Nonalcoholic Fatty Liver Disease Liver Research Articles

Simultaneously Screening for Liver Steatosis and Fibrosis in Romanian Type 2 Diabetes Mellitus Patients Using Vibration-Controlled Transient Elastography with Controlled Attenuation Parameter.

Non-alcoholic fatty liver disease (NAFLD) is a common finding among patients with type 2 diabetes mellitus (T2DM). Between NAFLD and T2DM exist a bidirectional relationship. Patients with T2DM are at high risk for NAFLD, and evidence suggests that T2DM is linked to progressive NAFLD and poor liver outcomes. NAFLD promotes the development of T2DM and leads to a substantial increase in the risk of T2DM complications. This study aimed to assess the prevalence of liver steatosis and fibrosis in patients with T2DM from north-eastern Romania by using Vibration-Controlled Transient Elastography (VCTE) with Controlled Attenuation Parameter (CAP), which is a non-invasive method and can assess simultaneously liver steatosis and fibrosis. In total, 424 consecutive patients with T2DM were enrolled and evaluated using VCTE with CAP from January 2020 to January 2022. Clinical and laboratory data were recorded in all patients. For the CAP score, we used the following cut-offs: mild steatosis (S1)—274 dB/m, moderate steatosis (S2)—290 dB/m, and severe steatosis (S3)—302 dB/m. For liver fibrosis, to differentiate between fibrosis stages, the cut-off values were F ≥ 8.2 kPa for significant fibrosis (F2), F ≥ 9.7 kPa for advanced fibrosis (F3), and F ≥ 13.6 kPa for cirrhosis (F4). In total, 380 diabetic patients (72.6%) had liver steatosis (51.3% females, the mean age of 55.22 ± 10.88 years, mean body mass index (BMI) 29.12 ± 5.64 kg/m2). Among them, 26 (8.4%) patients had moderate liver steatosis (S2) and 242 (78.5%) patients had severe hepatic steatosis (S3). According to VCTE measurements, 176 (57.14%) patients had liver fibrosis, 36 (11.7%) of them had advanced fibrosis (F3), and 42 (13.6%) diabetic patients had cirrhosis (F4). Univariate analyses showed that severe steatosis was significantly associated with ferritin (β = 0.223, p = 0.022), total cholesterol (β = 0.159, p = 0.031), and HDL-cholesterol (β = −0.120, p = 0.006). In multivariate analyses, BMI (β = 0.349, p < 0.001), fasting plasma glucose (β = 0.211, p = 0.006), and triglycerides (β = 0.132, p = 0.044) were predictors of S3. Patients with T2DM have a high prevalence of severe steatosis and advanced fibrosis which can lead to the development and progression of complications with high morbidity and mortality rates. Hence, it is necessary to implement screening strategies to prevent advanced liver disease in patients with T2DM.

MicroRNA-103a-3p promotes inflammation and fibrosis in nonalcoholic fatty liver disease by targeting HBP1

Background As a metabolic-associated disease, non-alcoholic fatty liver disease (NAFLD) development is tightly linked to lipid accumulation, inflammatory response, and fibrosis. Our study was intended to expound the role of microRNA (miR)-103a-3p in the pathogenesis of NAFLD. Methods First, potentially relevant genes in NAFLD were screened using microarray analysis. Expression of lipid metabolism-related, inflammatory and liver fibrosis indicators in the serum of patients with NAFLD was analyzed. We established a NAFLD mouse model and analyzed the serum level of lipid metabolism- and inflammation-related factors and fibrosis in the liver tissues of NAFLD mice. The targeting relationship between miR-103a-3p and HBP1 was examined by dual-luciferase reporter gene assay, RT-qPCR and Western blot. Finally, the simultaneous effects of miR-103a-3p and HBP1 on lipid metabolism, inflammatory response and liver fibrosis in NAFLD mice were analyzed by rescue experiments. Results MiR-103a-3p was upregulated in the serum of NAFLD patients and liver tissues of NAFLD mice, with increased lipid accumulation, inflammation and liver fibrosis. HBP1 was reduced in liver tissues of NAFLD mice, and miR-103a-3p bound to and negatively regulated HBP1. Inhibition of miR-103a-3p or promotion of HBP1 improved liver function, decreased lipid accumulation, suppressed inflammatory response, and reduced liver fibrosis in NAFLD mice. Moreover, sh-HBP1 partially reversed the effect of miR-103a-3p inhibitor on NAFLD mice, leading to increased lipid accumulation, elevated inflammation and liver fibrosis in the liver of mice. Conclusions miR-103a-3p inhibits the expression of HBP1, thus suppressing lipid metabolism, stimulating inflammatory responses, and promoting liver fibrosis in NAFLD.

