Extranodal natural killer/T-cell lymphoma (ENKTCL) is strongly associated with Epstein-Barr virus (EBV) infection. A 30-base-pair deletion in latent membrane protein 1 (del-LMP1) represents the most common variant in the EBV genome, but its clinicopathological significance in ENKTCL remains poorly elucidated. Some scholars suggested that the LMP1 protein product carrying the deletion gene reduced immunogenicity, allowed it to escape immune surveillance in immunocompetent hosts and confer a survival advantage. Therefore, simultaneous assessment of del-LMP1 and LMP1 protein expression may provide deeper insights into the potential role of LMP1 in ENKTCL tumorigenesis and progression. This study aimed to investigate the impact of del-LMP1 and LMP1 protein expression on the clinicopathological manifestations and prognosis of ENKTCL patients in Wenzhou. The clinical and histological characteristics of 42 ENKTCL cases were retrospectively evaluated. Del-LMP1 was detected using a nested polymerase chain reaction and Sanger sequencing, while LMP1 protein expression was assessed via immunohistochemistry. Overall survival (OS) was analyzed. The LMP1 gene was identified in 37/42 ENKTCL cases, including 2 wild-type (wt-LMP1), 35 del-LMP1 cases. LMP1 protein expression was positive in 21/42 cases. In the control group, the LMP1 gene was detected in 6/10 cases, all of which were del-LMP1, and the LMP1 protein was positive in 4/10 cases. Fisher’s exact test revealed no significant differences between the two groups in the LMP1 gene, del-LMP1, or LMP1 protein expression. Additionally, there was no significant correlation between del-LMP1 and LMP1 protein expression and clinical characteristics such as age, gender, or vascular invasion. However, LMP1 protein expression was significantly higher in necrotic tissues (p = 0.030) and younger patients with del-LMP1 (p = 0.004). Survival analysis showed no significant difference in OS between wt-LMP1 and del-LMP1 patients (p = 0.331) or between LMP1-positive and -negative cases (p = 0.592). In this retrospective cohort, we demonstrated that del-LMP1 might be the predominant variant rather than a phenotype-associated polymorphism in ENKTCL from a molecular epidemiological perspective. Moreover, LMP1 protein expression was associated with necrotic tissue and younger patients with del-LMP1, possibly due to the enhanced pathogenic effect of the mutated LMP1 isolate protein.

The complex etiological factors associated with metabolic dysfunction-associated fatty liver disease (MAFLD), including perturbed iron homeostasis, and the unclear nature by which they contribute to disease progression have resulted in a limited number of effective therapeutic interventions. Here, we report that patients with metabolic dysfunction-associated steatohepatitis (MASH), a pathological subtype of MAFLD, exhibit excess hepatic iron and that it has a strong positive correlation with disease progression. FerroTerminator1 (FOT1) effectively reverses liver injury across multiple MASH models without notable toxic side effects compared with clinically approved iron chelators. Mechanistically, our multi-omics analyses reveal that FOT1 concurrently inhibits hepatic iron accumulation and c-Myc-Acsl4-triggered ferroptosis in various MASH models. Furthermore, MAFLD cohort studies suggest that serum ferritin levels might serve as a predictive biomarker for FOT1-based therapy in MASH. These findings provide compelling evidence to support FOT1 as a promising novel therapeutic option for all stages of MAFLD and for future clinical trials.

