild oral thrush, a condition caused by Candida overgrowth, to design a prospective observational study comparing oral antifungal therapy (swish-and-spit nystatin; oral nystatin fungal targeting (ORNT); n = 18) with orogastrointestinal antifungal therapy (fluconazole; gastrointestinal and oral fluconazole fungal targeting (GIFT); n = 35). Among 53 patients with mild-to-moderate ulcerative colitis or Crohn's disease, fluconazole, but not nystatin, effectively reduced intestinal Candida burden and reshaped gut fungal-community composition. Fluconazole treatment was accompanied by increased bacterial diversity, expansion of short-chain fatty-acid-producing taxa, restoration of anti-inflammatory microbial metabolites and durable shifts in cross-kingdom microbial networks. These microbiome and metabolomic changes coincided with improved disease activity indices and a decreased risk of disease progression over the 8-week follow-up period. These findings demonstrate the feasibility of mycobiome-based patient stratification, provide evidence that targeted antifungal therapy can reshape the intestinal microbiota in IBD and establish a framework for implementing antifungal cotherapy in patients with fungal-associated disease manifestations.

The gut microbiome is a dynamic ecosystem in which microorganisms constantly adjust their transcriptional programmes. Here we developed metastrand, a framework that integrates strand-aware metatranscriptomics and metagenomics to quantify mRNAs and antisense RNAs (asRNAs) in complex microbial communities at gene-level resolution. In inflammatory bowel disease (IBD), microbial asRNA programmes converged across patients during active disease, correlated with faecal metabolites and calprotectin levels and remained stable during persistent inflammation, highlighting their potential as biomarkers of inflammatory activity in the gut. These programmes involved antisense-to-sense transcriptional shifts at insertion sequence elements with functionally diverse passenger genes and preceded their detection at new genomic locations, linking asRNA dynamics to structural genome rearrangements and redistribution of adaptive functions under selective pressure. Similar dynamics were observed in a mouse model of colitis, oxidative stress in vitro and in patients with pathogen-confirmed gastroenteritis, establishing asRNAs as an important dimension of microbial adaptation in health and disease.

A substantial proportion of persons with inflammatory bowel disease (IBD) in remission continue to experience abdominal pain, altered bowel habits, and bloating that resemble irritable bowel syndrome (IBS). Lack of standardized definitions and evidence-based management strategies leads to diagnostic ambiguity and potentially unnecessary escalation of IBD therapy. A joint Rome Foundation/International Organization for the Study of Inflammatory Bowel Disease Working Team developed consensus recommendations on nomenclature, evaluation, and treatment of IBD with IBS-like symptoms. A multidisciplinary international panel applied a modified RAND/UCLA Appropriateness Method. Systematic literature reviews informed statement generation across multiple domains: nomenclature, diagnostic and symptom assessment, dietary therapies, drugs, and brain-gut behavioral therapies. Panelists rated the appropriateness of candidate statements independently on a 9-point Likert scale, followed by anonymized feedback, discussion, and re-voting across 2 iterative rounds. Thirteen panelists reviewed 133 initial statements; 105 proceeded to final scoring. Of these, 86 were rated appropriate, 16 uncertain, and 3 inappropriate. The preferred term was "IBD with IBS-like symptoms," defined as abdominal pain, bowel habit change, and/or bloating not explained by active inflammation or structural disease. For clinical care, diagnosis should combine Rome clinical criteria with objective exclusion of inflammation. For research, candidate thresholds for endoscopic, histologic, biomarker, and imaging remission were endorsed. Appropriate therapies include psyllium (if no stricture), a short-term low fermentable oligosaccharides, disaccharides, monosaccharides, and polyols (FODMAP) diet, targeted drugs, and brain-gut behavior therapies. This first joint consensus provides standardized terminology, evaluation strategies, and treatment recommendations for IBD with IBS-like symptoms, supporting improved clinical management and guiding future mechanistic and therapeutic research.

Proteolytic imbalance involving host and microbial serine proteases and their inhibitors (serpins) contributes to intestinal barrier disruption and chronic inflammation in inflammatory bowel disease (IBD). However, the interplay among these components remains insufficiently characterized. Here, we survey 13,304 publications over five decades, to map protease-serpin-microbiome literature in IBD. Our analysis reveals a fragmented literature structure, with uneven coverage and limited cross-domain integration among host proteases, microbial proteases, and inhibitory pathways. We synthesize evidence linking these components to intestinal barrier integrity, mucosal immunity, extracellular-matrix remodeling, and microbial ecology. Considering the protease-serpin-microbiome axis may provide an integrative direction for future studies of IBD pathogenesis, including the investigation of mechanisms underlying disease heterogeneity, prioritizing future biomarker and therapeutic studies.

Drug resistance in inflammatory bowel disease (IBD) precision therapy remains a critical barrier to clinical outcomes, with traditional studies focusing on single molecules or isolated pathways but failing to systematically dissect the bidirectional, context-dependent dynamic crosstalk among gut microbiota, immunity, and epigenetic modifications. This review dissects resistance mechanisms of key biologics (e.g., antitumor necrosis factor-α agents, vedolizumab) and small-molecule drugs (e.g., janus kinase inhibitors), proposing and validating the "triple-loop hierarchical regulation model"-gut microbiota dysbiosis as the initiator, immune dysregulation as the amplifier, and epigenetic maintenance as the stabilizer, with a bidirectional feedback loop sustaining resistance. It identifies the interaction network as the central regulatory axis, outlining a cascade where altered microbiota composition, signature metabolites (e.g., short-chain fatty acids), epigenetic modification (e.g., acetylation), and Th17/Treg imbalance may collectively contribute to the emergence of drug-resistant phenotypes. Four distinct subtypes (immunogenic, metabolic, epigenetic, barrier-deficient) are defined, with targeted strategies: precise microbiota regulation (e.g., fecal microbiota transplantation), immunity-epigenetics intervention (e.g., histone deacetylase inhibitors), and synergistic schemes that could partially rescue resistant clinical presentations. It also discusses multi-omics biomarkers for early prediction and formulation translation challenges, emphasizing cutting-edge technologies (single-cell multi-omics, organoid-microbiota co-cultures) and interdisciplinary collaboration. This review provides a comprehensive framework for overcoming IBD drug resistance and advancing personalized therapies.

Patients with inflammatory bowel disease are at increased risk of cardiovascular disease, yet the mechanisms linking chronic intestinal inflammation to cardiac dysfunction remain poorly understood. Inflammatory bowel disease is characterized by profound gut microbiota dysbiosis, which we hypothesize drives systemic immune dysregulation and contributes to cardiac dysfunction. A chronic colitis mouse model was used to assess gut microbiota dysbiosis, systemic immune cell metabolism, and cardiac remodeling. Cardiac outcomes were evaluated by echocardiography, histology, and molecular analyses. Mechanisms were examined using fecal microbiota transplantation, immune cell depletion, exosome transfer, bone marrow chimeras, RNA sequencing, coimmunoprecipitation, confocal microscopy, and siRNA-mediated gene silencing. Chronic dextran sulfate sodium colitis induced cardiac dysfunction, hypertrophy, and fibrosis in mice. These changes were accompanied by sustained gut microbiota dysbiosis, metabolic reprogramming, and mitochondrial dysfunction in circulating immune cells. Fecal microbiota transfer experiments demonstrated that colitis-associated microbiota were sufficient to reprogram systemic immune cells and promote cardiac dysfunction. Immune cell depletion studies identified macrophages as key mediators of colitis-associated cardiac injury. Colitis increased systemic lipopolysaccharide translocation; bone marrow chimera experiments demonstrated that hematopoietic TLR4 (toll-like receptor 4) signaling was required for immune cell metabolic remodeling and cardiac dysfunction during chronic colitis. Transcriptomic analysis identified GBP2b (guanylate-binding protein 2b/GBP1, hereafter referred to as GBP1) as a key downstream effector of lipopolysaccharide TLR4 signaling. Upon lipopolysaccharide stimulation, GBP1 localized to mitochondria, where it interacted with DRP1 (dynamin-related protein 1) and FIS1 (fission 1 protein) to promote mitochondrial fission, oxidative stress, and enhanced immune cell migration into the heart. In addition, GBP1 was secreted via exosomes, which were taken up by cardiomyocytes and contributed to hypertrophic remodeling and cardiac dysfunction. These findings establish the lipopolysaccharide TLR4-GBP1 axis as a key driver of colitis-associated cardiovascular dysfunction and highlight this pathway as a promising therapeutic target for reducing cardiovascular risk in patients with inflammatory bowel disease.

Inflammatory bowel disease (IBD), which includes ulcerative colitis and Crohn's disease, represents a chronic, immune-mediated pathology in which genetic susceptibility, environmental exposures, and microbial perturbations converge to destabilize intestinal homeostasis. Despite a steady global rise in incidence affecting more than 10 million people worldwide, current single-pathway biologic and small-molecule therapies still leave 30-40% of patients without durable remission, with progression to stricturing, fistulizing, or transmural complications. Yet the mechanistic integration of innate and adaptive immune networks, epithelial barrier dysfunction, and microbiota‑driven inflammation remains fragmented. This review summarizes recent advances in understanding the pathophysiological mechanisms driving IBD, focusing on innate and adaptive immune networks. Current therapies, including cytokine or integrin-targeting biologics and small-molecule inhibitors like Janus Kinase (JAK) antagonists, are critically evaluated in light of their restricted pathway coverage. In response to these challenges, emerging strategies are examined targeting nonimmune pathways, including microRNA‑124-mediated gene regulation, fecal microbiota transplantation, and mesenchymal stromal cell therapies that combine immunomodulation with tissue repair. The development of effective IBD therapies is likely to benefit from combining complementary treatment strategies, guided by molecular and cellular profiling. This review underscores a paradigm shift from monotherapies to integrated, multitarget approaches as the new frontier in IBD management.

Excessive foraging of colonic mucin glycans by gut bacteria is associated with diseases such as inflammatory bowel disease. Although Akkermansia muciniphila is an important mucin degrader, the role of carbohydrate sulfatases that facilitate digestion of these heavily sulfated glycans remains unclear. Combining in vitro digestion assays, proteomics and structural biology, we show that A. muciniphila sulfatases, such as Amuc1755 and Amuc0953, have rare adaptations targeted towards known sulfated mucin structures. They show larger degrees of modularity, including a previously unknown mucin-binding domain. When grown on colonic mucin substrates, glycoproteins of reduced size were important for the growth of A. muciniphila. Further mutational analysis and localization studies revealed that desulfation of N-acetyl-D-glucosamine was periplasmic, while desulfation of D-galactose occurred extracellularly and in the periplasm. These data improve our understanding of contexts for the positive health correlations of A. muciniphila while metabolizing colonic mucin as its sole carbon source.

ATG16L1 (autophagy related 16 like 1) is a core macroautophagy/autophagy protein essential for autophagosome formation. It also functions in non-canonical autophagy pathways such as LC3-associated phagocytosis (LAP) and in other processes including immunity, inflammation, and membrane trafficking. This review synthesizes recent advances and proposes that ATG16L1 functions as a central molecular integrator governed by a multi-layered regulatory code. This framework includes genetic polymorphisms, transcriptional control, and diverse post-transcriptional and post-translational mechanisms. We detail how these regulatory layers collectively fine-tune ATG16L1 function in response to cellular stress. Dysregulation of this network contributes broadly to human diseases including inflammatory bowel disease, cancer, and neurodegenerative disorders. Notably, the functional impact of specific regulatory events is highly context dependent, a principle exemplified by the Crohn disease-associated T300A polymorphism. Deciphering this regulatory landscape and its crosstalk with both autophagy-dependent and autophagy-independent functions positions ATG16L1 as a pivotal node in cellular homeostasis and as an emerging therapeutic target.Abbreviations ATG: autophagy related; CASM: conjugation of Atg8-family proteins to single membranes; CCD: coiled-coil domain; CEBPA/CEBPα: CCAAT enhancer binding protein alpha; CHUK/IKKA: component of inhibitor of nuclear factor kappa B kinase complex; circRNA: circular RNA; CPT1A: carnitine palmitoyltransferase 1A; CREB: cAMP responsive element binding protein; CSNK2: casein kinase 2; FTO: FTO alpha-ketoglutarate dependent dioxygenase; GJA8/connexin 50: gap junction protein alpha 8; H/R: hypoxia-reoxygenation; HDAC: histone deacetylase; KAT2B/PCAF: lysine acetyltransferase 2B; KDM1A: lysine demethylase 1A; LAP: LC3-associated phagocytosis; lncRNA: long non-coding RNA; LRRK2: leucine rich repeat kinase 2; m6A: N6-methyladenosine; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; miRNA/MIR: microRNA; Mtb: Mycobacterium tuberculosis; ncRNA: non-coding RNA; PE: phosphatidylethanolamine; PI3K: phosphoinositide 3-kinase; PRKA/PKA: protein kinase cAMP-activated; PPP1: protein phosphatase 1; RAB33B: RAB33B, member RAS oncogene family; RB1CC1/FIP200: RB1 inducible coiled-coil 1; SETD7: SET domain containing 7, histone lysine methyltransferase; SQSTM1/p62: sequestosome 1; TNF/TNF-α: tumor necrosis factor; ULK: unc-51 like autophagy activating kinase; V-ATPase: vacuolar-type H+-translocating ATPase; VDR: vitamin D receptor; WIPI2B: WD repeat domain, phosphoinositide interacting 2B; YTHDF2: YTH N6-methyladenosine RNA binding protein F2; ZDHHC7: zDHHC palmitoyltransferase 7.