Improved body composition is associated with reduced steatosis in non-alcoholic fatty liver disease, a systematic review and meta-analysis of longitudinal studies

Abstract Background: Based on cross-sectional studies, there is a link between body composition parameters and steatosis in non-alcoholic fatty liver disease (NAFLD). However, whether long-term changes in different body composition parameters will result in NAFLD resolution is unclear. Therefore, we aimed to summarize the literature on longitudinal studies evaluating the association between NAFLD resolution and body composition change. Methods: Based on the recommendations of the Cochrane Handbook, we performed a systematic search on September 26th, 2021, in four databases: Embase, MEDLINE (via PubMed), and Cochrane Central Register of Controlled Trials (CENTRAL). Eligible studies reported on patients with NAFLD (liver fat &gt;5%) and examined the correlation between body composition improvement and a decrease in steatosis. We did not have pre-defined body composition or steatosis measurement criteria. Next, we calculated pooled correlation coefficient (r) with a 95% confidence interval (CI). Results: We identified 15 eligible studies, of which 5 five studies were included in our quantitative synthesis. Based on two studies with 85 patients, we found a pooled correlation coefficient of r=0.49 (CI: 0.22-0.69, Spearman’s correlation) between the change of visceral adipose tissue and liver steatosis. Similarly, based on three studies with 175 patients, the correlation was r=0.33 (CI: 0.19-0.46, Pearson’s correlation). On the other hand, based on two studies with 163 patients, the correlation between subcutaneous adipose tissue change and liver steatosis change was r=0.42 (CI: 0.29-0.54, Pearson’s correlation). Conclusion: Based on the included studies, body composition improvement is associated with a decrease in liver fat content in NAFLD.

Non-alcoholic fatty liver disease and sarcopenia is associated with the risk of albuminuria independent of insulin resistance, and obesity

BackgroundAlthough non-alcoholic fatty liver disease (NAFLD) is associated with metabolic disorders, its influence on albuminuria has not been determined. The aim of this study was to identify the relationship between NAFLD and albuminuria in the general Korean population. MethodsData from the Korea National Health and Nutrition Examination Surveys (KNHANES) of 2008–2011 were analyzed (n = 1795). Albuminuria was defined as an albumin-to-creatinine ratio of ≥30 mg/g in random spot urine samples. NAFLD was defined as a fatty liver index (FLI) ≥60 or NAFLD liver fat score (LFS) > −0.64. ResultsA total of 289 (16.1 %) subjects were classified as having albuminuria. Subjects with NAFLD exhibited a higher rate of albuminuria than subjects without NAFLD (crude odds ratios [ORs] = 2.60–2.95, all P < 0.001). Regardless of hypertension, insulin resistance, or obesity, the risk for albuminuria was higher in the NAFLD group than in the group without NAFLD (measured by either FLI or LFS; all P < 0.001). When subjects with NAFLD had sarcopenia, the risk of albuminuria further increased (OR = 4.33–4.64, all P < 0.001). Multiple logistic regression analyses also demonstrated that NAFLD was independently associated with albuminuria (OR = 2.58, 95 % confidence interval [CI] = 1.66–4.02, P < 0.001 for FLI, OR = 1.87, 95 % CI = 1.28–2.75, P = 0.001 for LFS). ConclusionsNAFLD was associated with an increased risk of albuminuria in the general Korean population. This association was independent of hypertension, insulin resistance, chronic kidney disease, diabetes and obesity, and stronger in subjects with sarcopenia.

Acetyl-CoA metabolism drives epigenome change and contributes to carcinogenesis risk in fatty liver disease

BackgroundThe incidence of non-alcoholic fatty liver disease (NAFLD)-associated hepatocellular carcinoma (HCC) is increasing worldwide, but the steps in precancerous hepatocytes which lead to HCC driver mutations are not well understood. Here we provide evidence that metabolically driven histone hyperacetylation in steatotic hepatocytes can increase DNA damage to initiate carcinogenesis.MethodsGlobal epigenetic state was assessed in liver samples from high-fat diet or high-fructose diet rodent models, as well as in cultured immortalized human hepatocytes (IHH cells). The mechanisms linking steatosis, histone acetylation and DNA damage were investigated by computational metabolic modelling as well as through manipulation of IHH cells with metabolic and epigenetic inhibitors. Chromatin immunoprecipitation and next-generation sequencing (ChIP-seq) and transcriptome (RNA-seq) analyses were performed on IHH cells. Mutation locations and patterns were compared between the IHH cell model and genome sequence data from preneoplastic fatty liver samples from patients with alcohol-related liver disease and NAFLD.ResultsGenome-wide histone acetylation was increased in steatotic livers of rodents fed high-fructose or high-fat diet. In vitro, steatosis relaxed chromatin and increased DNA damage marker γH2AX, which was reversed by inhibiting acetyl-CoA production. Steatosis-associated acetylation and γH2AX were enriched at gene clusters in telomere-proximal regions which contained HCC tumour suppressors in hepatocytes and human fatty livers. Regions of metabolically driven epigenetic change also had increased levels of DNA mutation in non-cancerous tissue from NAFLD and alcohol-related liver disease patients. Finally, genome-scale network modelling indicated that redox balance could be a key contributor to this mechanism.ConclusionsAbnormal histone hyperacetylation facilitates DNA damage in steatotic hepatocytes and is a potential initiating event in hepatocellular carcinogenesis.