CONTEXT:There is limited data on the clinical significance of metabolic hyperferritinemia (MHF) based on the most recent consensus. OBJECTIVE:We aimed to validate the clinical outcomes of MHF in the general population and patients with biopsy-proven metabolic dysfunction-associated fatty liver disease (MAFLD). METHODS:The NHANES database and PERSONS cohort were included. MHF was defined as elevated serum ferritin with metabolic dysfunction (MD) and stratified into different grades according to ferritin (grade 1: 200 [females]/300 [males]-550 ng/mL; grade 2: 550-1000 ng/mL; grade 3: >1000 ng/mL). The clinical outcomes, including all-cause death, comorbidities, and liver histology, were compared between non-MHF and MHF in adjusted models. RESULTS:In NHANES, compared with non-MHF with MD, MHF was related to higher risks of advanced fibrosis (P = .036), elevated albumin-creatinine ratio (UACR, P = .001), and sarcopenia (P = .013). Although the association between all grades of MHF and mortality was insignificant (P = .122), grades 2/3 was associated with increased mortality (P = .029). When comparing with non-MHF without MD, the harmful effects of MHF were more significant in mortality (P < .001), elevated UACR (P < .001), cardiovascular disease (P = .028), and sarcopenia (P < .001). In the PERSONS cohort, MHF was associated with more advanced grades of steatosis (P < .001), lobular inflammation (P < .001), advanced fibrosis (P = .017), and more severe hepatocellular iron deposition (P < .001). CONCLUSION:Both in the general population and in at-risk individuals with MAFLD, MHF was related with poorer clinical outcomes.

BACKGROUND:Fibrotic metabolic dysfunction-associated steatohepatitis (MASH) is a condition at risk of progressing to advanced liver disease. We examined whether an innovative exhaled nitric oxide (eNO) breath test (BT) can accurately diagnose fibrotic MASH without requiring blood tests. METHODS:One hundred and forty-seven patients with MASH were recruited, and all tests were undertaken within 1 week of recruitment. With fibrotic MASH (NAS ≥ 4 and fibrosis stage ≥ 2) as the main outcome indicator, the diagnostic efficacy of eNO in identifying fibrotic MASH was compared to other validated models for advanced fibrosis requiring venesection, namely FAST, Agile 3+, and FIB-4 scores. RESULTS:The mean age was 40.36 ± 12.28 years, 73.5% were men. Mean body mass index was 28.83 ± 4.31 kg/m2. The proportion of fibrotic MASH was 29.25%. The area under the receiver operating curve for eNO in diagnosing fibrotic MASH was 0.737 [95% CI 0.650-0.823], which was comparable to FAST (0.751 [0.656-0.846]), Agile 3+ (0.764 [0.670-0.858]), and FIB-4 (0.721 [0.620-0.821]) (all DeLong test p > 0.05). A cut-off of eNO <8.5 ppb gave a sensitivity of 86.0% and a negative predictive value of 88.5% for ruling-out fibrotic MASH. A cut-off of eNO >13.5 ppb provided a specificity of 91.3% and a positive predictive value of 65.4% for ruling-in fibrotic MASH. Sensitivity analyses demonstrated that the diagnostic efficacy of eNO was similar across characteristics such as age. Moreover, adding vibration-controlled transient elastography-LSM (liver stiffness measurement) reduced the uncertainty interval from 46.9% to 39.5%. CONCLUSIONS:The eNO-BT is a promising simple test for non-invasively identifying fibrotic MASH, and its performance is further improved by adding LSM measurement.

Although the host immune response is likely to be important for the prognosis of ENKTL, detailed information on the pre-treatment T lymphocyte subsets in ENKTL is lacking. To improve risk stratification for ENKTL patients, it is essential to look at the prognostic relevance of absolute CD3 + T cell counts (ACD3C), CD4 + T cell counts (ACD4C), and CD8 + T cell counts (ACD8C) for ENKTL. We retrospectively analyzed 46 ENKTL patients in the First Affiliated Hospital of Wenzhou Medical University between December 2016 and June 2022. Kaplan-Meier curves and log-rank tests were used to compare survival rates between groups according to the cut-off values of pre-treatment T lymphocyte subsets. Independent prognostic factors for survival were analyzed by Cox regression. ACD3C, ACD4C, and ACD8C were related to overall survival (OS) and progression-free survival (PFS) in ENKTL patients. Multivariate analyses identified pre-treatment ACD3C, ACD4C, and ACD8C as independent prognostic factors of survival, independent of the International Prognostic Index (IPI), prognostic index of natural killer lymphoma (PINK), and nomogram-revised risk index (NRI). The prognostic models incorporating pre-treatment T lymphocyte subsets and serum lactate dehydrogenase (LDH) could be used to stratify ENKTL patients into different prognostic groups with significantly different survivals. When superimposed on the IPI, PINK, or NRI categories, the ACD3C-LDH, ACD4C-LDH, and ACD8C-LDH models could better identify high-risk patients in the low-risk IPI, PINK, or NRI categories. In conclusion, the pre-treatment ACD3C, ACD4C, and ACD8C are effective prognostic survival indicators in ENKTL patients. When combined with LDH, they could better identify high-risk ENKTL patients.