Emerging insights into the gut microbiome have sparked interest in exploring microbial therapeutics for treating inflammatory bowel diseases (IBD). However, no microbial therapeutics have yet shown clinical efficacy for IBD. Escherichia coli Nissle 1917 (EcN), although effective for maintenance of remission, is only marginally effective for treating active colitis. We postulated that EcN effectiveness is hindered by the inflamed intestine, which prevents colonization because EcN lacks stress-resistance mechanisms necessary to persist during colitis. To address this, we introduced a fitness advantage, the ttr operon, to EcN (EcN::ttr), enabling tetrathionate, a byproduct of intestinal inflammation, to be used as fuel. We hypothesized that EcN::ttr bioengineered to bloom during colitis would effectively treat colitis. We evaluated the efficacy of EcN::ttr in murine colitis including an acute dextran sodium sulfate model and a chronic mucin 2-deficient model. To determine the role of interleukin (IL) 10 in EcN::ttr protection, we tested its efficacy in IL10-deficient mice. Finally, we co-incubated EcN::ttr with human colonoids to understand its effect on barrier proteins. EcN::ttr ameliorated colitis more effectively than EcN and 5-aminosalicylate. EcN::ttr bloomed during inflammation and promoted immunoregulatory responses reliant on IL10 that limited leukocyte infiltration and decreased tumor necrosis factor-α+ myeloid resident cells. EcN::ttr induced functional changes in the gut microbiome related to mucosal healing, increased butyric acid, reduced bacterial translocation, and improved zonula occludens-1 organization. We provide a proof-of-concept study that bioengineering ttr into EcN unlocks a robust therapeutic effect during colitis. EcN::ttr may be a novel microbiome therapeutic for IBD due to its enhanced ability to successfully colonize the inflamed gut.

Inflammatory bowel disease (IBD) is a chronic inflammatory disorder of the gastrointestinal tract that presents a significant global health challenge. Although its exact etiology remains unclear, growing evidence indicates that dysregulation of the complex regulatory network of serine proteases and their inhibitors is a major contributor to IBD pathogenesis. This imbalance leads to both pro-inflammatory and protective effects within the inflamed intestine, highlighting the need to better understand the underlying mechanisms. This review synthesizes recent advances in our understanding of the roles that host-derived serine proteases and their inhibitors play in intestinal inflammation. We assess current therapeutic approaches targeting this network, and explore the potential for developing novel serine protease inhibitors and more effective drug delivery systems. The review discusses the pathophysiological roles of serine proteases and serpins, highlighting their dual functions in both promoting and regulating intestinal inflammation. It then evaluates existing and emerging therapeutic strategies that modulate this enzymatic network for the treatment of IBD. Finally, the review discusses future directions, emphasizing the development of next-generation inhibitors and the critical need for translational research to accelerate the clinical application of these promising therapies.

Inflammatory bowel disease (IBD) and its frequent hepatic complication, metabolic-associated steatohepatitis (MASH), are intrinsically linked through the dysregulated gut-liver axis, with intestinal barrier dysfunction serving as a central pathological driver. Coordinated therapy that restores barrier integrity and interrupts pathogenic gut-liver crosstalk remains a significant challenge. To address this, a colon-targeted delivery system based on sodium alginate microspheres is developed for loading copper-kaempferol nanocomplexes (CuK@SA). Upon oral administration, this system prolongs the retention and release of CuK NCs in the colon. Through scavenging reactive oxygen and nitrogen species (ROS/RNS), regulating macrophage polarization, improving microbial homeostasis, and enhancing tight junction protein expression, it multi-dimensionally restores intestinal barrier integrity and effectively blocks the translocation of endotoxins to the liver via the gut-liver axis. In mouse models of dextran sulfate sodium (DSS)-induced colitis and high-fat-diet-induced MASH, CuK@SA significantly alleviates intestinal inflammation, repairs barrier structure, and simultaneously improves hepatic steatosis and inflammatory responses. This work provides a novel strategy for synchronously targeting IBD and its systemic complications through modulation of the gut-liver axis.

In individuals undergoing total pancreatectomy, an additional islet autotransplantation (total pancreatectomy with islet autotransplantation [TPIAT]) can be performed. We investigated the degree to which islet secretory function is preserved after TPIAT and assessed the relation to glycaemic control using continuous glucose monitoring. Eligibility for TPIAT was assessed by a multidisciplinary team. The cohort consisted of all individuals undergoing TPIAT from 2014 to 2024. To assess islet secretory function, participants were subjected to a 2 h liquid meal stimulation test prior to TPIAT, at 3 months and annually post TPIAT. Glycaemic control was assessed using continuous glucose monitoring and HbA1c. Twenty-six individuals were included, of whom 88.5% had chronic pancreatitis. At baseline, 57.7% had prediabetes (HbA1c 39-47 mmol/mol [5.7-6.4%] or a fasting glucose 5.6-6.9 mmol/l) or diabetes, and 46.2% had undergone previous pancreatic surgery. Islet secretory function determined by C-peptide area under the curve (AUCC-peptide) was 45% of baseline at 3 months post TPIAT. Preservation of islet secretory function was independent of preoperative glycaemic status and previous surgery. At 3 months, time in range was 75.2 ± 26.1%, and time in range was positively correlated with AUCC-peptide (p<0.0001). After 3 months islet function did not significantly change, which was reflected in a stable BETA-2 score. Nearly half of islet secretory function is retained after TPIAT. These results can facilitate shared decisions on the potential benefits of islet autotransplantation after total pancreatectomy, even in individuals with (pre)diabetes and previous pancreatic surgery.

Acute Pancreatitis (AP) is a common gastrointestinal emergency in which early fluid therapy is critical, yet optimal strategies remain debated. Although Hematocrit (HCT) and Blood Urea Nitrogen (BUN) are recommended to guide fluid management, the prognostic value of their dynamic changes is unclear. This multicenter retrospective cohort study utilized a development cohort from Jiangxi Province (n = 2027) and a validation cohort from the U.S. MIMIC-IV and eICU-CRD databases (n = 1381). Group-based multi-trajectory modeling (GBMTM) was applied to analyze the dynamic changes of HCT and BUN during the first week of ICU admission to identify distinct subphenotypes. Multivariate logistic regression and survival analysis were used to assess the association between each subphenotype and outcomes, including mortality and organ failure. The Boruta algorithm compared predictive importance. Five trajectory subphenotypes were identified: T1 (Renal Dysfunction Subphenotype, n = 287), T2 (Fluid-Responsive Subphenotype, n = 448), T3 (Volume-Deficient Subphenotype, n = 445), T4 (Stable Subphenotype, n = 516), and T5 (Hemodilution Subphenotype, n = 331). The T1 subphenotype had the highest mortality (36.6 % in the development cohort; adjusted OR = 16.27 (95 % CI 6.55-40.45) for in-hospital mortality in the validation cohort), while T4 had the lowest (2.7 %). Subphenotypes exhibited significantly different responses to fluid therapy: for T1, a fluid volume exceeding 4000 mL on the first day increased mortality risk; T3 tolerated a higher initial fluid load (up to 7000 mL on day one) but required fluid restriction on the second day. Consistent results in subgroup and sensitivity analyses demonstrated the robustness of this classification system. This study developed and validated a subphenotype classification system for the acute phase of AP based on the dynamic trajectories of HCT and BUN. It effectively distinguishes patient prognosis and responsiveness to fluid therapy, providing a crucial phenotypic framework for future randomized trials on individualized fluid management.

Biodegradable pancreatic stents are a potential alternative to conventional plastic stents for preventing post-endoscopic retrograde cholangiopancreatography (ERCP) pancreatitis (PEP), but comparative evidence remains limited. This multicenter study compared PEP incidence between biodegradable and plastic pancreatic stents using adjusted analyses to minimize confounding. This UK multicenter retrospective study included adults receiving biodegradable or plastic pancreatic stents between January 2020 and September 2025. Baseline differences were addressed using propensity score-based inverse probability of treatment weighting (IPTW). The primary outcome was PEP, analyzed using IPTW-adjusted risk differences with bootstrap confidence intervals. 355 patients were included: 177 received plastic stents and 178 received biodegradable stents. PEP occurred in 6.2% (11/177) and 7.3% (13/178), respectively. After IPTW, balance improved for most baseline variables, although procedure year remained imbalanced, reflecting temporal adoption of biodegradable stents. The adjusted PEP risk difference was +3.59% (95%CI -3.27 to +11.37), with no significant difference between groups. Repeat endoscopy within 3 months occurred in 45.2% (80/177) of plastic stent recipients and 0% (0/178) of biodegradable stent recipients (IPTW-adjusted risk difference -51.13%; 95%CI -60.61 to -40.86). In this largest multicenter retrospective study, biodegradable pancreatic stents were not associated with a significant difference in PEP risk compared with plastic stents after adjustment for confounders. Residual confounding remained possible, particularly due to temporal and procedural factors. Repeat endoscopy for stent removal was eliminated in the biodegradable group. Larger prospective multicenter studies are needed to evaluate the clinical and economic impact.

Direct endoscopic necrosectomy (DEN) is commonly performed for walled-off necrosis (WON), yet its optimal timing remains uncertain. Step-up strategies defer necrosectomy until clinical deterioration, whereas immediate DEN may provide earlier source control, but risks unnecessary intervention. We conducted a systematic review and meta-analysis of randomized controlled trials (RCTs) comparing immediate versus step-up DEN. Major databases were searched through to 26 April 2026 for RCTs comparing immediate versus step-up DEN. The primary outcome was clinical success. Secondary outcomes included technical success, necrosectomy sessions, total interventions, length of stay, adverse events, bleeding, organ failure, mortality, recurrence, and readmission. Random-effects meta-analysis was performed using the Hartung-Knapp adjustment. Certainty of evidence was assessed using Grading of Recommendations Assessment, Development and Evaluation (GRADE). 5 randomized trials, including 269 patients (134 immediate DEN; 135 step-up strategy), were analyzed. In four trials reporting standalone study-defined clinical success, immediate DEN was associated with higher success than the step-up strategy (odds ratio [OR] 2.56, 95%CI 1.03 to 6.37; P = 0.046; I 2 = 0%). A post hoc sensitivity analysis including the ACCELERATE trial yielded a similar estimate (OR 2.59, 95%CI 1.33 to 5.04; P = 0.02; I 2 = 0%). Immediate DEN was associated with a shorter length of stay (mean difference -8.02 days, 95%CI -15.00 to -1.05; P = 0.03; I 2 = 42%). Technical success, necrosectomy sessions, total interventions, adverse events, bleeding, organ failure, mortality, and readmission did not differ significantly. Immediate DEN was associated with higher clinical success and shorter hospital stay compared with a step-up strategy. These findings support selective consideration of immediate DEN in appropriate patients with WON, while the timing remains individualized.

[This corrects the article DOI: 10.1002/mco2.70350.].