Abstract 1372: Macrophages impair antigen specific CD8+ T cell response against HCC in NAFLD mice

Abstract Background: Liver cancer is a leading cause of cancer-related deaths. Non-alcoholic fatty liver disease (NAFLD) has become a burgeoning etiology of liver cancer. Emerging data shows that NAFLD alters adaptive T cell immunity and has a profound influence on both liver cancer development and efficacy of immunotherapy. Tumor antigen-specific T cells recognize tumor and are crucial for controlling malignancy. However, their populations are often low, and detection requires special tools. This study utilized a novel mouse model of HCC to investigate how the tumor antigen-specific T cell response is affected in NAFLD. Methods: We generated a doxycycline-inducible MHC-I OVA257-264 and MHC-II OVA323-339 antigen-expressing HCC cell line which allowed us to investigate tumor antigen-specific T cell responses. The cell line was seeded orthotopically into the livers of diet-induced (methionine choline-deficient [MCD]) and genetic (Ob/Ob [Ob]) NAFLD mouse models. OVA257-264 specific CD8+ T cells were detected by flow cytometry using H-2Kb/OVA257-264 tetramer. Pharmacological depletion experiments were used to identify additional effector cells. Results: The creation of an inducible MHC-I and -II OVA antigen-expressing HCC cell line was validated in both in-vitro and in-vivo settings. Tumor expression of highly immunogenic OVA antigens caused a strong tumor suppression which was mostly mediated by CD8+ T cells in mice fed with normal diet. The inverse correlation between percentage of OVA257-264 specific CD8+ T cells with tumor size further supported their critical role in tumor control. In contrast, in NAFLD mice fed MCD diet as well as Ob/Ob mice, tumor OVA antigens induction failed to reduce liver tumor growth. The presence of OVA257-264 specific CD8+ T cells in NAFLD mice suggested that NAFLD did not inhibit the generation of tumor antigen-specific T cells. This was supported by the finding that OVA257-264 specific CD8+ T cells from NAFLD mice had similar cytotoxic potential, activation, degranulation, PD-1 expression, and effector memory phenotype compared to controls, suggesting an indirect mechanism for the impaired anti-tumor immunity. Immunoprofiling revealed a drastic increase of macrophages in tumor-bearing NAFLD livers. Depletion of macrophages with clodronate reversed the NAFLD caused resistance to tumor-antigen dependent tumor suppression. It was observed that there were minimal changes to the amount of TAS CD8+ T cells in doxycycline-treated mice irrespective of clodronate depletion. There was, however, higher markers of activation such as CD69 and TNFα in clodronate-depleted mice compared to controls. Conclusions: NAFLD impairs antigen-specific CD8+ T cell immunity against liver tumor. The effect was not due to reduced generation or intrinsic functional changes of tumor antigen-specific CD8+ T cells but caused by accumulated macrophages in the liver environment. Citation Format: John C. McVey, Benjamin L. Green, Benjamin Ruf, Tim F. Greten, Chi Ma. Macrophages impair antigen specific CD8+ T cell response against HCC in NAFLD mice [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1372.

Risk assessment with gut microbiome and metabolite markers in NAFLD development.

A growing body of evidence suggests interplay between the gut microbiota and the pathogenesis of nonalcoholic fatty liver disease (NAFLD). However, the role of the gut microbiome in early detection of NAFLD is unclear. Prospective studies are necessary for identifying reliable, microbiome markers for early NAFLD. We evaluated 2487 individuals in a community-based cohort who were followed up 4.6 years after initial clinical examination and biospecimen sampling. Metagenomic and metabolomic characterizations using stool and serum samples taken at baseline were performed for 90 participants who progressed to NAFLD and 90 controls who remained NAFLD free at the follow-up visit. Cases and controls were matched for gender, age, body mass index (BMI) at baseline and follow-up, and 4-year BMI change. Machine learning models integrating baseline microbial signatures (14 features) correctly classified participants (auROCs of 0.72 to 0.80) based on their NAFLD status and liver fat accumulation at the 4-year follow up, outperforming other prognostic clinical models (auROCs of 0.58 to 0.60). We confirmed the biological relevance of the microbiome features by testing their diagnostic ability in four external NAFLD case-control cohorts examined by biopsy or magnetic resonance spectroscopy, from Asia, Europe, and the United States. Our findings raise the possibility of using gut microbiota for early clinical warning of NAFLD development.

