Hepatology

Reliance on invasive liver biopsy endpoints is a major limiting factor in new therapeutic development for metabolic dysfunction-associated steatohepatitis (MASH). While liver fat accumulation is considered a driver of early disease, liver fat reduction is not recognized as a surrogate endpoint by health authorities. Here, we conducted the largest-to-date genome-wide association study (GWAS) of liver fat in 79,642 people with magnetic resonance imaging and developed a polygenic score with liver fat-lowering alleles across 55 genomic loci. In a separate set of 714,886 individuals, a 30% relative liver fat reduction due to the polygenic score was associated with 39% lower odds of liver cirrhosis and 41% lower odds of hepatocellular carcinoma, consistent with a strong causal relationship between lifelong liver fat differences and advanced liver clinical outcomes. A systematic review and meta-analysis of 20 randomized controlled trials including a total of 3,502 MASH patients showed that treatment-induced liver fat reductions were associated with MASH resolution (meta-regression slope in % units of the outcome for each 1% relative reduction in liver fat from baseline, {beta} = -0.70%, p<0.001) and fibrosis improvement ({beta} = -0.24%, p<0.001). These findings demonstrate the profound etiologic impact of liver fat in MASH and provide compelling evidence for liver fat reductions as a reasonably likely surrogate endpoint for future clinical development.

Background: Focal liver lesions (FLLs) encompass a wide spectrum of benign and malignant pathologies, and accurate diagnosis is essential for appropriate management. Although advances in imaging have improved lesion characterization, histopathologic assessment remains the diagnostic gold standard for indeterminate lesions. Data on the histopathologic spectrum and diagnostic utility of ultrasound-guided percutaneous liver biopsy (US-PLB) in sub-Saharan Africa (SSA) are limited. This study aimed to characterize the histopathologic findings of US-PLB performed for FLLs at a tertiary referral center in SSA and to identify factors associated with hepatocellular carcinoma (HCC). Methods: We conducted a retrospective observational study of adult patients ([&ge;]18 years) who underwent US-PLB for FLL between January 2021 and December 2024 at Adera Medical and Surgical Center. Patients with indeterminate pathology results, incomplete records, biopsies performed for diffuse liver disease, or lesions classified as LI-RADS 1, 2, or 5 were excluded. Demographic, clinical, laboratory, imaging, histopathologic, and outcome data were extracted from medical records. Descriptive statistics were used to summarize patient characteristics and histopathologic diagnoses. Logistic regression analysis was performed to identify factors associated with HCC. Results: A total of 119 were included in the final analysis. The median age was 56 years (IQR 45-65), and 59.7% were male. No major biopsy-related complications were reported. HCC was the most common histopathologic diagnosis, accounting for 42.9% of cases, followed by secondary metastatic tumors (15.9%) and regenerative nodules (15.9%). Other diagnoses included chronic hepatitis (8.4%), cholangiocarcinoma (5.9%), and hepatic abscess (3.4%). Hepatitis B virus (HBV) and hepatitis C virus (HCV) infections were present in 14.3% and 12.4% of patients, respectively. On multivariate analysis, HBV infection (AOR 7.85, 95% CI 1.45-42.60; p=0.017), HCV infection (AOR 9.03, 95% CI 1.41-57.76; p=0.020), and larger tumor size (AOR 1.27, 95% CI 1.11-1.46; p<0.01) were significantly associated with HCC. Conclusion: Ultrasound-guided percutaneous liver biopsy demonstrated a favorable safety profile for the evaluation of FLL. HCC was the predominant histopathologic diagnosis, reflecting the substantial burden of primary liver cancer in this setting. Chronic viral hepatitis and larger tumor size were significantly associated with HCC. These findings support the continued role of US-PLB in the diagnostic evaluation of indeterminate focal liver lesions and underscores the importance of viral hepatitis prevention, surveillance, and early detection strategies in sub-Saharan Africa.