Acute pancreatitis (AP) is a life-threatening inflammatory disease with a rising incidence in which intestinal barrier dysfunction and bacterial translocation contribute to severe disease progression. Short-chain fatty acids (SCFAs), key microbial metabolites, have been linked to intestinal health, however, their role in AP and the underlying mechanisms remain unclear. This study aimed to investigate whether SCFAs alleviate AP by restoring intestinal barrier function and modulating inflammation, with a focus on the gut microbiota and the interleukin-17A (IL-17A) pathway. Mouse AP models were induced using caerulein, and mice were treated with exogenous mixed SCFAs, IL-17A/IL-17RA inhibitors, or recombinant IL-17A (rIL-17A). Germ-free mice, antibiotic-treated mice, and fecal microbiota transplantation (FMT) were used to assess the role of the gut microbiota. Fecal SCFA levels, pancreatic/intestinal histopathology, barrier function markers (Occludin, claudin-1, and D-lactate), inflammatory cytokines, and the IL-17A pathway were analyzed via GC-MS, immunohistochemistry, Western blotting, RNA sequencing, and 16S rRNA gene sequencing. Fecal SCFA levels were reduced in AP mice. Exogenous mixed SCFAs ameliorated pancreatic injury, decreased serum amylase/lipase and proinflammatory cytokines (IL-1β, TNF-α, and IFN-γ), and restored intestinal barrier integrity (increased villus length, mucus layer thickness, and tight junction proteins). The protective effects of SCFAs were abrogated in germ-free or antibiotic-treated mice, but FMT from SCFA-treated donors recapitulated these benefits, supporting a contribution of the gut microbiota to SCFA-mediated protection. 16S rRNA sequencing revealed that SCFAs increased Bifidobacterium pseudolongum (B. pseudolongum) abundance, and B. pseudolongum supplementation alone mitigated AP severity. RNA sequencing and functional assays revealed that SCFAssuppressed the intestinal IL-17A/IL-17RA pathway, reducing the Th17 cell proportion and IL-17A-driven inflammation. The inhibition of IL-17A or IL-17RA mimicked the protective effects of SCFAs, whereas rIL-17A exacerbated AP, which was reversed by SCFAs. SCFAs alleviate AP by improving intestinal barrier integrity and suppressing inflammation, with the gut microbiota contributing, at least in part, to these protective effects. These effects involve gut microbiota remodeling, enrichment of B. pseudolongum, and attenuation of the IL-17A/IL-17RA-related signaling. These findings support a microbiota-associated SCFA-IL-17A regulatory axis in AP and suggest that IL-17A-related signaling warrants further investigation as a potential therapeutic target.

Screening endoscopy can be spared in patients with compensated cirrhosis when spleen stiffness measurement (SSM) by vibration-controlled transient elastography (VCTE) is ≤40 kPa, as they have a low probability of high-risk varices (HRV). Conversely, endoscopy is required in all patients with chronic portal vein thrombosis (PVT) without cirrhosis. The objective was to evaluate the performance of SSM-VCTE to exclude HRV in patients with chronic PVT. We retrospectively included patients with chronic PVT without cirrhosis, who underwent an upper endoscopy within 2 years before or after SSM-VCTE in 16 VALDIG centers, divided into a derivation and a validation cohort. 159 patients were included in the derivation cohort; 43% had HRV. 187 patients were included in the validation cohort; 32% had HRV. By univariable analysis, myeloproliferative neoplasm, ascites, hemoglobin, bilirubin, albumin, splenomegaly, portosystemic collaterals, liver stiffness - spleen diameter to platelet ratio score, liver stiffness measurement and SSM-VCTE were associated with HRV in both cohorts. By multivariable binary logistic regression analysis, only SSM-VCTE (p <0.005) remained associated with HRV in both cohorts. In the derivation cohort, SSM-VCTE ≤ 40 kPa had a sensitivity of 97% to rule out HRV, and could spare 41% of endoscopies, with 3% of HRV missed, and a 97% negative predictive value (NPV). In the validation cohort, SSM-VCTE ≤ 40 kPa could spare 43% of endoscopies, with 5% of HRV missed, and a 96% NPV. This study gathering a total of 346 patients with chronic PVT without cirrhosis showed that SSM-VCTE ≤ 40 kPa can be used to identify patients with a probability of HRV ≤5%, in whom endoscopy can be spared. Patients with chronic portal vein thrombosis who do not have cirrhosis usually have low liver stiffness measurement values; the liver stiffness cut-offs used to rule out high-risk varices in patients with cirrhosis cannot therefore be used in this population. We show here that spleen stiffness measurement by vibration-controlled transient elastography ≤40 kPa is able to identify patients with chronic portal vein thrombosis with a very low probability of high-risk varices, in whom screening endoscopy can be spared. Annual surveillance of spleen stiffness measurement could reduce the need for repeated screening endoscopies throughout a person's lifetime. This approach could enhance quality of life while also reducing risks associated with endoscopic procedures, particularly those related to anesthesia.

Clinically significant portal hypertension (CSPH) is the main driver of hepatic decompensation, and its early identification allows timely initiation of preventive therapies. Hepatic Venous Pressure Gradient (HVPG) is the current gold standard for assessing portal hypertension (PH), but it may underestimate PH in conditions with a presinusoidal component. Endoscopic ultrasound-guided portal pressure gradient (EUS-PPG) enables direct measurement of portal pressure and may overcome these limitations. We evaluated the prognostic performance of EUS-PPG compared with HVPG for predicting hepatic decompensation in patients with suspected CSPH. This preliminary exploratory analysis of the ongoing prospective EVADIPP study included 90 patients who underwent paired HVPG and EUS-PPG measurements and were followed for decompensation. Mean EUS-PPG and HVPG values were 13.8 ± 5.8 mmHg and 8.9 ± 4.8 mmHg, respectively, with poor overall agreement (ICC 0.08, 95% CI -0.08 to 0.24). Agreement remained poor in porto-sinusoidal vascular disorder (PSVD), slight in metabolic dysfunction-associated steatotic liver disease, and substantial in alcohol- and viral-related liver disease. During follow-up, 28 patients (31%) developed decompensation; no events occurred among patients with EUS-PPG<10 mmHg, whereas 60.7% had HVPG <10 mmHg. In multivariable analysis, EUS-PPG (HR 1.19, 95% CI 1.10-1.30; p<0.001) and albumin were independently associated with decompensation. EUS-PPG demonstrated better discrimination than HVPG (C-index 0.78 vs 0.56), and time-dependent ROC analysis identified an optimal threshold of 12 mmHg. EUS-PPG is independently associated with hepatic decompensation and showed better prognostic discrimination than HVPG in this exploratory cohort.

Liver sinusoidal microthrombosis (LST) is recognized as an initiating event in fibrogenesis and portal hypertension in chronic liver diseases. Liver sinusoidal endothelial cell (LSEC)-derived chemoattractants recruit neutrophils and macrophages, promoting LST in congestive hepatopathy (CH). However, the driving molecules from LSECs for LST remain unclear. This study aims to elucidate that LSECs inflammatory via cyclooxygenase-2 (COX-2) upregulation triggers metabolic reprogramming promoting thrombospondin-1 (TSP-1)-mediated LST and portal hypertension. A murine model of LST and portal hypertension was established by partial ligation of inferior vena cava (pIVCL). LST and portal hypertension were suppressed in both LSEC-specific COX-2 knockout mice (Ptgs2ΔLSEC) and celecoxib-treated wild-type mice induced by pIVCL. RNA sequencing of mouse liver tissue and untargeted metabolomics of human hepatic sinusoidal endothelial cells (HHSECs) revealed that COX-2 inhibition was concurrent with downregulation of the AKT/mTOR pathway, reduced lactate, and decreased TSP-1. In vitro, COX-2-derived prostaglandin E2 (PGE2) activated the AKT/mTOR pathway, driving glycolytic reprogramming and lactate production. In turn, the accumulation of lactate induced by COX-2 upregulation enhanced histone H3K9 lactylation, which transcriptionally upregulated Thbs1 (encoding TSP-1), thereby instigating a prothrombotic phenotype in LSECs. Collectively, this study uncovers a novel pathogenic axis in LST formation, where COX-2 drives a prothrombotic switch in LSECs via AKT/mTOR-mediated metabolic reprogramming and lactate-dependent epigenetic upregulation of TSP-1. Targeting LSEC COX-2 may represent a promising therapeutic strategy for mitigating LST and portal hypertension.

Hepatopulmonary syndrome (HPS) is a condition characterized by pulmonary angiogenesis and refractory hypoxemia, often seen in patients with chronic liver disease. Its unclear mechanism means that liver transplantation is the only effective therapy. Agrimoniin, a compound from Pilosa ledeb, shows potential in protecting against liver cirrhosis via anti-angiogenic and anti-glycolytic effects. This study investigates agrimoniin as a potential integrated therapy for HPS-related liver and lung dysfunction. Using transcriptome data and an ICU cohort, we analyzed the role of glycolysis in chronic liver disease progression. HPS rats were established via common bile duct ligation, and serum metabolites were measured. The oxygen consumption rate and extracellular acidification rate were also detected. Rats were treated with agrimoniin (3 mg/kg/day or 8 mg/kg/day) at the early stage of HPS. Our results showed that imbalanced oxidative phosphorylation and glycolysis correlated with chronic liver disease progression and poorer outcomes. Decreased oxygen consumption rate and increased extracellular acidification rate, as well as increased glycolysis, were observed in the HPS group. Agrimoniin treatment improved liver and lung function by inhibiting pathological angiogenesis and glycolysis. Through TCM suite analysis, molecular docking, and dynamics simulations, PGC-1α was identified as a potential target of agrimoniin. Inhibiting PGC-1α blocked agrimoniin's benefits on angiogenesis and glycolysis flux. Thus, agrimoniin may be a potential integrated therapy for HPS by activating PGC-1α to inhibit glycolysis and angiogenesis.

Microbiome-derived deoxycholic acid (DCA) elevates serum 5-hydroxytryptamine (5-HT), a mediator of portal hypertension (PH). Rifaximin, a non-absorbable antibiotic, is known to reduce DCA levels. We aimed to elucidate the role of DCA in cirrhotic PH and evaluate the therapeutic potential of rifaximin. PH was induced in mice by thioacetamide (TAA) injection or bile duct ligation (BDL). Mice were treated with antibiotics (ABX) or rifaximin, with or without exogenous DCA supplementation. A cohort of 51 patients with cirrhosis and 19 healthy controls was analyzed to validate correlations among DCA, 5-HT, and hepatic venous pressure gradient (HVPG). Mice with tissue-specific knockout of gut epithelial Tph1 (Tph1VKO), vascular smooth muscle cell Htr1a (Htr1aΔVSMC), or Kcnj9 (Kcnj9ΔVSMC) were used for mechanistic studies. Fecal DCA positively correlated with portal pressure (PP) in TAA-induced PH mice (r = 0.631, p <0.001) and with HVPG in patients (r = 0.5874, p <0.001). ABX treatment reduced fecal DCA, serum 5-HT, and PP in TAA- or BDL-induced PH mice. Exogenous DCA reversed the ABX-induced reductions in serum 5-HT and PP, an effect abolished in Tph1VKO mice. GIRK3 (encoded by Kcnj9) was upregulated in portal veins from PH mice and patients. VSMC-specific Kcnj9 deletion attenuated PH and prevented 5-HT-induced PP elevation. Mechanistically, 5-HT triggered portal vein smooth muscle cell contraction via the HTR1A-GIRK3-Ca2+-MLC2 pathway. Rifaximin alleviated PH by reducing DCA in wild-type mice but showed no additional PP reduction in Tph1VKO, Htr1aΔVSMC, or Kcnj9ΔVSMC mice. Microbiome-derived DCA exacerbates PH by enhancing TPH1-dependent 5-HT biosynthesis, which activates portal vein smooth muscle cell contraction via HTR1A-GIRK3 signaling. Rifaximin alleviates cirrhotic PH by reducing DCA levels, highlighting a potential therapeutic strategy for clinical PH management. Portal hypertension is a key driver of cirrhosis-related complications, yet current therapies exhibit suboptimal efficacy. Herein, we elucidate the role of the gut microbial metabolite deoxycholic acid in the pathophysiology of cirrhotic portal hypertension through the TPH1-5-HT/HTR1A-GIRK3 axis and provide preclinical evidence that rifaximin ameliorates cirrhotic portal hypertension by reducing deoxycholic acid. These insights may open a new avenue for the clinical management of cirrhotic portal hypertension.

Aberrant activation of the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) pathway has been increasingly recognized as a key driver of autoimmune and inflammatory diseases. In recent years, accumulating evidence has highlighted the close association between cGAS-STING signaling and the pathogenesis of these disorders, suggesting that pharmacological targeting of this pathway may represent a promising therapeutic strategy. This review provides a comprehensive overview of the molecular mechanisms underlying cGAS-STING hyperactivation and its pathological roles across diverse diseases, including systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), non-alcoholic steatohepatitis (NASH), STING-associated vasculopathy with onset in infancy (SAVI), Aicardi-Goutières syndrome (AGS), COPA syndrome, Niemann-Pick disease type C (NPC), neurodegenerative diseases and cancer. We critically evaluate current and emerging pharmacological strategies targeting the cGAS-STING pathway, encompassing direct cGAS and STING inhibitors, protein degradation technologies, epigenetic modulation, regulation of biomolecular phase separation, and artificial intelligence (AI)-enabled drug discovery approaches. By integrating disease-specific pathogenic mechanisms with therapeutic opportunities, this review highlights key challenges and future directions in the development of cGAS-STING-targeted therapies. Collectively, these insights provide a translational perspective for precision targeting of pathological cGAS-STING activation.