1338-P: Glucoregulatory Disturbances in Autoimmune Liver Disease and Nonalcoholic Fatty Liver Disease Compared with Healthy Individuals

Gluco-regulatory disturbances such as hepatic insulin resistance, hyperinsulinemia and prediabetes are commonly present in patients with nonalcoholic fatty liver disease (NAFLD) and those individuals may over time develop full-blown type 2 diabetes. Chronic liver diseases­ such as NAFLD and autoimmune liver diseases (AILDs) are heterogenous but may affect glucose-metabolism similarly. It is, however, unknown if AILDs—such as primary biliary cholangitis (PBC) —display gluco-regulatory impairments. We therefore investigated glucose and hormonal responses during a 75 g oral glucose tolerance test (OGTT) in patients with biopsy-verified, non-cirrhotic PBC (n = 9, age 55 ± y (mean ± sd) , BMI 31 ± 6 kg/m2 (mean ± sd)) , NAFLD (n = 6, age 38 ± 17 y, BMI 31 ± 4 kg/m2) and healthy controls (n = 8, age 23 ± 3 y, BMI 23 ± 2 kg/m2) . None of the participants had diabetes. In the PBC group, 3 had NAFLD. Fasting glucose, c-peptide and insulin levels were significantly increased in PBC and NAFLD compared with healthy controls ([mean (95 % CI) ]; glucose (mM) 5.6 (4.7-6.7) , 5.7 (5.2-6.1) , 4.7 (3.9-5.6) ; c-peptide (pM) 993 (556-1773) , 1334 (1036-1719) , 483 (268-869) ; insulin (pM) 98 (33-298) , 166 (103-267) , 43 (14-136) ; respectively) . Hepatic insulin resistance (reflected by fasting homeostasis model assessment of insulin resistance (HOMA-IR)) was present in PBC (mean 4.0 (95 % CI 1.2-13.9)) and NAFLD (7.0 (4.1-11.9)) but not in healthy controls (1.5 (0.4-5.4)) . There was no significant difference in glucose levels between the groups. Beta-cell secretion (c-peptide) was significantly increased in PBC and NAFLD. Insulin responses were higher in PBC and NAFLD compared with healthy but only reached statistical significance in NAFLD. Our data suggest that patients with PBC have gluco-regulatory disturbances including hepatic insulin resistance and impaired beta-cell function. Metabolic dysfunction of PBC may be underestimated and warrant further investigation. Disclosure A.H. Jensen: None. H. Ytting: Other Relationship; Gilead Sciences, Inc. J. Grandt: None. M.P. Werge: None. E.B. Rashu: None. L.E. Hetland: None. A. Junker: None. L. Hobolth: None. C. Mortensen: None. F. Tofteng: None. M. Vyberg: None. R. Serizawa: Consultant; Merck Sharp &amp; Dohme Corp. L. Gluud: Advisory Panel; Novo Nordisk. Consultant; Pfizer Inc. Research Support; Alexion Pharmaceuticals, Inc., Gilead Sciences, Inc., Novo Nordisk, Sobi. N.J. Wewer Albrechtsen: Research Support; Mercodia AB, Novo Nordisk, Regeneron Pharmaceuticals Inc. Speaker’s Bureau; Merck &amp; Co., Inc., Mercodia AB. Funding Nicolai J. Wewer Albrechtsen were financed by NNF Excellence Emerging Investigator Grant – Endocrinology and Metabolism (Application No. NNF19OC0055001) , EFSD Future Leader Award (NNF21SA0072746) and DFF Sapere Aude

Aberrant cholesterol metabolic signaling impairs antitumor immunosurveillance through natural killer T cell dysfunction in obese liver.

Obesity is a major risk factor for cancers including hepatocellular carcinoma (HCC) that develops from a background of non-alcoholic fatty liver disease (NAFLD). Hypercholesterolemia is a common comorbidity of obesity. Although cholesterol biosynthesis mainly occurs in the liver, its role in HCC development of obese people remains obscure. Using high-fat high-carbohydrate diet-associated orthotopic and spontaneous NAFLD-HCC mouse models, we found that hepatic cholesterol accumulation in obesity selectively suppressed natural killer T (NKT) cell-mediated antitumor immunosurveillance. Transcriptome analysis of human liver revealed aberrant cholesterol metabolism and NKT cell dysfunction in NAFLD patients. Notably, cholesterol-lowering rosuvastatin restored NKT expansion and cytotoxicity to prevent obesogenic diet-promoted HCC development. Moreover, suppression of hepatic cholesterol biosynthesis by a mammalian target of rapamycin (mTOR) inhibitor vistusertib preceded tumor regression, which was abolished by NKT inactivation but not CD8+ T cell depletion. Mechanistically, sterol regulatory element-binding protein 2 (SREBP2)-driven excessive cholesterol production from hepatocytes induced lipid peroxide accumulation and deficient cytotoxicity in NKT cells, which were supported by findings in people with obesity, NAFLD and NAFLD-HCC. This study highlights mTORC1/SREBP2/cholesterol-mediated NKT dysfunction in the tumor-promoting NAFLD liver microenvironment, providing intervention strategies that invigorating NKT cells to control HCC in the obesity epidemic.