Background Gallstones are one of the most common gastrointestinal conditions closely associated with metabolic dysfunction. The metabolic dysfunction - associated fibrosis - 5 (MAF -5) score has been established as a non - invasive indicator for assessing liver fibrosis in individuals with metabolic abnormalities. However, comprehensive large - scale research examining the correlation between the MAF-5 score and gallstone occurrence remains limited. This study seeks to clarify the link between the MAF-5 score and gallstone prevalence using nationally representative data from the National Health and Nutrition Examination Survey (NHANES). Methods This study examined data from 15,560 NHANES 2017-2020 participants aged 20 years or older, ensuring complete records for MAF-5 scores and gallstone status. Gallstone presence was identified through self-reported physician diagnoses. To assess the relationship between MAF-5 scores and gallstone prevalence, weighted logistic regression models were applied, adjusting for demographic characteristics, lifestyle factors, and health conditions. Subgroup analyses were conducted to evaluate the stability of this association and detect possible interactions. Sensitivity analyses were performed by excluding extreme values ({+/-}3SD) to assess result robustness. Furthermore, MAF-5 scores were divided into quartiles to investigate gallstone prevalence trends, and a restricted cubic spline (RCS) model was employed to visualize response patterns. Results A total of 15,560 participants met the inclusion criteria, with 747 in the gallstone group and 6,367 in the non-gallstone group. MAF-5 scores were significantly higher in the gallstone group (P < 0.001). After adjusting for multiple covariates, each unit increase in MAF-5 score correlated with a 14% higher gallstone prevalence (OR = 1.14, 95% CI: 1.06-1.23). Quartile-based analysis indicated that individuals in the highest MAF-5 quartile had a 2.12-fold higher prevalence of gallstones than those in the lowest quartile (OR = 2.12, 95% CI: 1.13-3.98). RCS analysis confirmed a linear association between MAF-5 scores and gallstone prevalence. Subgroup analyses showed this association remained stable across age, sex, and racial/ethnic groups, with no significant interactions. Sensitivity analyses, excluding extreme values ({+/-}3SD), reinforced the reliability of these findings (OR = 1.15, 95% CI: 1.04-1.28). Conclusion The MAF-5 score is significantly and positively associated with gallstone prevalence, independent of demographic and lifestyle confounders. These findings indicate that the MAF-5 score may be a useful tool for assessing gallstone prevalence in individuals with metabolic dysfunction, offering valuable insights for early screening and targeted health management strategies. Keywords: Gallstones, MAF-5 score, Metabolic dysfunction, NHANES, Liver fibrosis.

Background and Aims: Vibration-controlled transient elastography (VCTE) and two-dimensional shear-wave elastography (2D-SWE) are used to assess liver stiffness in metabolic dysfunction-associated steatotic liver disease (MASLD); patients may be assessed by different methods over time. Because 2D-SWE fibrosis cut-offs are not uniformly validated and no biopsy or magnetic resonance elastography reference is available, we asked whether switching methods moves patients across decision thresholds. We evaluated agreement and decision-threshold interchangeability between VCTE and 2D-SWE in MASLD with obesity. Methods: In a retrospective cross-sectional agreement study at a tertiary-care center, 317 consecutive adults with MASLD underwent same-day VCTE (FibroScan) and GE LOGIQ E9/E10 2D-SWE. Continuous agreement was assessed by Bland-Altman analysis; the categorical analysis compared binary clinical decision thresholds (VCTE [&ge;]8.0/[&ge;]10.0 kPa; 2D-SWE [&ge;]7.204/[&ge;]8.060 kPa) using Cohen {kappa} and McNemar tests. Prespecified sensitivity analyses and a post-hoc recalibration were performed; VCTE served as operational reference. Results: VCTE yielded higher values (geometric mean VCTE/2D-SWE ratio, 1.23; ratio limits of agreement, 0.64 2.40; intraclass correlation coefficient, 0.41). At the lower threshold, 71 of 117 VCTE-positive patients (61%) were below the 2D-SWE threshold versus 5 of 200 (2.5%) reclassified upward (agreement 76.0%; {kappa} 0.417; P<.001); the upper threshold was similar (38 of 63, 60%, vs 8 of 254, 3.1%). Discordance persisted across cut-offs; agreement worsened descriptively across BMI strata. Recalibration removed the directional asymmetry but not the discordance (agreement unchanged, 76.0%). Conclusions: In MASLD patients, switching from VCTE to 2D-SWE reclassified decision-threshold status in ~60% of VCTE-positive patients; recalibration removed the directional bias but not the disagreement. Follow-up should use the same elastography modality and, when possible, the same platform.