In group decision-making (GDM) involving decision-makers (DMs) with heterogeneous interests and responsibilities, such as transboundary watershed governance, consensus formation is fundamentally return-driven. This study develops a directionally asymmetric maximum-return consensus model (MRCM) with nonnegative return constraints, which shifts from a moderator-cost perspective to an individual-return perspective. To this end, a feasibility diagnosis approach integrating the minimum-slack feasibility checking model (MSFCM) and the relaxed MRCM is proposed to determine whether the MRCM is feasible. When it is not feasible, it further reveals whether the infeasibility originates from individual- or collective-level return deficits. Based on the diagnosis results, a twofold adaptive consensus framework is further developed: 1) internal compensation-driven feedback is applied when individual return deficits coexist with a nonnegative total cooperative return, reallocating surplus via an asymmetric Nash bargaining game (ANBG) without modifying the relaxed consensus outcome and 2) external compensation-driven feedback is activated when the total cooperative return is negative, with the moderator providing the minimum compensation to ensure consensus with nonnegative returns. The novelty of this work lies in developing a unified return-driven consensus mechanism governed by feasibility diagnosis by refining the new return formulation and compensation scheme. A numerical study based on the Dongjiang River Basin demonstrates that the proposed framework adaptively selects compensation strategies and effectively enhances consensus feasibility and stability.

Reinforcement learning (RL), a key artificial intelligence technique, has been widely studied and applied over the past two decades to solve various optimization control problems. Generally speaking, there are two basic frameworks for RL-based control design, i.e., on-policy and off-policy RL (OffP-RL). The essential distinction between the two frameworks lies in whether the policy used to generate training data is the behavior policy or the target policy. In on-policy RL-based control methods, the data used for evaluating the target control policy at each iteration must be collected from the system under the target policy itself. In contrast, in OffP-RL methods, the system data is generated by other behavior control policies. It addresses the inadequate exploration problem in on-policy RL methods, making OffP-RL methods more practical and easier to implement. In this article, the recent advances in OffP-RL-based control methods are classified into three categories based on the number of controllers/players involved, i.e., single-/two-/multiplayer. For the single-player case, it is an optimal control problem, which aims to use OffP-RL to learn the optimal control policy, which minimizes the performance index. In the two-player case, most works focus on the $H_{\infty } $ control problem and the two-player zero-sum game, using learning to find the Nash equilibrium. In the multiplayer case, a multiplayer game involves a single system with multiple control inputs, while a multiagent system consists of multiple systems with independent control inputs. Finally, related applications of OffP-RL-based control and future work are analyzed.

This article investigates the problem of prescribed-time Nash equilibrium (NE) seeking for a multicluster pursuit-evasion game (PEG) subject to external disturbances. To mitigate the impact of disturbances and reach the NE within a user-defined prescribed time, a prescribed-time disturbance observer (PTDO) is devised to estimate and compensate for them. Based on this observation, a novel control algorithm is developed, which facilitates collaboration among multiple pursuers to capture multiple evaders within the prescribed time. It is theoretically demonstrated that the designed algorithm ensures prescribed-time convergence to the NE of the multicluster PEG with disturbances. Finally, numerical simulations are conducted to verify the effectiveness of the algorithm under different initial conditions, further demonstrating the flexibility of the user-specified convergence time.

This article investigates the distributed formation control of uncrewed surface vehicles (USVs) under aperiodic denial-of-service (DoS) attacks within a Stackelberg-Nash game (SNG) framework. An actor-critic (AC) reinforcement learning (RL) algorithm is developed to approximate these policies online, ensuring convergence to the Stackelberg-Nash equilibrium (SNE). To enhance resilience against communication interruptions, a consensus-based estimator is designed to reconstruct missing neighbor data using local information. Rigorous Lyapunov-based analysis guarantees the input-to-state stability (ISS) of the estimator and the semi-globally uniformly ultimately bounded (SGUUB) stability of the closed-loop system. Simulation results verify the framework's effectiveness in achieving accurate trajectory tracking and robustness against frequent DoS attacks.

Vegetables anchor healthy-diet recommendations yet are the principal nonoccupational route by which residues of organophosphate (OP), pyrethroid, neonicotinoid, organochlorine (OC), and glyphosate herbicide enter the diet, with the gut microbiota a plausible mediator linking these residues to functional gastrointestinal (GI) and inflammatory bowel outcomes. We conducted a scoping review following the PRISMA extension for Scoping Reviews (PRISMA-ScR), searching PubMed (2010 to April 2026) with mandatory backward and forward citation tracing for human studies of dietary or biomarker-confirmed pesticide exposure against gut microbiota, intestinal barrier and inflammation, functional GI disorders, or inflammatory bowel disease (IBD). Risk of bias (RoB) was appraised with ROBINS-E and Cochrane RoB 2.0, and evidence was synthesized in a 60-cell pesticide-class × outcome gap matrix. Twenty-eight papers were retained; 20 full-text-extracted human studies entered the synthesis, populating 20 of 60 cells (33%). The strongest convergent within-scope signal was OC exposure × IBD, where three cohorts agreed in direction (hazard ratios 1.56-1.61; Faroese risk ratio 3.04 on 37 cases). As a microbiota-mediation exemplar for an adjacent non-GI endpoint, the MCMC Nanjing pregnancy cohort linked a pesticide environmental risk score to gestational diabetes (adjusted odds ratio 4.5; 95% CI, 1.4-13.8; internal-validation area under the curve 0.83), with a Dorea-branch microbiota mediating part of the path. Glyphosate × IBD and neonicotinoid × any GI outcome remained empty. RoB centered on Moderate (11 of 22 rated cohorts), with six elevated-bias contributors. Biomarker-anchored prospective cohorts targeting the empty cells should be the next research priority.

Pyoderma gangrenosum (PG) is a neutrophilic dermatosis frequently associated with systemic comorbidities such as inflammatory bowel disease (IBD), arthritis, and hematologic disorders. Management remains challenging due to heterogeneous presentations and treatment responses. To propose a comorbidity-guided therapeutic framework. A clinical review of clinical trials, case series, and real-world reports on PG management was conducted. Emphasis was placed on immunopathologic pathways linking PG with major comorbidities and on therapeutic strategies employing biologic and small-molecule agents tailored to these associations. Comorbidity-directed therapy, such as TNF inhibition for IBD-associated PG, IL-1 blockade for autoinflammatory syndromes, and Janus kinase (JAK) inhibition for arthritis overlap, resulted in higher healing rates and lower relapse risk across studies. Real-world cases demonstrate that individualized therapy addressing the underlying systemic drivers of skin disease may yield more durable ulcer healing than empiric therapy alone. A comorbidity-guided approach personalizes PG therapy, aligning dermatologic and systemic management to improve healing, minimize recurrence, and optimize patient outcomes.

Macrophage immunometabolism is an important regulator of inflammatory resolution and angiogenic responses. Aberrant macrophage polarization, accompanied by metabolic disturbances such as enhanced glycolysis and mitochondrial dysfunction, has been implicated in impaired angiogenesis in chronic inflammatory conditions. Cajaninstilbene acid (CSA), a natural stilbene derived from Cajanus cajan, exhibits anti-inflammatory, antioxidant, and metabolic regulatory activities. In non-diabetic inflammatory settings, including liver injury, ischemia-reperfusion injury, and inflammatory bowel disease, CSA promotes macrophage functional reprogramming and activates AMPK/Nrf2 signaling. Notably, these effects may facilitate a shift from glycolysis-associated inflammatory states toward oxidative phosphorylation-supported reparative macrophage states. VEGF, PDGF-BB, and SDF-1 are established mediators of macrophage-associated angiogenic responses. Their involvement in CSA-related regulation has yet to be examined. Several limitations warrant consideration. Direct experimental validation in disease-relevant contexts exhibiting sustained metabolic stress (e.g., diabetic wounds or chronic ischemia) has not yet been conducted. In addition, causal relationships between metabolic remodeling and macrophage phenotypic transition remain incompletely defined. The proposed involvement of CSA in exosomal miRNA regulation also awaits experimental confirmation. Throughout the article, experimentally supported findings are distinguished from mechanistic interpretation, with unresolved questions highlighted as priorities for future research. CSA may represent a potential modulator of immunometabolic dysfunction, although systematic validation in pathophysiologically relevant disease models is still required.

The microbiota shapes postnatal gut development and physiology. In the small intestine, epithelial-to-endothelial crosstalk governs the microbiota-induced remodeling of villus capillary networks essential for nutrient transport. The intestinal epithelial enzyme dual oxidase 2 (DUOX2), an established regulator of the microbiome-host interaction, exerts microbicidal functions through the generation of reactive oxygen species. However, its role in intestinal vascular development remains poorly understood. Here, we demonstrate a Toll-like receptor 2 (TLR2)-dependent regulatory pathway controlling DUOX2 expression that influences villus vascularization in the small intestine. Mice globally lacking DUOX2 activity exhibited a notable reduction in vascularization in the small intestine, accompanied by alterations in gut microbial community structure. Conversely, mice with an intestinal epithelial-specific deficiency of TLR2 displayed an increase in villus vascularization along with elevated expression levels of DUOX2. Notably, DUOX2 expression was strongly upregulated in intestinal epithelial biopsies from patients with Crohn's disease. Similarly, inflammatory conditions induced by dextran sulfate sodium (DSS) treatment in mice resulted in increased epithelial Duox2 expression accompanied by enhanced villus vascularization. Together, our findings suggest a microbiota-TLR2-DUOX2 signaling axis in intestinal epithelial cells that promotes villus vascularization. This mechanism links microbial sensing in the intestinal epithelium to structural remodeling of the villus microvasculature during homeostasis and inflammation.

Linerixibat (LYNAVOY®) is an orally administered reversible ileal bile acid transporter (IBAT) inhibitor developed by GSK for the treatment of cholestatic pruritus. In March 2026, linerixibat received its first approval for the treatment of cholestatic pruritus associated with primary biliary cholangitis (PBC) in adult patients in the USA. It is the first US FDA-approved therapy for this indication. Subsequently, linerixibat was approved in May 2026 in the UK for the treatment of cholestatic pruritus in adult patients with PBC. A regulatory review of linerixibat is currently underway in Canada, China and the EU for the treatment of cholestatic pruritus associated with PBC. This article summarizes the milestones in the development of linerixibat leading to these first approvals.

Drug development in primary biliary cholangitis (PBC) has led to the conditional approval of three second-line therapies, yet the transition to full regulatory approval remains challenging due to (1) the reliance on liver biochemistry and non-invasive measures of liver fibrosis as biomarkers of clinical outcomes, which are not accepted by regulatory authorities as validated efficacy endpoints; (2) the limited integration of real-world data and real-world evidence to complement clinical trials; and (3) the absence of harmonised and standardised use of patient-reported outcomes (PROs). To address these limitations, the European Association for the Study of the Liver (EASL) and the American Association for the Study of Liver Diseases (AASLD) commissioned an international, multi-stakeholder consensus panel to conduct a modified Delphi process to review current evidence, identify unmet research needs, and create a framework aligned with regulatory expectations to guide ongoing and future research in PBC. A total of 62 panellists, including clinicians, methodologists, regulators, industry partners, and patient representatives, were divided into three working groups and participated in regular online meetings, one in-person conference, and two online Delphi voting rounds. Response rates across the two voting sessions were 88% and 60%, respectively. The Delphi process resulted in agreement on 16 statements and 42 recommendations across the three thematic domains. Collectively, these consensus recommendations provide a comprehensive and pragmatic framework to support and expand the existing regulatory pathway for the development and approval of new therapies in PBC.