A new detection system for serum fragmented cytokeratin 18 as a biomarker reflecting histologic activities of human nonalcoholic steatohepatitis.

Caspase‐generated fragmented cytokeratin 18 (fCK18) is recognized as a useful noninvasive biomarker in the diagnosis of nonalcoholic fatty liver disease (NAFLD), particularly nonalcoholic steatohepatitis (NASH). However, fCK18 measurement is not applied clinically due to widely variable cut‐off values under the current enzyme‐linked immunosorbent assay platform. Therefore, we developed a highly sensitive chemiluminescent enzyme immunoassay using newly developed monoclonal antibodies against fCK18 and investigated its relevance in NASH diagnosis. Serum fCK18 levels were measured in the derivation and validation cohort. The correlation between serum fCK18 levels and NAFLD activity score (NAS), fibrosis stage, and liver function was examined. Serum fCK18 levels were significantly correlated with alanine aminotransferase (ALT), aspartate aminotransferase (AST), and gamma‐glutamyl transpeptidase. Serum fCK18 levels were significantly associated with NAS, Brunt's grade/stage, Matteoni's classification, portal inflammation, and fat accumulation in the liver. Notably, hepatocyte ballooning was the only independent variable significantly associated with serum fCK18 in the multivariate linear regression analysis. Serum fCK18 levels were significantly elevated in patients with NAFLD and nonalcoholic fatty liver (NAFL) compared to healthy individuals. They were also significantly elevated in patients with NAFL compared to NASH defined by NAS or Matteoni's classification, with area under the curve values being 0.961 (NAFLD vs. healthy), 0.913 (NAFL vs. healthy), 0.763 (NASH vs. NAFL), and 0.796 (NASH type 3–4 vs. NAFL type 1–2). These results were confirmed by a validation cohort. Notably, changes over time in serum fCK18 levels were significantly correlated with changes in ALT, AST, and the fibrosis‐4 index in 25 patients who underwent lifestyle modification. Serum fCK18 levels were significantly correlated with liver damage associated with NASH pathology. Serum fCK18 levels are accurate in distinguishing patients with NAFL or NASH from healthy individuals and may be useful to monitor NASH over time.

The Role of Interferon Regulatory Factors in Non-Alcoholic Fatty Liver Disease and Non-Alcoholic Steatohepatitis

Non-alcoholic fatty liver disease (NAFLD) is becoming the most common chronic liver disease with many metabolic comorbidities, such as obesity, diabetes, and cardiovascular diseases. Non-alcoholic steatohepatitis (NASH), an advanced form of NAFLD, accompanies the progression of hepatic steatosis, inflammation, cell death, and varying degree of liver fibrosis. Interferons (IFNs) have been shown to play important roles in the pathogenesis of NAFLD and NASH. Their regulating transcriptional factors such as interferon regulatory factors (IRFs) can regulate IFN expression, as well as genes involved in macrophage polarization, which are implicated in the pathogenesis of NAFLD and advanced liver disease. In this review, the roles of IRF-involved signaling pathways in hepatic inflammation, insulin resistance, and immune cell activation are reviewed. IRFs such as IRF1 and IRF4 are also involved in the polarization of macrophages that contribute to critical roles in NAFLD or NASH pathogenesis. In addition, IRFs have been shown to be regulated by treatments including microRNAs, PPAR modulators, anti-inflammatory agents, and TLR agonists or antagonists. Modulating IRF-mediated factors through these treatments in chronic liver disease can ameliorate the progression of NAFLD to NASH. Furthermore, adenoviruses and CRISPR activation plasmids can also be applied to regulate IRF-mediated effects in chronic liver disease. Pre-clinical and clinical trials for evaluating IRF regulators in NAFLD treatment are essential in the future direction.

New inflammatory biomarkers (lymphocyte and monocyte percentage to high-density lipoprotein cholesterol ratio and lymphocyte to monocyte percentage ratio) and their association with some cardiometabolic diseases : Results from alarge Kurdish cohort study in Iran.