BACKGROUND:Metabolic dysfunction-associated fatty liver disease (MAFLD) is an emerging risk factor for chronic kidney disease (CKD). N-terminal propeptide of collagen type 3 (PRO-C3) is a biomarker of advanced fibrosis in MAFLD and PRO-C3 may be involved in renal fibrosis. We aimed to use PRO-C3 measurements to generate a new algorithmic score to test the prediction of MAFLD with chronic kidney disease (MAFLD-CKD).METHODS:A derivation and independent validation cohort of 750 and 129 Asian patients with biopsy-confirmed MAFLD were included. Serum PRO-C3 concentration was measured and regression analyses were performed to examine associations with MAFLD-CKD. A derivative algorithm for MAFLD-CKD risk prediction was evaluated with receiver operator characteristic (ROC) curve analysis.RESULTS:The study included two Asian cohorts (n = 180 with MAFLD-CKD; mean-eGFR: 94.93 mL/min/1.73 m2; median-urinary albumin-to-creatinine ratio: 6.58 mg/mmol). PRO-C3 was associated with the severity of MAFLD-CKD and independently associated with MAFLD-CKD (adjusted odds ratio = 1.16, 95% confidence interval [CI]: 1.08-1.23, p < .001). A new non-invasive score (termed PERIOD) including PRO-C3 efficiently predicted MAFLD-CKD (AUROC = .842, 95% CI: .805-.875). Accuracy, specificity and negative predictive values were 80.2%, 85.1% and 88.4%, respectively. In the validation cohort, the PERIOD score had good diagnostic performance (AUROC = .807, 95% CI: .691-.893) with similar results in all patient subgroups. In the MAFLD-CKD subgroup, the accuracy for identifying advanced fibrosis was further improved by combining the PRO-C3-based ADAPT with the Agile 3+ scores (AUROC = .90, 95% CI: .836-.964).CONCLUSIONS:The PERIOD score is helpful for accurately predicting the risk of MAFLD-CKD. PRO-C3 can also be used to assess liver fibrosis in people with MAFLD-CKD.

BACKGROUND AND AIMS:Lifestyle intervention is the mainstay of therapy for metabolic dysfunction-associated steatohepatitis (MASH), and liver fibrosis is a key consequence of MASH that predicts adverse clinical outcomes. The placebo response plays a pivotal role in the outcome of MASH clinical trials. Second harmonic generation/two-photon excitation fluorescence (SHG/TPEF) microscopy with artificial intelligence analyses can provide an automated quantitative assessment of fibrosis features on a continuous scale called qFibrosis. In this exploratory study, we used this approach to gain insight into the effect of lifestyle intervention-induced fibrosis changes in MASH. METHODS:We examined unstained sections from paired liver biopsies (baseline and end-of-intervention) from MASH individuals who had received either routine lifestyle intervention (RLI) (n = 35) or strengthened lifestyle intervention (SLI) (n = 17). We quantified liver fibrosis with qFibrosis in the portal tract, periportal, transitional, pericentral, and central vein regions. RESULTS:About 20% (7/35) and 65% (11/17) of patients had fibrosis regression in the RLI and SLI groups, respectively. Liver fibrosis tended towards no change or regression after each lifestyle intervention, and this phenomenon was more prominent in the SLI group. SLI-induced liver fibrosis regression was concentrated in the periportal region. CONCLUSION:Using digital pathology, we could detect a more pronounced fibrosis regression with SLI, mainly in the periportal region. With changes in fibrosis area in the periportal region, we could differentiate RLI and SLI patients in the placebo group in the MASH clinical trial. Digital pathology provides new insight into lifestyle-induced fibrosis regression and placebo responses, which is not captured by conventional histological staging.