We report 104-week placebo-controlled data and long-term open-label extension (OLE) data from the ongoing ELATIVE® phase III trial (NCT04526665) of elafibranor in primary biliary cholangitis (PBC). 161 patients were randomized 2:1 to elafibranor 80 mg or placebo. The double-blind period (DBP) consisted of 52-week common (Part 1) and variable (Part 2) periods. Patients completing Part 1 continued into Part 2 until all patients completed Part 1, or for a maximum of 104 weeks. All patients completing Part 1 could enter the OLE and receive elafibranor. At Week 104 in the DBP Part 2, 64.3% (18/28) and 10.7% (3/28) of elafibranor-treated patients achieved biochemical response and alkaline phosphatase (ALP) normalization, versus no placebo-treated patients. In patients with moderate-to-severe fatigue or pruritus at baseline, mean (SE) changes to Week 104 in PROMIS Fatigue Short Form 7a (PFSF 7a) and PBC Worst-Itch Numeric Rating Scale (PBC WI NRS) were -6.3 (2.2) versus -0.4 (1.2) and -4.1 (1.0) versus 0.3 (1.2) with elafibranor versus placebo. 138 patients entered the OLE (continuous elafibranor: n=93; crossover elafibranor: n=45). In continuous patients at Weeks 104 and 156, 58.8% (47/80) and 65.0% (13/20) achieved biochemical response, and 15.0% (12/80) and 25.0% (5/20) achieved ALP normalization. In crossover patients, 51.2% (21/41) and 22.0% (9/41) achieved biochemical response and ALP normalization after 52 weeks. In continuous patients with baseline moderate-to-severe symptoms, mean (SE) changes to Week 130 in PFSF 7a and PBC WI NRS were -4.8 (1.5) and -4.0 (0.7). There were no unexpected safety findings. Through three years of treatment, elafibranor led to sustained biochemical improvements and was generally well tolerated, with numerical improvements in fatigue and pruritus. NCT04526665 (https://clinicaltrials.gov/study/NCT04526665); first registered 08/26/2020 IMPACT AND IMPLICATIONS: • Given the chronic, progressive nature of primary biliary cholangitis (PBC), the long-term efficacy and tolerability of treatments is important.• Here, we present two-year results from the double-blind period, and long-term data from the ongoing open-label extension of the phase III ELATIVE® trial, wherein elafibranor (a peroxisome proliferator-activated receptor-α/δ agonist) treatment led to sustained biochemical improvements, stable non-invasive tests of fibrosis, and numerical improvements in fatigue and pruritus, through three years.• Elafibranor demonstrated a favorable safety profile up to a maximum treatment exposure of 3.5 years, including in patients crossing over from placebo.• These findings support elafibranor's role as a durable long-term treatment for patients with PBC, and are particularly relevant for clinicians managing patients with inadequate response or intolerance to first-line treatments.

Pregnane X receptor (PXR), a nuclear receptor superfamily member, maintains bile acid homeostasis by regulating metabolic enzymes [e.g., cytochrome P450 3A4 (CYP3A4), uridine diphosphate glucuronosyltransferase 1A1 (UGT1A1)] and multidrug resistance protein 1 transporter, and alleviates liver/intestinal inflammation via inhibiting the nuclear factor kappa-B pathway, serving as a critical therapeutic target for cholestatic liver diseases and inflammatory bowel disease. In this study, we established a novel structure-based machine learning strategy to identify PXR agonists from a natural product database. With generated bioactive conformations binding with PXR, we integrated ligand-based and structure-based features to construct a comprehensive machine learning model using light gradient boosting machine. This model illustrated an R2 of 0.874 for the internal validation set and an R2 of 0.845 for the external test set, superior to the performance of other machine learning models, e.g. random forest regression, support vector regression, gradient boosting regression, K-nearest neighbors, and extreme gradient boosting. The model was used to predict the PXR agonistic activity of the candidate molecules screened out by the pharmacophore model. Promising candidates were selected out for further assay with HepG2 cell culture combined with a dual-luciferase reporter. Ultimately, natural products like schisantherin A, rhynchophylline, and irigenin were identified as potent PXR agonists, with half-maximal effective concentrations (EC50) of 1.58 μM, 2.57 μM, and 20.67 μM, respectively. These PXR agonists act as potential candidates for targeted therapies against PXR-related diseases. We anticipate that this work will provide support for the design and discovery of PXR modulators.

Reduced-frequency oral antiretroviral therapy may lessen treatment burden, but the exposure thresholds required to maintain virological suppression and the role of the HIV reservoir remain undefined. BETAF-RED was a 48-week, single-centre, randomised, open-label, controlled phase 2 pilot trial conducted at Hospital Clínic, Barcelona, Spain. Eligible participants were adults aged 18 years or older with HIV-1 RNA <50 copies per mL for at least 6 months while receiving once-daily bictegravir, emtricitabine, and tenofovir alafenamide. Key eligibility criteria included stable clinical condition and CD4 count >350 cells per μL; key exclusions included previous virological failure or documented resistance to study drugs, active hepatitis B or C infection, and conditions judged to jeopardise adherence. Participants were assigned 1:1:1:1 to receive the oral fixed-dose single-tablet regimen bictegravir, emtricitabine, and tenofovir alafenamide 50/200/25 mg once daily, three times weekly, twice weekly, or once weekly for 48 weeks. The primary endpoint was maintenance of HIV-1 RNA <50 copies per mL (virological success) at weeks 12 and 48, assessed by Food and Drug Administration Snapshot in the intention-to-treat exposed population, defined as all randomised participants receiving study drug, and in the prespecified on-treatment analysis. Secondary outcomes were virological failure and no virological data. Safety analyses included all participants who received at least one dose of study drug. This was not a non-inferiority study, and no non-inferiority margin was specified. ClinicalTrials.gov, NCT05602506. Between November 7, 2022, and July 3, 2023, 40 participants were enrolled; 39 received study medication. In the intention-to-treat exposed Food and Drug Administration Snapshot analysis, virological success was observed at week 12 in 9/9, 9/10, 10/10, and 8/10 participants in the once-daily, three-times-weekly, twice-weekly, and once-weekly arms, respectively. At week 48, virological success was observed in 8/9, 9/10, 9/10, and 7/10 participants in the once-daily, three-times-weekly, twice-weekly, and once-weekly arms, respectively. At week 12, virological failure and no virological data were observed in 0/9 and 0/9, respectively, in the once-daily arm; 1/10 and 0/10, respectively, in the three-times-weekly arm; 0/10 and 0/10, respectively, in the twice-weekly arm; and 2/10 and 0/10, respectively, in the once-weekly arm. At week 48, virological failure and no virological data were observed in 0/9 and 1/9, respectively, in the once-daily arm; 1/10 and 0/10, respectively, in the three-times-weekly arm; 0/10 and 1/10, respectively, in the twice-weekly arm; and 2/10 and 1/10, respectively, in the once-weekly arm. In the prespecified on-treatment analysis, virological success was observed in 36/37 participants at week 12 and in all 33 participants remaining on their assigned regimen at week 48; the only virological failure occurred at week 12 in the three-times-weekly arm, and no virological data were missing at either timepoint. All three participants with protocol-defined confirmed virological failure regained suppression after resuming once-daily therapy, without emergent resistance. Most participants met the primary endpoint of maintained virological suppression at weeks 12 and 48 in the intention-to-treat exposed FDA Snapshot analysis. Early protocol-defined confirmed virological failures occurred in the once-weekly arm, and one confirmed low-level viraemia event occurred in the three-times-weekly arm. Because the trial was not powered for formal between-arm comparisons, these findings should be interpreted descriptively. Once-weekly dosing is not supported as a maintenance strategy; twice-weekly and three-times-weekly dosing require confirmation in larger, adequately powered studies with close virological monitoring before any clinical role can be considered. Instituto de Salud Carlos III and Institut d'Investigacions Biomèdiques August Pi i Sunyer.

Hepatitis B virus (HBV) infection remains a severe global public health challenge, with hepatocellular carcinoma being a primary cause of HBV-related mortality. Occult HBV infection (OBI) represents a distinct type of HBV infection that has been increasingly linked to hepatocellular carcinoma development, yet the precise molecular mechanisms underlying this association remain poorly elucidated. Although HBV pre-S deletion mutations have been shown to enhance cell proliferation and contribute to hepatocarcinogenesis, the biological functions of other types of pre-S mutations, particularly point mutations, are still mostly unexplored. In our prior studies, we identified several high-frequency pre-S point mutations from OBI blood donors. Within this research, we systematically explored the effects of these OBI-associated pre-S mutations on host cell proliferation and assessed their potential oncogenic properties. Cell proliferation assays revealed that several pre-S mutations significantly enhanced the proliferative capacity of host cells. Mechanistically, five pre-S mutations (E39K, D44N, N98T, H128R, and I161T) activated the Akt/mTOR signaling cascade, up-regulated Cyclin D1 expression, and induced G1-to-S phase cell cycle progression. Further analyses suggested that the large HBV surface protein (LHBs) likely acts as the key mediator linking pre-S mutations to signaling activation and cellular proliferation. These findings provide novel mechanistic understandings of the oncogenic potential of pre-S point mutations in hepatocarcinogenesis and may facilitate the identification of high-risk individuals within OBI populations as well as the development of treatment strategies for hepatocellular carcinoma linked to HBV.

Hepatitis B virus (HBV) integration represents a major obstacle to curing HBV; however, the landscape of HBV integration and local immune response to transcriptionally active viral integration in children with chronic HBV infection remain unclear. Herein, we aimed to elucidate this landscape in this population. Genomic analyses using a probe-based capture strategy were performed on 18 children and 28 adults with chronic HBV infection. Spatial transcriptomics (ST) was performed on 12 children from our cohort and 3 adults from a public database. All patients were hepatitis B e antigen (HBeAg)-positive and treatment-naïve. Genomically, children exhibited significantly lower clonal expansion level of HBV-integrated hepatocytes than adults, despite comparable unique breakpoint counts. After adjusting for confounding variables, age was identified as an independent risk factor for total frequency of unique integration breakpoints (b = 3.22, P = 0.005). Spatially, ST revealed that spots with transcriptionally active viral integration exhibited a sparse distribution and accounted for a low proportion of all spots in children. Notably, at these spots, children showed reduced adaptive immune cells (e.g., CD8+ T cells) but increased innate components (myeloid cells, Kupffer cells, activated dendritic cells) and APC co-stimulation, whereas adults exhibited a uniform reduction of immune cell populations. Compared with adults, children exhibit lower clonal expansion of HBV-integrated hepatocytes and distinct immune profiles in response to transcriptionally active viral integration, offering new insights into their differing clinical course. Key Laboratory of Molecular Biology for Infectious Diseases (Ministry of Education).

Hepatitis A vaccination is used for routine prevention and outbreak control, but its effectiveness and population-level impact have not been comprehensively synthesised. We aimed to assess vaccine effectiveness, certainty of evidence, and population-level impact. In this systematic review and meta-analysis, we searched MEDLINE, Embase, Biological Abstracts, and CENTRAL for studies published from 1st January 1985 to 1st April 2026. Randomised controlled trials and observational studies with an unvaccinated comparison group were eligible if they assessed pre-exposure or post-exposure hepatitis A vaccination and reported effect estimates or sufficient data for their calculation. Estimates were pooled using a DerSimonian-Laird random-effects model. Risk of bias was assessed with RoB 2 and ROBINS-I, and certainty of evidence with GRADE. Ecological studies evaluating vaccine impact before and after implementation of paediatric vaccination programmes were analysed separately. The review was registered with PROSPERO (CRD420261308908). We included 21 publications in the meta-analysis and 19 studies in the population-level impact analysis. Pooled effectiveness of pre-exposure vaccination was 95·8% (95% CI 93·1-97·4; I2 = 29·9%), supported by moderate-certainty evidence. In paediatric populations, effectiveness was 97·5% (92·8-99·1; I2 = 20·1%) in randomised trials and 95·9% (93·0-97·6; I2 = 0·0%) in cohort studies. Effectiveness was 97·2% (93·8-98·8; I2 = 4·3%) for inactivated vaccines and 96·0% (93·2-97·6; I2 = 0·0%) for live attenuated vaccines. Pooled effectiveness of post-exposure vaccination was 96·9% (89·3-99·1; I2 = 89·7%), but certainty of evidence was very low. Ecological studies showed consistently high population-level impact, generally ranging from 80% to 100%. Hepatitis A vaccination provides very high protection in pre-exposure settings. Post-exposure vaccination may also be effective, but confidence in this estimate is limited by very low-certainty evidence and substantial heterogeneity. Population-level findings support the public health value of routine immunisation and suggest potential benefit in outbreak response. Cooperatio 31 fund, Health Sciences, Charles University, Prague, Czech Republic.