The liver is organized into spatial zones with distinct metabolic roles, but how this architecture contributes to disease remains unclear. Glucose production is concentrated in periportal hepatocytes and depends on G6PC1; loss of this enzyme causes glycogen storage disease type Ia (GSD1a). We tested whether G6PC1 loss in specific zones, including its primary periportal location, is sufficient to cause disease. Unexpectedly, loss of G6pc1 in any single zone caused local glycogen accumulation but did not produce the systemic metabolic abnormalities or liver tumors seen after whole-liver deletion. Instead, disease developed only after near-complete loss of G6pc1 across the entire liver. These findings show that organ-wide compensation preserves metabolic homeostasis and suppresses tumorigenesis despite localized disruption. TeaserSpatial G6pc1 loss causes local glycogen storage without systemic metabolic disease.

Background & Aims: Immune-mediated liver injury from immune checkpoint inhibitors (ILICI) is a major immune-related adverse event that limits cancer immunotherapy, yet its tissue-level immunobiology is poorly defined and its management is largely extrapolated from autoimmune hepatitis (AIH). We previously identified a tri-cellular CD8+ T cell-macrophage-hepatocyte injury niche in a murine model of ILICI; here, we tested whether this niche is recapitulated in human disease. Methods: We applied imaging mass cytometry with a 32-marker panel to liver biopsies from patients with ILICI (n = 12), AIH as a disease comparator (n = 14), and healthy controls (n = 2), profiling approximately 297,000 single cells across 144 regions of interest with spatially resolved detection of apoptosis (cleaved caspase-3, cC3) and pyroptosis (cleaved gasdermin D, cGSDMD). Results: We detected histiocyte-rich granulomas in ILICI consisting of macrophages and CD8+ T cells, including activated memory-effector subsets. Permutation-based spatial analysis identified CD8+ T cell-macrophage co-localization as the most frequent significant interaction in ILICI, organizing into integrated innate-adaptive cellular neighborhoods that concentrated cC3- and cGSDMD-positive cells. Descriptively, this contrasted with AIH, in which immune cells and stroma were more spatially compartmentalized. CD8+ T-cell and macrophage densities correlated with Ishak necroinflammation scores, jaundice, and granuloma formation. Conclusions: These findings provide the first single-cell spatial proteomic characterization of human ILICI in situ; they recapitulate the tri-cellular CD8-macrophage-hepatocyte niche we previously defined in a murine model and characterize ILICI as a spatially organized innate-adaptive inflammatory process, nominating myeloid signaling and CD8-macrophage interactions as candidate liver-directed targets to uncouple hepatotoxicity from anti-tumor immunity.

Background Small common bile duct stones may pass after endoscopic sphincterotomy, but leaving the duct without attempting extraction also leaves clearance uncertain. We compared same-session stone clearance after sphincterotomy alone with clearance after sphincterotomy followed by balloon-based and/or basket extraction. Methods We conducted a single-center observational study of 203 patients with common bile duct stones 5 mm or smaller who underwent therapeutic endoscopic retrograde cholangiopancreatography. The endoscopist chose the treatment during the procedure. Forty-seven patients underwent sphincterotomy alone, and 156 underwent sphincterotomy followed by balloon-based and/or basket extraction. The primary outcome was fluoroscopically confirmed clearance during the same procedure. Intraprocedural bleeding was a secondary outcome. We calculated unadjusted risk ratios and used Fisher's exact test for sparse events. Results The mean age was 58.62 years (standard deviation 20.10), and 90 patients (44.3%) were men. Same-session clearance was documented in 21 of 47 patients (44.7%) after sphincterotomy alone and 130 of 156 (83.3%) after adjunctive extraction (risk ratio 1.87, 95% confidence interval 1.35 to 2.58; P < 0.001). A prominent ampulla was more common in the sphincterotomy-alone group (25.5% vs 10.9%; P = 0.012). Intraprocedural bleeding occurred in 2 patients after sphincterotomy alone and none after adjunctive extraction (4.3% vs 0%; Fisher's exact P = 0.053). Conclusions Adjunctive extraction was associated with a higher rate of documented same-session clearance than sphincterotomy alone. Treatment was not randomized, the groups differed at baseline, and only two bleeding events occurred. These findings therefore do not prove that adjunctive extraction is causally superior or safer. Prospective studies using standardized techniques and follow-up are needed.