The incidence of metabolic heart diseases has increased significantly in Middle Eastern countries such as Iran. The present study aimed to investigate the association between monocyte percentage to high-density cholesterol ratio (MHR), lymphocyte percentage to high-density cholesterol ratio (LHR), and lymphocyte to monocyte percentage ratio (LMR) and cardiometabolic diseases in aKurdish population in the west of Iran. This study recruited 9803 individuals, 4728 (48.2%) were male and 5084 (51.8%) were female from Ravansar, Iran. All biomarkers were analyzed by the standard methods. The prevalence of cardiometabolic diseases was higher in overweight/obese participants and increased with age. MHR and LHR increased significantly in cardiometabolic individuals compared with healthy controls. Individuals in the fourth quartiles of LHR and MHR had higher odds ratio (ORs) for metabolic syndrome (MetS) and diabetes mellitus (DM) than the first quartiles. The LMR had astatistical association with non-alcoholic fatty liver disease (NAFLD) ORs and FLI. Besides, all these associations were stronger for females, and increased physical activity decreased inflammatory biomarkers. The present study showed MHR and LHR had significant associations with ORs of MetS and DM. Also, MHR and LHR had asignificant positive correlation with cardiometabolic risk factors. The LMR only had astatistical association with NAFLD and fatty liver index (FLI). Besides, the strong correlation between inflammatory biomarkers and cardiometabolic risk factors in females might be relevant to higher fat accumulation and metabolic inflammation background, and lower physical activity.

Association of thyroid autoimmunity with nonalcoholic fatty liver disease in euthyroid middle-aged subjects: A population-based study.

The association between thyroid autoimmunity and nonalcoholic fatty liver disease (NAFLD) remains unclear. In this study, we aimed to investigate the relationship between thyroid autoimmunity and NAFLD in a large cohort of euthyroid subjects. We analyzed clinical and biochemical data from a nationwide, population-based, cross-sectional survey (KNHANES VI). A total of 1589 middle-aged participants aged 45-65years, with normal thyroid function, were included in this study. NAFLD was defined as a hepatic steatosis index of >36. We estimated the odds ratios (ORs) for NAFLD according to anti-thyroid peroxidase antibody (TPOAb) positivity by using logistic regression models, and adjusted for potential confounders. Overall, 24% (n=378) of the subjects had NAFLD. Subjects with NAFLD showed a higher positivity for TPOAb (11% vs 7%, P=0.014) compared with those without NAFLD. TPOAb positivity was a significant risk factor for NAFLD [OR 1.668, 95% confidence interval (CI) 1.019-2.730, P=0.042] even after adjusting for confounding variables, including age, sex, household income, education, smoking, alcohol consumption, walking activity, abdominal obesity, elevated blood pressure, dyslipidemia and hyperglycemia. In addition, TPOAb positivity predicted the risk of advanced liver fibrosis (OR 3.112, 95% CI 1.256-7.713, P=0.014) in subjects with NAFLD, independent of the confounding variables. In euthyroid subjects, thyroid autoimmunity is associated with NAFLD and advanced liver fibrosis, independent of known metabolic risk factors. Large longitudinal studies in the future will help clarify the causality.

Label-free Assessment of the Nascent State of Rat Non-alcoholic Fatty Liver Disease Using Spontaneous Raman Microscopy.

Spontaneous Raman microscopy, which can detect molecular vibrations in cells and tissues, could be a useful tool for the label-free assessment of non-alcoholic fatty liver disease (NAFLD). However, it is unclear whether it can be used to evaluate the nascent state of NAFLD. To address this, we analyzed the Raman spectra of rat liver tissues in the nascent state of NAFLD upon excitation at 532 nm. Raman and histochemical analyses were performed of liver tissues from rats fed a high-fat, high-cholesterol diet (HFHCD). Raman microscopic imaging analysis of formalin-fixed thin tissue slices showed hepatic steatosis, as revealed by the Raman band at 2,854 cm−1, whereas lipid droplets were not detectable by hematoxylin-eosin staining of images until 3 days after feeding a HFHCD. Raman signals of retinol at 1,588 cm−1 emitted from hepatic stellate cells were distributed alongside hepatic cords; the retinol content rapidly decreased after feeding a HFHCD, whereas hepatic lipid content increased inversely. Raman microscopic analysis of the surface of fresh ex vivo livers enabled early detection of lipid accumulation after a 1-day feeding a HFHCD. In conclusion, spontaneous Raman microscopy can be applied to the label-free evaluation of the nascent state of NAFLD liver tissues.

Liver Fibrosis Marker and Postoperative Mortality in Patients Without Overt Liver Disease.