HDV requires HBsAg for viral assembly. The siRNA JNJ-73763989 (JNJ-3989) targets all HBV transcripts and reduces HBsAg in patients with chronic hepatitis B. We evaluated whether HBsAg reduction with JNJ-3989 leads to a decline in HDV RNA and ALT in patients with chronic hepatitis D (CHD). REEF-D, a two-part, phase II, randomized, double-blind, placebo-controlled study, enrolled 22 adults with CHD in Part 1. Participants were randomized 4:1 to receive JNJ-3989 (100 mg subcutaneously every 4 weeks) + nucleos(t)ide analogue (NA) for 144 weeks (active arm, n = 17) or for 96 weeks after 52 weeks of placebo + NA (deferred treatment arm, n = 5). During the 48-week follow-up period, NA therapy was continued. The primary endpoint was achievement of a ≥2 log10 IU/ml reduction of HDV RNA from baseline (or 10,000 IU/ml) at screening experienced unexpected ALT elevations, sometimes severe, with a return to baseline levels after discontinuation of JNJ-3989. Both HDV RNA reduction and ALT elevation are critical observations for a better understanding of HDV biology and the interplay between viral antigens and the infected hepatocyte. Despite the low sample size and majority of participants not completing the planned treatment course, these findings are important for caregivers and researchers developing novel treatment strategies for patients with chronic hepatitis D.

Chronic liver necroinflammation induced by hepatitis B virus (HBV) infection plays a major causative role in the development of end-stage liver diseases; however, mechanisms contributing to its initiation remain unclear. Analysis of the hepatic transcriptome from HBV-replication mice or HBV-infected patients revealed that significantly down-regulated mitochondrial oxidative phosphorylation function was the salient transcriptional feature at the early stage of liver inflammation compared with the stage without liver inflammation. In cell models, persistent HBV replication-induced progressive impairment of mitochondrial respiration resulted in increased reactive oxygen species (ROS) levels. We further discovered that HBV replication-induced ROS accumulation was essential for the up-regulation of nuclear factor erythroid 2-related factor 2 (Nrf2)-associated interleukin (IL)-6/IL-8 production, mediating the activation of Janus kinase 2 (Jak2)/signal transducer and activator of transcription (Stat3) signaling, and then the expression of downstream inflammatory genes. These observations were also identified in HBV-replication mice at the early stage of liver inflammation, which exhibited elevated hepatic oxidative stress, Nrf2 expression, IL-6 and IL-8 production, and Jak2/Stat3 activation, alongside hepatic inflammatory cell infiltration. In vivo, ROS scavenging with N-acetylcysteine (NAC) mitigated these effects. Our findings underscore the critical role of ROS-dependent Jak2/Stat3 pathway activation in the occurrence of HBV-induced liver inflammation, providing new insights into the pathogenesis of chronic hepatitis B.

The effect of computer-aided detection (CADe) on performance of endoscopists with different experience levels is not well understood. This study assessed whether CADe use promotes learning or deskilling. We performed a prospective, multicenter, registry-based, pragmatic clinical trial. The primary end point was change in the proportion of colonoscopies with detection of at least one polyp ≥5 mm (PDR-5). Each endoscopist performed colonoscopies in three successive phases: 1) before CADe exposure, 2) during CADe use, and 3) after CADe removal. To assess the impact of CADe on PDR-5, we compared phase 1 with phases 2 and 3, adjusting for patient and endoscopist characteristics and endoscopy performance measures. Generalized linear mixed models were used and presented according to endoscopist experience. 13 endoscopists (7 inexperienced, 6 experienced) performed 5013 colonoscopies; median patient age was 59 years (interquartile range 44-71) and 2703 (53.9%) were women. Patient sex and age were similar across the two experience groups, whereas indications differed (e.g. more inflammatory bowel disease in the inexperienced group). CADe temporarily increased PDR-5 among inexperienced endoscopists from phase 1 to 2 (31.9% [216/678] to 39.5% [257/651]; odds ratio [OR] 1.43, 95%CI 1.11-1.84). There was no significant change between phases 1 and 3 (31.9% [216/678] to 36.3% [173/476]; OR 1.03, 95%CI 0.79-1.34). There was no significant CADe effect on PDR-5 among experienced endoscopists between phases. PDR-5 of inexperienced endoscopists increased when CADe was used. In non-CADe-assisted colonoscopy, no upskilling or deskilling was observed following a period of CADe exposure.

Endoscopic retrograde cholangiopancreatography-guided transpapillary biliary drainage (ERCP-BD) is the standard for primary palliation of malignant distal biliary obstruction (MDBO), but endoscopic ultrasound-guided choledochoduodenostomy (EUS-CDS) has demonstrated improved technical success, efficiency, and safety in randomized trials. However, cost-effectiveness data are lacking. In this modeling study, we analyzed the cost-effectiveness of EUS-CDS with lumen-apposing metal stent (LAMS) and ERCP-BD with self-expandable metal stent (SEMS) for primary MDBO palliation. A state-transition Markov model compared EUS-CDS and ERCP-BD over a 1-year time horizon from a US healthcare perspective. The base case was a 70-year-old with locally advanced, unresectable pancreatic cancer, common bile duct dilation >15 mm, and MDBO. Probabilities were derived from meta-analyses of randomized trials. Outcomes were incremental cost-effectiveness ratios (ICERs), with a willingness-to-pay (WTP) threshold of $100 000/quality-adjusted life year (QALY). Extensive sensitivity analyses were performed. EUS-CDS with LAMS was cost effective versus ERCP-BD with SEMS for primary treatment of MDBO at an ICER of $47 711/QALY. In one-way sensitivity analyses, EUS-CDS remained cost effective if it cost <$15 502 or if ERCP-BD cost >$11 174. ERCP-BD would become cost effective if technical success was >91%, reintervention <11%, or postprocedural pancreatitis <4%. Probabilistic sensitivity analysis showed EUS-CDS remained cost effective in 74.1% of iterations at a $100 000/QALY WTP threshold. In patients with MDBO and biliary dilation >15 mm, EUS-CDS with LAMS may be not only a clinically preferred option but also an economically viable primary approach. Continued efforts to minimize LAMS costs, decrease stent dysfunction, and identify optimal anatomic indications are warranted to facilitate wider adoption.

Post-endoscopy upper gastrointestinal cancer (PEUGIC) is an emerging quality metric, but varying definitions complicate benchmarking. We characterized PEUGIC rates and outcomes by anatomical site and coded high-risk condition (HRC) status in a large US cohort. Retrospective cancer-first cohort study of 14,814 patients with upper gastrointestinal cancer (UGIC) from a multicenter database (2016-2024). Detected UGIC was diagnosed within 6 months of EGD. PEUGIC was defined as UGIC diagnosed 6-36 months after a non-diagnostic EGD. The primary outcome was the PEUGIC rate by cancer type (esophageal [EC], gastric [GC], duodenal [DC]) and HRC status. Secondary outcomes included 3-year overall survival. The overall PEUGIC rate was 9.4% (EC 9.6%, GC 8.6%, DC 10.7%). Rates were higher with HRC (18.2%) versus without HRC (5.0%, P < .001). PEUGIC rate without HRC plateaued (4.1-5.1% in 2022-2024), while PEUGIC rate with HRC for EC (16.8%) and GC (14.4%) decreased over time. In exploratory analysis, PEUGIC was associated with better survival versus Detected UGIC for EC (adjusted hazard ratio [aHR] 0.82, 95% CI 0.67-0.99) and GC (aHR 0.77, 0.62-0.96), and PEUGIC with HRC was associated with lower mortality versus PEUGIC without HRC for EC (aHR 0.51, 0.36-0.74) and GC (aHR 0.50, 0.34-0.73). One in 10 upper GI cancer patients had a prior non-diagnostic endoscopy within 3 years. PEUGIC rates varied by anatomical site and HRC status, suggesting site- and HRC-stratified benchmarks may better capture endoscopic performance than aggregate metrics.

Fecal microbiota transplantation (FMT) holds therapeutic promise beyond recurrent Clostridioides difficile infection, but clinical outcomes remain unpredictable and donor-selection strategies remain limited, in part because the role of donor‒recipient metabolic interactions in shaping the post-FMT community remains poorly understood. Here, we leverage metagenome-scale metabolic modeling to quantify metabolic niche complementarity between donor and recipient microbiomes and predict post-FMT community composition. Using MICOM-derived metabolic models, we show that donor genomes whose metabolic flux profiles are more dissimilar from the recipient community colonize at significantly higher rates in a murine FMT model. In a human IBS trial, the same metric predicted post-FMT community composition via leave-one-out cross-validation and captured known disease-associated alterations in short-chain fatty acid, sulfur, and gas metabolism. We then performed 2,548 in silico FMT simulations between IBS-D/M patients and donors from the OpenBiome biobank to evaluate personalized donor screening, identifying super-donors characterized by high taxonomic diversity, broad metabolic niche coverage, and community interaction networks dominated by cross-feeding rather than competition. Together, these results support metabolic niche complementarity as a potential determinant of post-FMT community composition and provide a mechanistic basis for evaluating donor-recipient metabolic compatibility. This framework offers a scalable approach for generating testable hypotheses for personalized donor selection.

Alterations in the gut microbiota accompanied by intestinal inflammation are early features of Parkinson's disease (PD). Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene represent a common genetic risk factor for PD and inflammatory bowel disease. Parabacteroides goldsteinii has been reported to alleviate intestinal and systemic inflammation. However, whether modulation of the gut microenvironment at early disease stage can attenuate PD progression remains unclear. To investigate the impact of P. goldsteinii colonization prior to the onset of motor dysfunction on PD progression. We established a germ-free PD mouse model carrying the LRRK2 G2019S mutation and administered P. goldsteinii orally at the pre-symptomatic stage to evaluate its effects on motor performance and PD-related neuropathology. Spatial and bulk RNA transcriptomic analyses of brain tissue, together with cytokine profiling, were conducted to assess central changes. To investigate gut immunomodulatory mechanisms, we performed intestinal bulk and single-cell RNA sequencing, spectral flow cytometry as well as cellular bioenergetic analyses. Germ-free conditions partially alleviated PD-like phenotypes in LRRK2 G2019S mice. Colonization with P. goldsteinii at 5-months of age, prior to motor symptom onset, further improved locomotor performance, reduced neuronal α-synuclein aggregations, and mitigated microglial activation and dopaminergic neurodegeneration. Neuroprotection was mediated through enhanced noncanonical neuronal IL-12 receptor-dependent neurotrophic support without activating the canonical STAT4 phosphorylation pathway, along with suppression of microglial activation and downregulation of LRRK2 kinase activity. At the intestinal level, P. goldsteinii suppressed TLR4-driven inflammation, expanded anti-inflammatory intraepithelial CD4+CD8αα+ T cells, promoted dendritic cell and macrophage differentiation, upregulated epithelial tight-junction genes, and improved mitochondrial bioenergetics in intestinal cells. P. goldsteinii colonization attenuates the progression of LRRK2-associated parkinsonism by restoring intestinal homeostasis and reducing neuroinflammation. These findings underscore the therapeutic potential of modulating the gut-immune-brain axis during the prodromal stage of PD.

Alcohol-related hepatitis (AH) is a complex disease associated with numerous unmet needs, particularly in diagnosis and treatment. The epidemiology of AH has evolved in recent years, reflecting changes in alcohol consumption during the COVID-19 pandemic and the increasing incidence of AH following bariatric surgery. Advances have also been made in the non-invasive diagnosis of AH, helping to reduce the need for liver biopsy, as well as in the management of infection. Several novel therapeutic strategies have been evaluated, including faecal microbiota transplantation, IL-1 receptor antagonists, oxysterols, and reduced exposure to corticosteroids or antibiotics. Although the results of these trials have been relatively disappointing, they have helped identify promising directions for future research. In patients with the most severe form of AH, particularly those who do not respond to corticosteroids, several studies have suggested that the indications for early liver transplantation could be expanded. Overall, developments over recent years have generated increased optimism regarding the management of patients with severe AH.