Liver cancer remains a major global health burden with high mortality and limited treatment response prediction tools. Patient-derived cancer organoids have emerged as promising preclinical models that recapitulate tumor heterogeneity; however, the biological significance of morphological diversity within established organoids remains poorly characterized in hepatocellular carcinoma (HCC). In this exploratory study, we investigated whether distinct organoid growth phenotypes reflect underlying tumor biology and correlate with clinical outcomes. We established liver cancer organoids from resected tumor tissues of 27 patients and analyzed their clinical, histological, and genomic characteristics. Organoids were classified morphologically into cystic and solid types. Whole exome sequencing (WES) was conducted on six matched tumor-organoid pairs to assess genomic fidelity. Associations between organoid establishment, growth characteristics, and clinical parameters were statistically evaluated. Progression-free survival (PFS) was analyzed using the Kaplan-Meier method and univariate Cox proportional hazards regression. Organoids were successfully established in 13 of 27 cases (48.1%). Solid-type organoids were significantly associated with shorter PFS compared to cystic types (HR = 13.91; p = 0.0039, log-rank test). Organoid establishment was more frequent in older patients (>70 years), those with HBV infection, and tumors with positive {beta}-catenin expression. WES analysis demonstrated high concordance in somatic mutation profiles and variant allele frequency distributions between tissues and corresponding organoids. In univariate Cox regression, organoid growth pattern (solid vs. cystic) showed a significant association with PFS within this exploratory cohort (p = 0.0207). Patient-derived liver cancer organoids preserved the genomic and histopathological features of the original tumors. Notably, solid morphology was associated with shorter PFS, suggesting that organoid growth phenotype may serve as a supplementary indicator of tumor biological behavior in HCC. Given the modest cohort size and the absence of multivariate analysis, these preliminary findings should be interpreted with caution and warrant further large-scale validation.

Hospital readmission is used in cirrhosis research as if a return to hospital meant the disease had worsened: it is a trial outcome, a quality indicator and a prediction target. We asked what those readmissions were. Sixty-one adults with alcohol-related cirrhosis, selected because they had been hospitalized more than once, contributed all 326 later admissions to one department. A clinician read each full record and classified it for acute decompensation and for admission mode, blinded to diagnostic codes and structured flags. Fewer than two in five admissions (125, 38.3%) were decompensations. One in three (114) had been booked in advance; none of these was a decompensation, yet they used 29.1% of all bed-days. Among 124 returns within 30 days, planned admissions outnumbered decompensations, 48 to 43. Counting all admissions tracked a patients decompensations only moderately (correlation 0.50) whereas counting urgent admissions tracked them closely (0.93). In crude analysis the sicker patients had fewer admissions, not more, because planned care goes to those well enough to receive it. A liver severity score, computable for 165 admissions, did not separate planned from urgent admissions or decompensations from other stays, even within one patient. Readmission was not the same thing as decompensation.

Background & aims: TP53 (p53) coordinates diverse cellular stress response programmes including pro-survival activities, senescence, and cell death. During tissue damage, p53 can shape both the local cellular response to injury, including the fibrotic response, and influence distal organ biology. Fibrosis in the liver is a major driver of hepatocellular carcinoma (HCC) risk within metabolic dysfunction-associated steatohepatitis (MASH). It is also an important determinant of dysfunction in multiple distal tissues including the kidneys, lungs, and heart. Despite significant clinical burden, our understanding of the molecular determinants of fibrotic MASH and its relationship to multiorgan fibrosis remain incomplete. Here, we investigate local and systemic effects of hepatocellular p53 activity and cholesterol during MASH development, with implications for disease prevention. Methods: This study utilised a genetic model of stabilised p53, diet-induced MASH models with varying cholesterol compositions, and an in vitro obesogenic system to investigate p53 activity during liver disease development. Non-invasive imaging and histopathological analyses were employed to monitor p53 activity, MASH, and multiorgan fibrosis in vivo. Complementary approaches, including in vitro human multicomponent liver spheroids, cytokine arrays, and analyses of human MASH transcriptomic and proteomic datasets, were used to examine molecular drivers and patient relevance. Results: Using an inducible mouse model of MDM2 E3 ubiquitin ligase deficiency to stabilise p53, we report that hepatocellular MDM2 E3 loss results in progressive fibrotic damage, robust hepatocellular expression of the p53 target gene CDKN1A/p21 (p21),and induces p21 and fibrosis in the kidneys of male mice in a sex-specific manner. In diet-induced MASH, we observe cholesterol and p53-dependent development of liver fibrosis, high expression of hepatocellular p21, and induction of p21 and fibrosis in the kidneys of male mice-reminiscent of features observed in MDM2 E3-deficient mice. We also observe fibrosis in the lungs and heart of male MASH mice. Both a cholesterol-free obesogenic diet and liver-specific loss of p53 mitigate hepatic fibrosis and systemic induction of p21 and fibrosis. Mechanistically, p53 induces hepatic expression of senescence-associated secretory phenotype (SASP) factors, including GDF15, in vivo. A human multicomponent LiverACE spheroid model showed a concordant trend towards increased GDF15 protein abundance under steatotic stress, while in humans, elevated circulating GDF15 levels in advanced MASH correlate with increased TNFRSF1A and EPHA2, circulating markers linked to kidney injury. Conclusions: Our work identifies undue p53 activity within the liver as a driver of multiorgan fibrosis in a sex-specific manner, affecting male but not female mice. We implicate cholesterol in promoting this pro-fibrotic environment in vivo and highlight circulating factors that could identify at-risk patients for multiorgan fibrosis in MASH.