Nonalcoholic fatty liver disease (NAFLD) can progress to advanced fibrosis, which, in the nonsurgical population, is associated with poor hepatic and extrahepatic outcomes. Despite its high prevalence, NAFLD and related liver fibrosis may be overlooked during the preoperative evaluation, and the role of liver fibrosis as an independent risk factor for surgical-related mortality has yet to be tested. The aim of this study was to assess whether fibrosis-4 (FIB-4), which consists of age, aspartate aminotransferase (AST), alanine aminotransferase (ALT), and platelets, a validated marker of liver fibrosis, is associated with postoperative mortality in the general surgical population. A historical cohort of patients undergoing general anesthesia at an academic medical center between 2014 and 2018 was analyzed. Exclusion criteria included known liver disease, acute liver disease or hepatic failure, and alcohol use disorder. FIB-4 score was categorized into 3 validated predefined categories: FIB-4 ≤1.3, ruling out advanced fibrosis; >1.3 and <2.67, inconclusive; and ≥2.67, suggesting advanced fibrosis. The primary analytic method was propensity score matching (FIB-4 was dichotomized to indicate advanced fibrosis), and a secondary analysis included a multivariable logistic regression. Of 19,861 included subjects, 1995 (10%) had advanced fibrosis per FIB-4 criteria. Mortality occurred intraoperatively in 15 patients (0.1%), during hospitalization in 272 patients (1.4%), and within 30 days of surgery in 417 patients (2.1%). FIB-4 ≥2.67 was associated with increased intraoperative mortality (odds ratio [OR], 3.63; 95% confidence interval [CI], 1.25-10.58), mortality during hospitalization (OR, 3.14; 95% CI, 2.37-4.16), and within 30 days from surgery (OR, 2.46; 95% CI, 1.95-3.10), after adjusting for other risk factors. FIB-4 was related to increased mortality in a dose-dependent manner for the 3 FIB-4 categories ≤1.3 (reference), >1.3 and <2.67, and ≥2.67, respectively; during hospitalization (OR, 1.89; 95% CI, 1.34-2.65 and OR, 4.70; 95% CI, 3.27-6.76) and within 30 days from surgery (OR, 1.77; 95% CI, 1.36-2.31 and OR, 3.55; 95% CI, 2.65-4.77). In a 1:1 propensity-matched sample (N = 1994 per group), the differences in mortality remained. Comparing the FIB-4 ≥2.67 versus the FIB-4 <2.67 groups, respectively, mortality during hospitalization was 5.1% vs 2.2% (OR, 2.70; 95% CI, 1.81-4.02), and 30-day mortality was 6.6% vs 3.4% (OR, 2.26; 95% CI, 1.62-3.14). A simple liver fibrosis marker is strongly associated with perioperative mortality in a population without apparent liver disease, and may aid in future surgical risk stratification and preoperative optimization.

Variants in mitochondrial amidoxime reducing component 1 and hydroxysteroid 17-beta dehydrogenase 13 reduce severity of nonalcoholic fatty liver disease in children and suppress fibrotic pathways through distinct mechanisms.

Genome‐wide association studies in adults have identified variants in hydroxysteroid 17‐beta dehydrogenase 13 (HSD17B13) and mitochondrial amidoxime reducing component 1 (MTARC1) as protective against nonalcoholic fatty liver disease (NAFLD). We aimed to test their association with pediatric NAFLD liver histology and investigate their function using metabolomics. A total of 1450 children (729 with NAFLD, 399 with liver histology) were genotyped for rs72613567T>TA in HSD17B13, rs2642438G>A in MTARC1, and rs738409C>G in patatin‐like phospholipase domain‐containing protein 3 (PNPLA3). Genotype–histology associations were tested using ordinal regression. Untargeted hepatic proteomics and plasma lipidomics were performed in a subset of children. We found rs72613567T>TA in HSD17B13 to be associated with lower odds of NAFLD diagnosis (odds ratio, 0.7; 95% confidence interval, 0.6–0.9) and a lower grade of portal inflammation (p < 0.001). rs2642438G>A in MTARC1 was associated with a lower grade of hepatic steatosis (p = 0.02). Proteomics found reduced expression of HSD17B13 in carriers of the protective ‐TA allele. MTARC1 levels were unaffected by genotype. Both variants were associated with down‐regulation of fibrogenic pathways. HSD17B13 perturbs plasma phosphatidylcholines and triglycerides. In silico modeling suggested p.Ala165Thr disrupts the stability and metal binding of MTARC1. Conclusion: Both HSD17B13 and MTARC1 variants are associated with less severe pediatric NAFLD. These results provide further evidence for shared genetic mechanisms between pediatric and adult NAFLD.

Pharmacological systemic analysis of gardenia fructus against non-alcoholic fatty liver disease and validation of animal models.