AIMS:To explore the associations between cuprotosis-related genes (CRGs) across different stages of liver disease in metabolic dysfunction-associated fatty liver disease (MAFLD), including hepatocellular carcinoma (HCC). MATERIALS AND METHODS:We analysed several bulk RNA sequencing datasets from patients with MAFLD (n = 331) and MAFLD-related HCC (n = 271) and two MAFLD single-cell RNA sequencing datasets. To investigate the associations between CRGs and MAFLD, we performed differential correlation, logistic regression and functional enrichment analyses. We also validated the findings in an independent Wenzhou PERSONS cohort of MAFLD patients (n = 656) used for a genome-wide association study (GWAS). RESULTS:GLS, GCSH and ATP7B genes showed significant differences across the MAFLD spectrum and were significantly associated with liver fibrosis stages. GLS was closely associated with fibrosis stages in patients with MAFLD and those with MAFLD-related HCC. GLS is predominantly expressed in monocytes and T cells in MAFLD. During the progression of metabolic dysfunction-associated fatty liver to metabolic-associated steatohepatitis, GLS expression in T cells decreased. GWAS revealed that multiple single nucleotide polymorphisms in GLS were associated with clinical indicators of MAFLD. CONCLUSIONS:GLS may contribute to liver inflammation and fibrosis in MAFLD mainly through cuprotosis and T-cell activation, promoting the progression of MAFLD to HCC. These findings suggest that cuprotosis may play a role in MAFLD progression, potentially providing new insights into MAFLD pathogenesis.

Background:Animal organ meat (offal) is a food with high nutrient density that is popular in different parts of the world, but its relationship with nonalcoholic steatohepatitis (NASH) is unclear. We aimed to examine whether daily animal organ meat consumption is associated with the presence of NASH in individuals with nonalcoholic fatty liver disease (NAFLD).Methods:A total of 136 Chinese adults with biopsy-proven NAFLD were included. Definite NASH was defined as NAFLD activity score ≥4 and at least one point for steatosis, ballooning, and lobular inflammation. Daily animal organ meat consumption was estimated using a self-administered validated food frequency questionnaire. Logistic regression analysis was performed to assess the association between animal organ meat intake and liver disease severity.Results:The 136 participants (80.9% men) of the study had a mean ± standard deviation (SD) age of 39.0±12.5 years and body mass index of 27.4±3.6 kg/m2. Prevalence of definite NASH was 65.4%. Daily median organ meat consumption was 1.30 g/1,000 kcal. Animal organ meat consumption was inversely associated with the presence of NASH even after adjustment of demographics, lifestyle variables, metabolic and dietary factors, as well as liver fibrosis stage; adjusted-odds ratios (95% confidence intervals) for NASH were 0.15 (0.03, 0.69) for the highest tertile and 0.18 (0.05, 0.70) for the medium tertile, compared to the lowest (reference) tertile of animal organ meat intake (P value for trend =0.024).Conclusions:Our results suggest for the first time that higher animal organ meat consumption is associated with a lower prevalence of NASH in Chinese individuals with biopsy-proven NAFLD.