Background & Aims: Fatigue is a central symptom of chronic liver disease (CLD), substantially impacting health-related quality of life (HRQoL). This study aimed to further understand CLD symptomatology, including fatigue, and its impact on HRQoL from a patient perspective. Methods: Abbott Global Assessment of Patients unmet needs (aGAP) was a multinational, cross-sectional survey in adults with compensated CLD in China, India and Mexico, conducted between July and November 2024. Adult participants who self-reported that they had physician-diagnosed CLD and were experiencing fatigue completed a quantitative survey to assess symptom burden and included three HRQoL patient-reported outcome (PRO) questionnaires (Patient-Reported Outcomes Measurement Information System [PROMIS]-29+2, Work Productivity and Activity Impairment - Specific Health Problem version 2.0 [WPAI: SHP], Multidimensional Fatigue Inventory [MFI]). Results: Overall, 505 participants (China: 200; Mexico: 105; India: 200) completed the study. Participants reported that their CLD-related fatigue sometimes, often or always affected their self-esteem/confidence (45.1%) and ability to maintain or acquire new employment (38.6%). Most participants reported moderate (51.3%) or serious (26.9%) fatigue, with 33.5% experiencing fatigue every day or almost every day. Many participants felt their social life was negatively impacted by their fatigue (47.3%) and that there were related financial difficulties (53.9%). Use of validated PRO tools demonstrated severe fatigue (MFI: overall mean [SD] 13.9 [3.4] general fatigue and 13.4 [3.6] physical fatigue) as well as substantial levels of work and activity impairment (WPAI: SHP overall mean [SD] 53.0 [26.4]) and high levels of anxiety, pain interference, depression and sleep interference (PROMIS T-scores [&ge;]54). Conclusions: Fatigue has a substantial impact on HRQoL among adults with CLD across several countries, highlighting a global unmet need for targeted interventions to effectively identify and manage the condition.

Background & Aims. Primary sclerosing cholangitis (PSC) is a cholestatic liver disease of unknown etiology whose prevalence varies >30-fold worldwide, peaking in Northern Europe and the U.S. Upper Midwest. This geographic distribution is not fully explained by recognized risk factors. We examine its correlation with Ixodes tick exposure. Approach & Results. PSC incidence across North America, Europe, and Oceania was compared with Lyme incidence, HLA-DRB1*03 frequency, latitude and other environmental factors. Autoimmune hepatitis (AIH) and primary biliary cholangitis (PBC) were included as controls. A U.S. analysis (MarketScan, 2018-2022; 110.7 million person-years) correlated age and sex-standardized rates against 24 exposures, including Ixodes density and tick-borne infections, using ancestry-adjusted partial correlations. Cross-country PSC incidence tracked Lyme incidence (Spearman rho = 0.71-0.87); HLA-DRB1*03, AIH, and PBC did not. Alaska Native and Greenlandic populations, high-latitude but without established human exposure to Ixodes-borne pathogens, report no PSC despite high autoimmune liver disease and IBD. In the U.S., PSC was clustered and tracked Ixodes-borne pathogen incidence (ancestry-adjusted partial r, log scale: anaplasmosis +0.50, babesiosis +0.56, Powassan virus disease +0.52; in the Northeast-Midwest block, ancestry- and latitude-adjusted r = +0.72, +0.84, and +0.78, respectively). Non-Ixodes infections (Ehrlichia chaffeensis, spotted fever, tularemia), AIH, and PBC were null-to-negative; rural, agricultural, pollution, and healthcare-access also did not correlate. Conclusions. These ecological analyses are consistent with the hypothesis that Ixodes-borne pathogen exposure may trigger PSC. These ecological data cannot establish causation; they are hypothesis-generating, yielding falsifiable predictions for case control, serologic, and animal-model studies.