[Purpose]We aimed to investigate the systemic pharmacological analysis of gardenia fructus (GF) and the proof of concepts. We examined the antioxidant and anti-inflammatory effects in high-fat (HF) diet mice.[Methods]The active compounds of GF and the target genes were identified using the Traditional Chinese Medicine Database and Analysis Platform (oral bioavailability ≥ 30%, Caco-2 permeability ≥ -0.4, and drug-likeness ≥ 0.18). The rats were divided into four groups: untreated group, HF group, HF and metformin (17 mg/kg) treated group, and HF and treated with GF (28 mg/kg) for 8 weeks group. Hepatic lesion changes and markers were analyzed using hematoxylin and eosin staining and immunohistochemistry assay.[Results]In the systemic analysis, we identified 14 active compounds including A, B, and C. From these 14 compounds, 242 biological target genes were identified. The top 10 Gene Ontology were analyzed using GO-biological process analysis: removal of superoxide radicals, regulation of endothelial cell apoptotic process, and cellular response to lipopolysaccharide. GF extracts in high-fat diet-induced non-alcoholic fatty liver disease (NAFLD) mice models significantly regulated hepatic lesion markers, such as mTOR, 8-Hydroxy-2'-deoxyguanosine as well as oxidative stress activities, TGF-β, and phosphorylation of ERK1/2.[Conclusion]These results suggest that GF, as an exercise supplement, can alleviate NAFLD disease or fatty liver inflammation. Further studies are required to verify the synergistic effect of GF treatment combined with exercise, which is known to alleviate NAFLD and fatty liver inflammation.

Recent Epidemiology and Risk Factors of Nonalcoholic Fatty Liver Disease.

Because of the global obesity epidemic, the incidence and prevalence of nonalcoholic fatty liver disease (NAFLD) have increased worldwide, including among Koreans. Recently, the incidence rate of NAFLD in Korea was reported to be 45.1 per 1,000 person-years, and the prevalence as approximately 30% depending on the diagnostic methods used. The incidence of advanced fibrosis and hepatocellular carcinoma, as well as all-cause and liver-related mortality in NAFLD patients has increased substantially, imposing considerable public health costs in Korea. Genetic, demographic, environmental, and clinical factors are involved in the pathogenesis of NAFLD. Some genetic variants, such as patatin-like phospholipase domain-containing 3 (PNPLA-3) and sorting and assembly machinery component 50 (SAMM-50), play a major role in the occurrence of NAFLD. The risk of NAFLD and fibrosis increases with advancing age and in men. Nutritional factors, inadequate exercise, and sleep duration are also associated with increased risk of NAFLD. Obesity is a major risk factor for NAFLD; however, NAFLD in lean individuals has been noted in recent studies. Insulin resistance, type 2 diabetes, and metabolic syndrome and its components are closely associated with NAFLD development and liver fibrosis with various underlying mechanisms. Sarcopenia likely shares a common pathophysiology with NAFLD. The rapidly increasing incidence and prevalence of NAFLD and its complications, as well as the associated healthcare burden, warrant early assessment of NAFLD and its risk factors to prevent NAFLD-related complications in high risk groups.

Transcriptomic profiling of a multiethnic pediatric NAFLD cohort reveals genes and pathways associated with disease.

Nonalcoholic fatty liver disease (NAFLD) is the most common type of chronic liver disease in children. The mechanisms that drive NAFLD disease progression in this specific patient population remain poorly defined. In this study, we obtained liver biopsy samples from a multiethnic cohort of pediatric patients with NAFLD (n = 52, mean age = 13.6 years) and healthy liver controls (n = 5). We analyzed transcriptomic changes associated with NAFLD stages using high‐throughput RNA sequencing. Unsupervised clustering as well as pairwise transcriptome comparison distinguished NAFLD from healthy livers. We identified perturbations in pathways including calcium and insulin/glucose signaling occurring early in NAFLD disease, before the presence of histopathologic evidence of advanced disease. Transcriptomic comparisons identified a 25‐gene signature associated with the degree of liver fibrosis. We also identified expression of the insulin‐like growth factor binding protein (IGFBP) gene family (1/2/3/7) as correlating with disease stages, and it has the potential to be used as a peripheral biomarker in NAFLD. Comparing our data set with publicly available adult and adolescent transcriptomic data, we identified similarities and differences in pathway enrichment and gene‐expression profiles between adult and pediatric patients with NAFLD. Regulation of genes including interleukin‐32, IGFBP1, IGFBP2, and IGFBP7 was consistently found in both NAFLD populations, whereas IGFBP3 was specific to pediatric NAFLD. Conclusion: This paper expands our knowledge on the molecular mechanisms underlying pediatric NAFLD. It identifies potential biomarkers and directs us toward new therapies in this population.

The Negative Bidirectional Interaction Between Climate Change and the Prevalence and Care of Liver Disease: A Joint BSG, BASL, EASL, and AASLD Commentary

The Negative Bidirectional Interaction Between Climate Change and the Prevalence and Care of Liver Disease: A Joint BSG, BASL, EASL, and AASLD Commentary