Severe alcohol-related hepatitis (sAH) is associated with high short-term mortality. However, 30-40% of patients fail to respond to corticosteroids, the only proven pharmacological treatment. The pathophysiological mechanisms underlying sAH and the marked heterogeneity in treatment response remain incompletely understood. We aimed to define cellular changes associated with corticosteroid response in sAH and to identify baseline markers predictive of treatment outcome. Single-nucleus RNA sequencing was performed on liver biopsies from patients with biopsy-proven sAH (n = 17), including paired baseline and day 8 biopsies in a subset of patients (n = 8). Patients were classified as corticosteroid responders (sAH-R; Lille score <0.45) or non-responders (sAH-NR; Lille score ≥0.45). Liver biopsies from patients with acute decompensation of alcohol-related cirrhosis (AD; n = 5) and healthy controls (n = 4) were included for comparison. Findings were validated using immunohistochemistry and spatial proteomics in a large multicenter validation cohort (n = 172). At baseline, sAH-R exhibited a significantly higher proportion of liver-infiltrating S100A8+ monocytes compared to sAH-NR, a difference that persisted at day 8. sAH livers showed a marked reduction in mature hepatocytes and an expansion of stressed and intermediate hepatocyte populations, indicating progressive loss of mature hepatocyte identity. This loss was more pronounced in sAH-NR, who also exhibited fewer cycling hepatocytes at day 8, consistent with impaired regenerative capacity. Expression of SULT2A1, a marker of mature hepatocyte identity, was significantly reduced in sAH-NR at baseline. Finally, liver biopsies with ≥50% SULT2A1-positive hepatocytes at baseline were strongly predictive of corticosteroid response. This study delineates distinct immune and hepatocyte changes associated with corticosteroid response in sAH. Baseline SULT2A1 expression may facilitate stratified treatment approaches in sAH. Severe alcohol-related hepatitis (sAH) represents one of the most devastating manifestations of alcohol-related disorders. sAH is characterized by acute hepatic inflammation and high short-term mortality. Clinical management remains challenging, as corticosteroids - the only pharmacological therapy currently applied - are ineffective in a substantial proportion of patients and are associated with significant adverse effects. These limitations highlight the need for a more detailed understanding of sAH pathophysiology and for approaches that enable improved patient stratification and therapeutic decision-making. In this study, we identify distinct pre-treatment differences between corticosteroid responders and non-responders at the level of both hepatocytes and myeloid cells, providing new insight into the cellular mechanisms underlying treatment heterogeneity in sAH. Furthermore, leveraging these findings, we developed a histology-based SULT2A1 scoring system using a commercially available antibody, which predicts corticosteroid response at baseline and may support stratified treatment approaches in clinical practice.

Computer-aided diagnosis (CADx) may support optical diagnosis of diminutive (≤5 mm) colorectal polyps, replacing histopathology. We described CADx usage and diagnostic performance under real-life, discretionary adoption. This prospective implementation study enrolled patients undergoing colonoscopy at our tertiary center (2021-2025). Patients aged <40 years, with inflammatory bowel disease, polyposis syndromes, or no diminutive polyp were excluded. CADx use was at the endoscopists' discretion. Endoscopists' optical diagnoses with and without CADx assistance, and the CADx output were documented. Main outcomes were CADx usage and sensitivity for adenomas of CADx-assisted and CADx-unassisted optical diagnosis, using histopathology as reference. 868 patients (age 66.1 years, 47.0% female) with 1660 diminutive polyps (62.0% adenomas) were included. Endoscopists recorded CADx-assisted and CADx-unassisted optical diagnoses for 657 and 864 polyps, respectively, showing sensitivity of 85.1% (95%CI 78.3-90.1) and 85.1% (81.2-88.4), specificity 55.2% (45.8-64.3) and 61.1% (54.5-67.3), positive predictive value 76.2% (69.1-82.1) and 79.3% (75.5-82.9), negative predictive value (NPV) 60.8% (52.6-68.5) and 67.1% (60.7-73.0), and accuracy 62.7% (56.5-68.4) and 66.4% (63.0-69.7), respectively. In a simulated resect-and-discard strategy, CADx assistance marginally increased the proportion of polyps that could avoid histopathology (84.9% vs. 81.1%). For high-confidence rectosigmoid diagnoses, NPV for adenomas was 95.5% (86.9-98.5) for CADx-assisted and 93.8% (85.9-97.4) for CADx-unassisted predictions. Both CADx-assisted and CADx-unassisted optical diagnoses showed good diagnostic performance in real-world practice. Assessment of the relationship between CADx use and optical diagnosis was limited by unavoidable selection bias relating to discretionary use and variable endoscopist behavior.

Colonoscopy quality and diagnostic outcomes in Lynch syndrome have historically been heterogeneous. Following the introduction of the Lynch syndrome Bowel Cancer Screening Programme (BCSP), we assessed colonoscopy quality and diagnostic yield in Lynch syndrome using the National Endoscopy Database (NED), comparing outcomes between BCSP and non-BCSP providers. We performed a cross-sectional analysis of UK Lynch syndrome colonoscopies from NED between June 2024 and June 2025. Colonoscopy quality indicators and endoscopic detection rates for polyp (PDR) and advanced neoplasia (eANDR; polyps ≥10 mm or cancer; histopathology linkage was unavailable) were calculated and compared between BCSP and non-BCSP providers. Factors associated with neoplasia detection were identified using logistic regression. Among 3196 colonoscopies, most were performed in England (90.2%), with 34.1% undertaken within BCSP. Overall colonoscopy quality was high: 97.6% were complete, 89.0% had adequate bowel preparation, and 96.3% had adequate tolerance. BCSP procedures demonstrated higher rates of completion (98.5% vs. 97.1%; P = 0.02) and adequate bowel preparation (92.7% vs. 86.7%; P < 0.001) than non-BCSP colonoscopies. PDR and eANDR were 46.8% and 4.7%, respectively, and were higher in BCSP than non-BCSP procedures (PDR 53.7% vs. 43.3%; eANDR 5.7% vs. 3.9%). Higher polyp detection was associated with increasing age (odds ratio [OR] 1.03, 95%CI 1.03-1.04), male gender (OR 1.48, 95%CI 1.26-1.73), BCSP status (OR 1.50, 95%CI 1.28-1.77), adequate bowel preparation (OR 1.81, 95%CI 1.40-2.34), and England nation (OR 1.99, 95%CI 1.22-3.36). Our NED evaluation demonstrated high colonoscopy performance in Lynch syndrome, with superior quality and endoscopic yield in BCSP procedures, supporting national integration of surveillance to establish Lynch syndrome-specific quality benchmarks.

In the duodenum of patients with irritable bowel syndrome (IBS), acute food-induced mucosal reactions have been visualized using confocal laser endomicroscopy (CLE). In uncontrolled studies, a diet excluding trigger foods led to symptomatic improvement. Here, we aim to assess the association of food-induced alterations identified using CLE and the efficacy of a targeted diet in a controlled study. Double-blind, controlled, crossover study in patients with IBS. Following CLE, patients excluded up to two foods that did (= real diet) and up to two foods that did not (= sham diet) cause alterations in a randomized order. Diets were followed for four weeks each. Clinical response was defined as improving ≥50 points on the IBS symptom severity score (IBS-SSS). Healthy controls underwent the same CLE protocol as patients with IBS. Altered mucosa was visualized in all patients either before (11%) or after food administration. Clinical response rates were not different between real and sham diet (42% vs 36%, OR = 1.33, p = 0.6). Symptoms improved both on the real and the sham diet (-30 vs -20; p = 0.7). Also, in 15 healthy volunteers, altered mucosa was visualized in 100% of the exams either at baseline (13%) or after food administration. Acute alterations visualized using CLE have poor sensitivity and specificity for dietary responses in IBS patients. Moreover, altered mucosa was present in 100% of healthy volunteers and likely represents a non-specific mucosal reaction to food exposure rather than pathology, challenging the use of CLE in clinical practice to identify culprit foods. (clinicaltrials.gov: NCT05097872).

1: Before commencing hands-on upper gastrointestinal bleeding (UGIB) training, trainees should have thorough knowledge of: pathology, vascular anatomy, technical use of devices, clinical care pathways, and all relevant components of pre-, intra-, and postprocedural patient care. 2: Preadoption technical skills required for training in the management of UGIB include adequate scope handling, intubation technique, washing and suctioning of residue, mucosal examination, and handling of accessories. 3: Preadoption technical skills required for training in the management of UGIB should be assessed individually based on a competency framework and not solely on numerical thresholds. 4: The integrative skills required for the management of UGIB do not essentially differ from those needed in other endoscopic procedures and should include adequate situational awareness, collaboration, team leadership, and patient communication in order to ensure short- and long-term goals are achieved. 5: The trainee should reach minimum recommended standards for key performance indicators in upper GI endoscopy before starting training in the management of UGIB. 6: Attendance of at least one half-day training session on a dedicated simulator that provides GI bleeding training at the beginning of training for management of UGIB is advised. 7: Trainees should be supervised directly and carefully during UGIB training for a minimum of 20 procedures with endoscopic stigmata of recent hemorrhage in order to prevent failure of hemostasis and ensure adequate trainee skill acquisition. 8: Trainers should take into account pre-endoscopic and intraprocedural factors predicting outcome, technical complexity, and risk of hemostatic failure when deciding appropriateness and degree of trainee involvement in case management. 9: During their training, trainees should be exposed to all hemostatic modalities, as per ESGE guideline recommendations, and the opportunities for teaching arising from the specifics of each UGIB case. 10: Trainers should use "successful hemostasis," defined as the absence of any further bleeding (persistent or recurrent bleeding), as the ultimate goal of training in the endoscopic management of UGIB. 11: Patients with recurrent bleeding should be treated by experienced endoscopists or by a trainee under their direct supervision owing to the higher risk of failure of conventional endoscopic treatment. 12: Trainers who are teaching management of UGIB should fulfil the same standards as any trainer of basic endoscopy procedures. 13: Trainees should be exposed to multidisciplinary team discussions and care pathways for failed endoscopic treatment of UGIB. 14: Training centers with limited UGIB case volumes are encouraged to offer short-term immersive training in centers with high volume caseloads of UGIB in order to ensure sufficient exposure for trainees. 15: Competency in managing UGIB is defined as the ability to assess the need for endoscopy, and plan and carry out successful hemostasis. 16: Trainees should manage an indicative number of 30 cases in which successful hemostasis is achieved before evaluation of competence in management of UGIB. 17: UGIB-CAT is advised as a formative assessment tool during training to track acquisition of competence and provide trainee feedback. 18: Trainees should undergo a formal summative assessment of competence in managing UGIB during their training. 19: Endoscopists should continue a period of tracking results and mentored practice with an experienced colleague for at least 6 months after achieving competence in managing UGIB. 20: As trainees move to independent practice, they should have established access to or referral pathways for key supporting specialties involved in the nonendoscopic management of acute UGIB (including emergency medicine, surgery, and interventional radiology).

The incidence of esophageal adenocarcinoma (EAC) is rapidly increasing in Western countries. Gastroesophageal reflux, containing acidic bile salts (ABS), is the main risk factor for EAC. Cancer cells develop adaptive abilities to recalibrate redox balance via hijacking the antioxidant systems to maintain reactive oxygen species (ROS) below lethal levels. We investigated the role of PRDX2 and its regulation in EAC chemoresistance under reflux conditions. We analyzed public databases to identify PRDX2's aberrant overexpression and potential role in chemoresistance in EAC. To model acute and chronic GERD in vitro, we applied transient and repeated ABS exposures. Using 2D and 3D organotypic culture models of EAC cells, we investigated PRDX2's anti-ferroptosis, pro-chemoresistance functions, and regulatory mechanisms. In addition, we also utilized patient derived organoids and xenografts, cell line-derived xenografts, and human EAC tissue microarrays. Aberrant expression of PRDX2 was detected in both human EAC tissues and cell lines under ABS exposure. PRDX2 was regulated via an APE1-redox-dependent transcription activation of NF-kB. The knockdown of PRDX2 impaired the recovery of EAC cells from ABS-induced ROS and ROS-dependent lipid peroxidation. Silencing PRDX2 sensitized the chemo-resistant EAC cells to oxaliplatin by enhancing ferroptosis. Mechanistically, we found that PRDX2 inhibits ferroptosis by stabilizing GPX4, a crucial ferroptosis suppressor. PRDX2 enhances GPX4 stability through OTUB1-dependent deubiquitination, preventing its degradation. The APE1-redox inhibitor APX2009 significantly sensitized EAC cells to oxaliplatin by downregulating PRDX2 and inducing ferroptosis. Importantly, the combination of oxaliplatin and APX2009 showed synergistic tumor-suppressive effects in xenograft models. Our findings revealed a novel link between reflux-induced redox rebalance and ferroptosis-related chemoresistance in EAC via APE1-redox/NF-kB/PRDX2/OUTB1/GPX4 signaling cascade. Targeting redox with APE1-redox-specific inhibitors is a potential novel strategy for drug combination in refractory EAC, via inhibition of PRDX2.