More than two million deaths annually are attributed to liver-related conditions, making primary human hepatocytes (PHH) an invaluable in vitro model for studying liver pathophysiology and the molecular mechanisms underlying hepatic diseases. However, because PHH do not proliferate in culture, CRISPR gene editing has been highly inefficient. Here, we report lipofection- and lentivirus-mediated protocols for CRISPR-Cas9 delivery in mouse-passaged primary human hepatocytes (mpPHH), a system that enables PHH expansion in liver-humanized mice. We achieve robust gene editing efficiencies exceeding 90% in mpPHH while maintaining cell viability. We demonstrate the utility of these protocols by disrupting CYP3A4 to impair xenobiotic metabolism and by showing that edited mpPHH efficiently engraft and expand in mice, generating liver-humanized animals. We establish the feasibility of arrayed CRISPR screening in mpPHH using an 85-gene screen to identify host factors influencing hepatitis B virus (HBV) infection, and validate key findings in humanized mice by targeting the HBV entry receptor SLC10A1 (NTCP), which reduced viral infection in vivo. Our methodology enables scalable genetic manipulation of mpPHH, opening new avenues for HBV research and liver disease modeling.

Background: Metabolic adaptation is a recognized feature of therapeutic resistance in hepatocellular carcinoma (HCC), but it is unclear whether transcriptional states exposed during acquired resistance are restricted to drug adaptation or reflect broader aggressive tumor biology. We tested whether metabolic programs derived from a lenvatinib-resistance model identify a clinically adverse transcriptional state in an independent HCC patient cohort. Methods: The discovery framework was based on GSE186191, comprising parental and acquired lenvatinib-resistant Hep3B and Huh7 cells. A pre-specified 33-gene lipid-source ledger served as a biological anchor, and three discovery-derived programs, MYC Targets V2, mTORC1 Signaling, and Fatty Acid Metabolism, were frozen before patient-level evaluation. In TCGA-LIHC, single-sample enrichment scores for the three programs were population-standardized and summed to generate an integrated metabolic score. Overall survival was assessed by Kaplan-Meier and Cox analyses. Whole-transcriptome differences between high- and low-score tumors were characterized by preranked gene set enrichment analysis (GSEA). Results: The survival cohort comprised 282 patients (118 deaths), with 141 patients in each median-defined score group. High-score patients had shorter overall survival (log-rank P=0.000419). The continuous score was associated with mortality in univariable analysis (HR 1.86, 95% CI 1.33-2.61; P=0.000293) and in the frozen model adjusted for age, sex, and stage indicators (HR 1.93, 95% CI 1.35-2.76; P=0.000350; n=277). In 327 primary tumors, Fatty Acid Metabolism was strongly depleted in high-score tumors (NES -2.06; FDR<0.001). MYC Targets V2 (NES 1.18; FDR=0.232) and mTORC1 Signaling (NES 1.11; FDR=0.229) showed positive directional enrichment without FDR significance. Conclusions: A lenvatinib-resistance-derived transcriptional program is associated with an adverse-survival state in HCC. The strongest patient-level pathway feature is depletion of canonical fatty-acid metabolism, accompanied by directional MYC/mTORC1 features rather than statistically established pathway activation. These findings support a testable model of metabolic identity remodeling but do not establish causality or clinical prediction of lenvatinib response.