Background:There is an unmet need for accurate non-invasive methods to diagnose non-alcoholic steatohepatitis (NASH). Since impedance-based measurements of body composition are simple, repeatable and have a strong association with non-alcoholic fatty liver disease (NAFLD) severity, we aimed to develop a novel and fully automatic machine learning algorithm, consisting of a deep neural network based on impedance-based measurements of body composition to identify NASH [the bioeLectrical impEdance Analysis foR Nash (LEARN) algorithm].Methods:A total of 1,259 consecutive subjects with suspected NAFLD were screened from six medical centers across China, of which 766 patients with biopsy-proven NAFLD were included in final analysis. These patients were randomly subdivided into the training and validation groups, in a ratio of 4:1. The LEARN algorithm was developed in the training group to identify NASH, and subsequently, tested in the validation group.Results:The LEARN algorithm utilizing impedance-based measurements of body composition along with age, sex, pre-existing hypertension and diabetes, was able to predict the likelihood of having NASH. This algorithm showed good discriminatory ability for identifying NASH in both the training and validation groups [area under the receiver operating characteristics (AUROC): 0.81, 95% CI: 0.77-0.84 and AUROC: 0.80, 95% CI: 0.73-0.87, respectively]. This algorithm also performed better than serum cytokeratin-18 neoepitope M30 (CK-18 M30) level or other non-invasive NASH scores (including HAIR, ION, NICE) for identifying NASH (P value <0.001). Additionally, the LEARN algorithm performed well in identifying NASH in different patient subgroups, as well as in subjects with partial missing body composition data.Conclusions:The LEARN algorithm, utilizing simple easily obtained measures, provides a fully automated, simple, non-invasive method for identifying NASH.

Dysregulated bile acid (BA) metabolism has been linked to steatosis, inflammation, and fibrosis in nonalcoholic fatty liver disease (NAFLD).

Hepatocytic ballooning is a key histological feature in the diagnosis of non-alcoholic steatohepatitis (NASH) and is an essential component of the two most widely used histological scoring systems for diagnosing and staging non-alcoholic fatty liver disease (NAFLD) [namely, the NAFLD activity score (NAS), and the steatosis, activity and fibrosis (SAF) scoring system]. As a result of the increasing incidence of NASH globally, the diagnostic challenges of hepatocytic ballooning are unprecedented. Despite the clear pathological concept of hepatocytic ballooning, there are still challenges in assessing hepatocytic ballooning in 'real life' situations. Hepatocytic ballooning can be confused with cellular oedema and microvesicular steatosis. Significant inter-observer variability does exist in assessing the presence and severity of hepatocytic ballooning. In this review article, we describe the underlying mechanisms associated with hepatocytic ballooning. Specifically, we discuss the increased endoplasmic reticulum stress and the unfolded protein response, as well as the rearrangement of the intermediate filament cytoskeleton, the appearance of Mallory-Denk bodies and activation of the sonic Hedgehog pathway. We also discuss the use of artificial intelligence in the detection and interpretation of hepatocytic ballooning, which may provide new possibilities for future diagnosis and treatment.

BACKGROUND:Liver fibrosis is the strongest histological risk factor for liver-related complications and mortality in metabolic dysfunction-associated fatty liver disease (MAFLD). Second harmonic generation/two-photon excitation fluorescence (SHG/TPEF) is a powerful tool for label-free two-dimensional and three-dimensional tissue visualisation that shows promise in liver fibrosis assessment.AIM:To investigate combining multi-photon microscopy (MPM) and deep learning techniques to develop and validate a new automated quantitative histological classification tool, named AutoFibroNet (Automated Liver Fibrosis Grading Network), for accurately staging liver fibrosis in MAFLD.METHODS:AutoFibroNet was developed in a training cohort that consisted of 203 Chinese adults with biopsy-confirmed MAFLD. Three deep learning models (VGG16, ResNet34, and MobileNet V3) were used to train pre-processed images and test data sets. Multi-layer perceptrons were used to fuse data (deep learning features, clinical features, and manual features) to build a joint model. This model was then validated in two further independent cohorts.RESULTS:AutoFibroNet showed good discrimination in the training set. For F0, F1, F2 and F3-4 fibrosis stages, the area under the receiver operating characteristic curves (AUROC) of AutoFibroNet were 1.00, 0.99, 0.98 and 0.98. The AUROCs of F0, F1, F2 and F3-4 fibrosis stages for AutoFibroNet in the two validation cohorts were 0.99, 0.83, 0.80 and 0.90 and 1.00, 0.83, 0.80 and 0.94, respectively, showing a good discriminatory ability in different cohorts.CONCLUSION:AutoFibroNet is an automated quantitative tool that accurately identifies histological stages of liver fibrosis in Chinese individuals with MAFLD.