Infection with hepatitis B virus (HBV) remains a severe concern to public health, with roughly 292 million people worldwide suffering from the chronic form of the disease, for which there is no cure. Chronic HBV infections frequently lead to hepatocellular carcinoma (HCC), one of the world's leading causes of cancer-related deaths. Although the process of hepatocarcinogenesis is complex and not fully understood, various studies have identified numerous long non-coding RNAs (lncRNAs) as contributing to the formation of HCC. These host-derived lncRNAs are frequently dysregulated as a result of viral infection. Numerous lncRNAs have been linked to HBV carcinogenesis and replication, particularly those that are dysregulated in HBV-associated HCC. HBV X protein regulates the majority of these dysregulated lncRNAs. Certain lncRNAs have been found to exert regulatory functions in HBV replication and carcinogenesis. The prognosis for HCC remains poor, and early detection of novel tumor markers is critical for effective HCC therapy. Understanding the biological activities and regulatory mechanisms of HCC-associated lncRNAs will aid in disease diagnosis and therapy and help elucidate the disease etiology. In HBV-related HCC, certain dysregulated lncRNAs may develop into biomarkers for early detection or potential targets for HCC treatment. This review provides a brief overview of the recent findings on lncRNAs in HBV with a focus on current developments. We also investigated the possible relevance of dysregulated lncRNAs in HCC as biomarkers for diagnosis and treatment and assessed their carcinogenic and tumor-suppressive effects.

Achalasia appears to increase the risk of esophageal squamous cell carcinoma, but existing evidence relies on small studies that did not adjust for this cancer's main risk factors (ie, smoking and alcohol overconsumption). Whether achalasia increases the risk of esophageal adenocarcinoma, whose main risk factor is gastroesophageal reflux disease (GERD), is even more uncertain. This study aimed to clarify these associations. A population-based case-control study was conducted in 5 Nordic countries from 1987 to 2023. Esophageal cancer cases (n = 35,604) were matched to 10 times as many background population controls by age, sex, calendar year, and country (n = 355,869). Conditional logistic regression provided odds ratios (ORs) with 95% confidence intervals (CIs), adjusted for smoking and alcohol, and additionally for GERD in a mechanistic model. All variables came from national health data registries. Achalasia was present in 137 cases (96 squamous cell carcinomas and 41 adenocarcinomas) and 230 controls (99 squamous cell carcinomas and 131 adenocarcinomas). Achalasia was associated with a strongly increased risk of esophageal squamous cell carcinoma (OR, 9.46; 95% CI, 7.07-12.65). A moderate association was found for esophageal adenocarcinoma (OR, 3.05; 95% CI, 2.14-4.35), which attenuated after adjustment for GERD (OR, 1.44; 95% CI, 0.99-2.09). Achalasia treated with myotomy or dilation was associated with an even stronger association with esophageal squamous cell carcinoma (OR, 16.33; 95% CI, 11.26-23.67), but not with esophageal adenocarcinoma after adjustment for GERD (OR, 1.32; 95% CI, 0.76-2.30). Achalasia appears strongly and independently associated with esophageal squamous cell carcinoma. The association with esophageal adenocarcinoma is less strong and seems to be largely explained by GERD.

Bulevirtide is approved in several countries and regions for the treatment of compensated chronic hepatitis D (CHD). However, long-term outcomes after treatment discontinuation remain unknown. Patients with CHD (n = 150) were randomized to immediate treatment with bulevirtide 2 mg/day (n = 49) or 10 mg/day (n = 50) for 144 weeks (W), or a 48W delay before starting treatment (DT; n = 51) followed by bulevirtide 10 mg/day for 96W (DT/10 mg; n = 50), and 96W of post-treatment follow-up (FU96) in the MYR301 study. Efficacy endpoints included virologic response (VR; undetectable HDV RNA or ≥2 log10 IU/ml decline from baseline), combined response (CR; VR and alanine aminotransferase [ALT] normalization), ALT normalization, and undetectable HDV RNA. At the end of treatment (EOT), response rates in the 2, 10, and DT/10 mg groups were: VR, 73%, 76%, and 92%; ALT normalization, 59%, 60%, and 58%; CR, 57%, 54%, and 56%; and HDV RNA undetectability, 29%, 50%, and 52%. At FU96, VR rates declined to 33%, 30%, and 32%, respectively; CR rates were 24% across all groups. HDV RNA undetectability rates were 20%, 22%, and 20% at FU96. Sustained undetectability through follow-up was observed in 23/64 (36%) patients with undetectable HDV RNA at EOT, with weeks continuously undetectable at EOT being the most important predictor of sustained undetectability. Post-treatment hepatic serious adverse events occurred in 20/142 (14%) patients and resolved in 17/20 (85%). Bulevirtide treatment for CHD for up to 144W was safe and effective. Response rates decreased after treatment discontinuation; however, some patients had sustained undetectable HDV RNA throughout 2 years of follow-up. Although bulevirtide is approved for treatment of chronic hepatitis D (CHD) in several countries and regions including the United States, the European Economic Area, the United Kingdom, Switzerland, the Russian Federation, Australia, and Canada, treatment outcomes beyond 2 years and after bulevirtide discontinuation remain unknown. In this analysis, we demonstrate that efficacy was maintained with bulevirtide monotherapy for up to 144 weeks compared with that at 96 weeks, while rates of HDV RNA undetectability continued to improve with extended treatment duration. Virologic and biochemical responses decreased after treatment discontinuation, but some patients with undetectable HDV at the end of treatment maintained HDV RNA undetectability post-treatment, with duration of continuous HDV RNA undetectability at the end of treatment being the strongest predictor of non-relapse. While most patients benefit from continued bulevirtide therapy, a subset of those who achieve undetectable HDV RNA may be able to discontinue treatment without loss of response even in the absence of HBsAg loss. NCT03852719.

Gastric cancer remains a major global health burden, with high mortality driven by late-stage diagnoses that limit treatment options and reduce survival. Current diagnostic methods such as endoscopy and biopsy are invasive, resource-intensive, and impractical for large-scale early detection. This study aimed to develop and validate an ensemble machine learning model integrating four cell-free DNA (cfDNA) fragmentomic feature classes derived from 5 × whole genome sequencing (WGS) data to non-invasively differentiate malignant gastric cancer from benign gastric lesions in high-risk or symptomatic patients. A total of 681 plasma samples were prospectively collected, comprising 329 from patients with gastric cancer or high-grade intraepithelial neoplasia (HGIN) and 352 from individuals with benign gastric conditions. The dataset was divided into a training cohort (n = 333) and a temporally independent validation cohort (n = 348). An external validation cohort of 305 participants was also included. The ensemble model achieved an AUROC of 0.920 in cross-validation testing on the training cohort, 0.912 in the independent validation cohort, and 0.896 (95% CI 0.860-0.932) in the external cohort. At a pre-specified prediction threshold of 0.402, the model demonstrated 93.3% sensitivity and 71.9% specificity in the validation cohort, yielding a PPV of 71.3% and an NPV of 93.5%. In the external cohort, sensitivity and specificity were 91.7% and 69.1%, respectively (PPV 75.7%, NPV 88.8%). Model scores correlated with clinical stage, tumor grade, and histopathological subtype. Approximately 71% of non-cancer patients could have been spared unnecessary endoscopy. The cfDNA fragmentomics-based ensemble model enables accurate, non-invasive differentiation between gastric cancer and benign gastric lesions in high-risk or symptomatic patients. This approach demonstrates strong potential as a pre-endoscopy triage tool, supporting earlier detection and more efficient use of diagnostic resources.

HBsAg is currently used as the key serum biomarker to define functional cure of chronic hepatitis B (CHB). Declining HBsAg is often interpreted as evidence of cccDNA elimination or durable transcriptional silencing. However, HBsAg can also be produced from transcriptionally active HBV DNA integrated into the host genome (iDNA), which can make serum HBsAg an unreliable indicator of the cccDNA activity or amount. We combined intrahepatic HBsAg immunostaining, quantitative serum HBsAg levels (qHBsAg), and digital droplet PCR-based profiling of HBV DNA and RNA in liver biopsies to identify patients in whom HBsAg expression was predominantly driven by cccDNA or by iDNA. We then compared HBsAg abundance, isoform composition, secretion, and intrahepatic staining patterns between these groups. All three envelope proteins (L, M, S) were detectable in liver tissue and serum in both groups. The relative abundance of L and S, assessed by western blot and immunofluorescence, did not differ between iDNA- and cccDNA-driven cases. Across the combined cohort, intrahepatic HBsAg burden correlated with serum qHBsAg, indicating similar intracellular retention and secretion behavior. Importantly, all established intrahepatic HBsAg staining patterns (membranous and cytoplasmic subtypes) occurred in both groups, with no pattern uniquely associated with either source. Thus, iDNA can sustain HBsAg expression profiles that are indistinguishable from those generated by active cccDNA. Serum HBsAg alone may therefore fail to identify patients who have achieved cccDNA elimination or durable silencing. These findings suggest that the use of serum qHBsAg as a component of the functional cure endpoint in clinical studies warrants re-evaluation. Serum hepatitis B surface antigen (HBsAg) is the central biomarker used to define and monitor "functional cure" in chronic hepatitis B. This study shows that in patients with substantial HBV DNA integration, HBsAg is produced predominantly from integrated sequences, and that integration-derived HBsAg is indistinguishable from cccDNA-derived HBsAg in isoform composition, secretion behaviour, and histological distribution. Long-read sequencing confirmed that the integrated sequences generate full-length envelope transcripts. These findings are most relevant to virologists, hepatologists, clinical trialists, and regulators developing and evaluating HBV cure strategies. They indicate that serum HBsAg measurement cannot report cccDNA burden or transcriptional activity, and that focusing on HBsAg loss could miss "partial cure" states with favourable clinical outcomes. This warrants re-evaluation of HBsAg loss as a main functional cure endpoint, and supports the use of complementary or source-specific markers to assess treatment response.

Deep learning models often struggle with class imbalance and low-resolution medical images, where critical spatial details and minority-class features are underrepresented. We introduce the Adaptive Distribution-aware Vision Transformer (AdaptiveViT), a novel hybrid CNN-Transformer architecture that unifies fine-grained local feature extraction with global contextual modelling. AdaptiveViT incorporates a distribution-aware modulation mechanism that adaptively adjusts feature emphasis according to the severity of class imbalance. In addition, a Distribution-aware Adaptive (DA) Loss incorporates the dataset imbalance ratio into an adaptive focusing scheme, enhancing minority-class sensitivity. Experiments on five skin lesion datasets with varying image resolutions and imbalance ratios (as high as 1:10 for melanoma versus non-melanoma) demonstrate that AdaptiveViT consistently outperforms state-of-the-art CNN, Transformer, and hybrid baselines in F1 and AUC, while maintaining stable convergence across imbalance levels. Validation on gastrointestinal endoscopy datasets further demonstrates AdaptiveViT's domain-agnostic generalisation beyond skin lesion data, which share similar imbalance characteristics. All experiments are conducted using patient-disjoint splits, with a threshold-free evaluation protocol to ensure fair, unbiased, and clinically reliable comparisons. Overall, AdaptiveViT establishes a hybrid framework for medical image classification under class imbalance and image-resolution variability. The code is available at https://github.com/mmu-dermatology-research/AdaptiveViT.

The human HepaRG™ cell line is the closest surrogate to primary culture of hepatocytes (PHH) for toxicology studies. However, differentiated HepaRG™ cells express low levels of the cytochrome P450 2D6 (CYP2D6) involved in the biotransformation of many drugs. Herein, progenitor HepaRG™ cells were transduced using lentiviral particles encoding both human CYP2D6 and GFP proteins. The resulting transgenic HepaRG™ cells stably expressed catalytically active CYP2D6 at levels close to those observed in PHH from rapid metabolizers and HepaSH™ hepatocytes. In CYP2D6 transgenic HepaRG™ cells, tramadol was metabolized into both N- and O-desmethyl tramadol as seen in humans while parental HepaRG™ cells produced only N-desmethyl tramadol. Following treatment with perhexiline, the CYP2D6 expressing HepaRG™ cells exhibited higher IC50 values and reduced mitochondrial damages compared to those found in parental cells. Transcriptomic analysis revealed that the expression of CYP2D6 did not significantly affect the cells' ability to proliferate and differentiate or compromise key hepatocyte-specific functions. However, we identified a small number of genes, including NXF3 and TRIM63, which were up-regulated in transgenic cells. Using CRISPR/Cas9-mediated knockdown of GFP and/or CYP2D6 sequences, we demonstrated that NXF3 mRNA and protein inductions were triggered by the lentiviral mRNA encoding GFP and CYP2D6 rather than by genomic transgene integration. Together, these findings establish CYP2D6-transgenic HepaRG™ cells as an optimized and reliable hepatocyte-like model for studying the metabolism and toxicity of CYP2D6 substrates. Our results also support the hypothesis that the NXF3 gene may be a marker of cellular response to the expression of a lentiviral chimeric mRNA.