Background: To date, no precision oncology approach has been established for HCC. Despite the diverse underlying causes, HCC development exhibits a uniform pathophysiology characterised by chronic hyper-proliferation, resulting from hepatocyte apoptosis and compensatory liver regeneration. This chronic hyper-proliferative pressure, termed regeneration stress, drives genomic instability during HCC onset, yet its therapeutic potential remains poorly explored. This study aimed to identify targetable vulnerabilities tied to regeneration stress and establish clinically applicable markers for treatment stratification. Methods: Weighted gene co-expression network analysis (WGCNA) was applied on external bulk RNA-seq datasets to define a LIVer REgeneration Stress Signature (LIVRESS). The signature was functionally validated using HCC patient-derived organoids (HCC-Org), and vulnerabilities were mapped using mid-throughput drug screening, single-molecule and single-cell assays, and multi-omic integration. Results: High LIVRESS scores, characterised by enrichment in replication, mitotic and DNA damage repair pathways, identified a subset of HCC patients with aggressive disease and poorer survival across aetiologies. HCC-Org with high LIVRESS scores displayed exquisite sensitivity to multiple inhibitors of the checkpoint kinase ATR. Although HCC-Org models exhibited a baseline reduction in replication fork speed, sensitivity to ATR inhibitor (ATRi) was decoupled from replication fork dynamics and rather linked to intrinsic mitotic instability. ATR inhibition triggers mitotic failure and apoptosis in LIVRESSHigh HCC-Org. This killing effect was significantly potentiated by combining ATRi with PARPi or WEE1i. Multi-omic integration identified KPNA2 as a surrogate biomarker of ATRi sensitivity. Conclusion: Our findings demonstrate that a subset of HCC-Org, characterised by high liver regeneration-associated stress, is vulnerable to ATRi-based therapies. By focusing on a comprehensive regenerative stress model, we establish a framework to stratify HCC patients and implement biomarker-driven, ATR-based therapies for HCC patients with advanced disease. Impact and implications: Regeneration stress is a key factor that drives genomic instability in HCC, providing a basis for the LIVRESS to identify patients dependent on ATR-mediated checkpoints. These findings reveal a conceptual shift for researchers and trialists: ATRi efficacy is decoupled from replication fork dynamics and instead leverages mitotic fragility. Practically, the LIVRESS and its IHC surrogate marker (KPNA2) offer a scalable roadmap for physicians to improve patient stratification in ATRi-based precision oncology trials. While requiring prospective validation, these results pave the way toward biomarker-driven therapies for advanced HCC.

Hepatic fibrosis remains a major cause of morbidity and mortality in patients with schistosomiasis without effective therapy. Here we show that vitamin D receptor (VDR) signaling in macrophages is essential for limiting fibrosis, acting through a previously unrecognized metabolic epigenetic circuit. Hepatic macrophages exhibit the highest VDR expression among liver resident cells, and pharmacological VDR activation with paricalcitol selectively expands a protective SPP1+ macrophage subset derived from circulating monocytes. Myeloid specific VDR knockout exacerbates fibrosis and abrogates paricalcitol's hepatoprotective effects, whereas SPP1+ macrophage depletion worsens disease. Mechanistically, VDR activation synergizes with hypoxia and lactate to drive SPP1 expression via glycolytic reprogramming. Moreover, lactate and HIF1alpha cooperatively induce VDR transcription by binding to the Vdr promoter and enhancing histone lactylation, forming a positive feedback loop that amplifies the antifibrotic response. Our findings establish the lactate-HIF1alpha-VDR-SPP1 axis as an endogenous defense mechanism and identify macrophage VDR as a promising therapeutic target for fibrotic liver diseases.

Progressive hepatic fibrosis is the principal determinant of morbidity and mortality in metabolic dysfunction-associated steatotic liver disease and steatohepatitis (MASLD/MASH). Mitochondrial dysfunction is a hallmark of MASH, and the release of mitochondrial damage-associated molecular patterns (mito-DAMPs) from injured hepatocytes can promote fibrosis. However, how mitochondrial dynamics and quality control shape the fibrotic response in MASLD/MASH remains unclear. Here, through large-scale genomic analyses of mitochondrial genes governing mitophagy, fusion and fission in human MASLD, with a power-equivalent sample size of approximately 700,000 individuals, we identify a strong association between hepatic fibrosis and the mitochondrial fusion factor dynamin-like GTPase optic atrophy 1 (OPA1). OPA1 transcripts and protein abundance in the liver epithelium were progressively dysregulated with advancing fibrosis. In mice, hepatocyte-specific OPA1 loss alone was sufficient to induce hepatic stellate cell activation and fibrosis in zone 3, promoted the release of mito-DAMPs into the circulation and exacerbated fibrosis in experimental MASH. These findings identify OPA1 as a central regulator of the hepatic fibrotic response and connect defective mitochondrial homeostasis to mito-DAMP release, hepatic stellate cell activation and fibrosis in MASLD.