Background and aim Copper is an essential trace element involved in oxidative stress reactions and energy metabolism. While nonalcoholic fatty liver disease (NAFLD) is closely related to metabolic dysfunction, the role of copper in the development of simple steatosis (NAFL) and nonalcoholic steatohepatitis (NASH) is still unclear. We aimed to compare serum copper levels between patients with simple steatosis and those with NASH. Methods and results We studied 102 patients with biopsy-proven NASH (cases) and 102 NAFL controls, who were matched for age, sex, and residential city. Multivariable conditional logistic analysis was performed to explore associations between serum copper levels and the presence of NASH. Serum copper levels were significantly lower in patients with NASH than in those with matched NAFL controls (15.53 ± 2.41 μmol/l vs. 16.34 ± 3.23 μmol/l; P = 0.029). This intergroup difference in serum copper levels was more pronounced in men than in women. The per unit, per SD, and per doubling of serum copper levels were associated, respectively, with an approximately 20, 40, and 90% decrease in risk of having NASH, even after adjustment for potential confounding factors. Conclusion Lower serum copper concentrations are significantly associated with higher prevalence of NASH among biopsied-proven NAFLD patients, particularly in men.

With metabolic dysfunction-associated fatty liver disease (MAFLD) incidence and prevalence sharply increasing globally, there is an urgent need for non-invasive diagnostic tests to accurately screen high-risk MAFLD patients for liver inflammation and fibrosis. We aimed to develop a novel sequential algorithm based on N-terminal propeptide of type 3 collagen (PRO-C3) for disease risk stratification in patients with MAFLD. A derivation and independent validation cohort of 327 and 142 patients with biopsy-confirmed MAFLD were studied. We compared the diagnostic performances of various non-invasive scores in different disease states, and a novel sequential algorithm was constructed by combining the best performing non-invasive scores. For patients with high-risk progressive steatohepatitis (i.e., steatohepatitis + NAFLD activity score ≥ 4 + F ≥ 2), the AUROC of FAST score was 0.801 (95

Background: With metabolic dysfunction-associated fatty liver disease (MAFLD) incidence and prevalence increasing, it is necessary to identify patients with advanced fibrosis (F3-F4 stages). We evaluated the performance of new biomarkers and algorithms for diagnosing advanced fibrosis in an Asian population. Methods: Data from two Asian cohorts (including 851 biopsy-proven MAFLD [578 from Wenzhou, 273 from Hong Kong]) were studied. The association between N-terminal propeptide of type 3 collagen (PRO-C3) and the histologic stage of liver fibrosis was analyzed by multivariable linear regression. The area under the receiver operating characteristic curve (AUROC) was used to test the diagnostic performance of serum PRO-C3 and the ADAPT score for advanced fibrosis and compared them to other established non-invasive tests. Results: Serum PRO-C3 levels increased progressively across liver fibrosis stages and correlated with advanced fibrosis (P < 0.001). The ADAPT score had an AUROC of 0.865 (95% confidence interval 0.829-0.901) for advanced fibrosis; the accuracy, sensitivity and negative predictive values were 81.4%, 82.2% and 96.1%, respectively. This result was better compared to that of PRO-C3 alone or other non-invasive fibrosis biomarkers (aspartate aminotransferase-to-platelet ratio index, Fibrosis-4, BARD, and NAFLD fibrosis score). In subgroup analyses (including sex, age, diabetes, NAFLD activity score, body mass index or serum alanine aminotransferase levels), the ADAPT score had good diagnostic performance. Conclusion: PRO-C3 and the ADAPT score reliably exclude advanced fibrosis in MAFLD patients and reduce the need for liver biopsy. (c) 2021 Elsevier Inc. All rights reserved.