Nuclear bile acid (BA) signaling plays a central role in liver homeostasis and represents a major therapeutic axis in fibrotic liver diseases. The farnesoid X receptor (FXR), a master nuclear effector of BA signaling, is expressed in several liver-resident cell types, suggesting that it may regulate distinct biological programs beyond the hepatocyte (HC) compartment. Using complementary pharmacological, genetic, and computational approaches across in vitro, ex vivo, and in vivo models of mouse and human origin, we investigated the role of hepatic stellate cell (HSC) FXR (FXRHSC) in both unchallenged and injured livers, which has remained controversial. FXR is robustly expressed in both HCs and HSCs with distinct isoform distributions, and these isoforms exhibited differential capacities to activate gene expression in an HSC context. We found that the potent selective FXR agonist tropifexor triggers a transcriptional program reminiscent of that observed after partial hepatectomy and associated with HC proliferation. This cell cycle-related response was also observed in HSCs and did not require intestinal FXR expression. An HSC-specific response to tropifexor was observed for several genes, including members of the glutathione-S-transferase (GST) family or Scube1. FXRHSC was sufficient to observe the anti-fibrotic effects of tropifexor in precision-cut liver slices, an ex-vivo model of fibrosis. Finally, we identified the regulation of the chemerin-encoding gene Rarres2 as a relevant example of FXRHSC-dependent control of hepatic intercellular communication. Together, these findings identify FXRHSC as an important contributor to hepatic adaptation and therapeutic response to BA analogs and confirmed HSCs as a significant site of nuclear bile acid signaling in liver biology.

Neutrophils infiltrate the liver as metabolic dysfunction-associated steatotic liver disease (MASLD) progresses from simple steatosis to steatohepatitis, but whether neutrophil effectors merely mark this transition or cause it is unknown. Here we show that human neutrophil peptides (HNPs), or alpha-defensins, are drivers rather than bystanders of disease. In patients surgically treated for hepatic hemangioma who displayed overt MASLD, the abundance of HNPs in their liver biopsy samples localized to diseased tissue correlates with the histological severity of steatosis. In vitro, HNPs enter hepatocytes to induce multimerization of voltage-dependent anion channel 1 (VDAC1) in the mitochondrial outer membrane, causing mitochondrial dysfunction and lipid accumulation. Under lipid overload, but not on a standard diet, HNP1-transgenic mice develop increased adiposity and steatohepatitis, which is reversed by an orally available prodrug that inhibits HNP-induced VDAC1 multimerization. These findings link neutrophil innate immunity to hepatocyte mitochondrial dysfunction, recast host-protective HNPs as pathogenic effectors when hepatocyte mitochondria are lipid-burdened, and establish the HNP-VDAC1 axis as a therapeutic target in MASLD.

Background Alcohol-associated liver disease (ALD) has emerged as a major cause of chronic liver disease and liver-related mortality in China. This study aimed to project the future burden of ALD in Chinese adults from 2020 to 2050, including prevalence of ALD, number of alcoholic steatohepatitis (ASH) cases, incident hepatocellular carcinoma (HCC) cases, liver transplantation (LT) demand, liver-related deaths, and disability-adjusted life years (DALYs). Methods We developed an agent-based state-transition microsimulation model with yearly cycles and a lifetime horizon. The model simulated 5,678,912 representative Chinese adults (mean age 36.2 years, 51.2% male). Health states included no steatosis, alcohol-associated steatotic liver, ASH, fibrosis stages F0-F4, decompensated cirrhosis, HCC, LT, and liver-related death. Model inputs were derived from the China Kadoorie Biobank, Global Burden of Disease Study 2021, China's national surveys, published meta-analyses, and transplant registry data. Projections incorporated demographic shifts, alcohol consumption trends, and calibrated transition probabilities. Uncertainty was assessed via 1,000 Monte Carlo simulations generating 95% uncertainty intervals. Results ALD prevalence was projected to increase from 4.8% (55 million individuals) in 2020 to 8.5% (94 million individuals) by 2050. ASH cases rose from approximately 18 million to 20 million. Annual incident HCC cases nearly doubled from 20,500 in 2020-2025 to 45,200 by 2046-2050. LT demand quadrupled from 2,300 to 9,800 cases. Liver-related deaths increased from 50,000 in 2020 to 85,000 in 2050, while DALYs rose from 1.5 million to 2.6 million. Conclusions In the absence of strengthened alcohol control policies, ALD will impose a substantial and growing burden on China's health system by 2050, with marked increases in HCC incidence, LT demand, and liver-related mortality.