Background and Aims Non-alcoholic fatty liver disease (NAFLD) is a usual chronic liver disease and lacks non-invasive biomarkers for the clinical diagnosis and prognosis. Extracellular vesicles (EVs), a group of heterogeneous small membrane-bound vesicles, carry proteins and nucleic acids as promising biomarkers for clinical applications, but it has not been well explored on their lipid compositions related to NAFLD studies. Here, we investigate the lipid molecular function of urinary EVs and their potential as biomarkers for non-alcoholic steatohepatitis (NASH) detection. Methods This work includes 43 patients with non-alcoholic fatty liver (NAFL) and 40 patients with NASH. The EVs of urine were isolated and purified using the EXODUS method. The EV lipidomics was performed by LC-MS/MS. We then systematically compare the EV lipidomic profiles of NAFL and NASH patients and reveal the lipid signatures of NASH with the assistance of machine learning. Results By lipidomic profiling of urinary EVs, we identify 422 lipids mainly including sterol lipids, fatty acyl lipids, glycerides, glycerophospholipids, and sphingolipids. Via the machine learning and random forest modeling, we obtain a biomarker panel composed of 4 lipid molecules including FFA (18:0), LPC (22:6/0:0), FFA (18:1), and PI (16:0/18:1), that can distinguish NASH with an AUC of 92.3%. These lipid molecules are closely associated with the occurrence and development of NASH. Conclusion The lack of non-invasive means for diagnosing NASH causes increasing morbidity. We investigate the NAFLD biomarkers from the insights of urinary EVs, and systematically compare the EV lipidomic profiles of NAFL and NASH, which holds the promise to expand the current knowledge of disease pathogenesis and evaluate their role as non-invasive biomarkers for NASH diagnosis and progression.

BACKGROUND AND AIMS:Zinc is an essential trace element that plays an important role in maintaining health, and affecting gene expression, signal transduction and regulation of apoptosis. It is uncertain whether serum zinc levels are altered in patients with metabolic dysfunction-associated fatty liver disease (MAFLD). We aimed to investigate the association between serum zinc levels and the severity of hepatic necro-inflammation (HN) in patients with MAFLD. METHODS AND RESULTS:Liver disease severity was graded histologically using the NAFLD activity score. HN was defined as the sum of ballooning and lobular inflammation. We used a smooth function regression model to analyze the relationship between serum zinc levels and HN. A total of 561 (76.5% men) patients with biopsy-confirmed MAFLD were enrolled. They had a mean age of 41.3 years, and a mean serum zinc level of 17.0 ± 4.1 μmol/L. Compared to those with mild hepatic necro-inflammation (MHN, grades 0-2; n = 286), patients with severe hepatic necro-inflammation (SHN, grades 3-5; n = 275) had lower serum zinc concentrations (16.3 ± 4.2 vs. 17.6 ± 4.0 μmol/L; p < 0.001). However, a threshold saturation effect analysis showed that there was an inflection in serum zinc levels at 24 μmol/L. After adjustment for potential confounders, serum zinc levels <24 μmol/L were inversely associated with SHN (adjusted-odds ratio 0.88, 95%CI 0.83-0.93; p < 0.001), whereas serum zinc levels >24 μmol/L were positively associated with SHN (adjusted-odds ratio 1.42, 95%CI: 1.03-1.97; p = 0.035). CONCLUSIONS:There is a J-shaped relationship between serum zinc levels and the severity of hepatic necro-inflammation in patients with biopsy-proven MAFLD.