[Intervention mechanism of ginsenoside Rb_1 on liver steatosis in db/db obese mice based on TLR4/MyD88/NF-κB signaling pathway].

Based on the Toll-like receptor 4(TLR4)/myeloid differentiation factor 88(MyD88)/nuclear factor kappaB(NF-κB) signaling pathway, this study observed the regulatory effect of ginsenoside Rb_1(Rb_1) on liver lipid metabolism in db/db obese mice and explored its potential mechanism. Thirty 6-week-old male db/db mice were randomly divided into a model group, a metformin group, and Rb_1 groups with low, medium, and high doses, with six mice in each group. Additionally, six age-matched male db/m mice were assigned to the normal group. The intervention lasted for five weeks. Body weight, fasting blood glucose, and food intake were mea-sured weekly. At the end of the experiment, serum lipid levels and liver function were detected. Hematoxylin-eosin(HE) staining and oil red O staining were performed to observe pathological changes in liver tissue. Real-time quantitative PCR and immunohistochemistry on paraffin sections were used to detect the mRNA and protein expression of TLR4, MyD88, and NF-κB p65. RESULTS:: showed that compared with the normal group, the model group exhibited significant increases in body weight, liver weight, liver index, epididymal fat mass, epididymal fat index, total cholesterol, low-density lipoprotein cholesterol, liver function parameters, and fasting blood glucose levels. Liver lipid accumulation significantly increased, along with elevated mRNA and protein expression of TLR4, MyD88, and NF-κB p65 in the liver. After Rb_1 treatment, the above-mentioned parameters in the intervention groups showed significant reversals. In conclusion, Rb_1 can improve obesity and obesity-related hepatic steatosis in mice while regulating abnormal lipid and glucose meta-bolism. Mechanistically, Rb_1 may improve liver steatosis in db/db obese mice by modulating the TLR4/MyD88/NF-κB signaling pathway.

Polygenic risk score of metabolic dysfunction-associated steatotic liver disease amplifies the health impact on severe liver disease and metabolism-related outcomes.

BACKGROUND: Although the inherited risk factors associated with fatty liver disease are well understood, little is known about the genetic background of metabolic dysfunction-associated steatotic liver disease (MASLD) and its related health impacts. Compared to non-alcoholic fatty liver disease (NAFLD), MASLD presents significantly distinct diagnostic criteria, and epidemiological and clinical features, but the related genetic variants are yet to be investigated. Therefore, we conducted this study to assess the genetic background of MASLD and interactions between MASLD-related genetic variants and metabolism-related outcomes. METHODS: Participants from the UK Biobank were grouped into discovery and replication cohorts for an MASLD genome-wide association study (GWAS), and base and target cohorts for polygenic risk score (PRS) analysis. Autosomal genetic variants associated with NAFLD were compared with the MASLD GWAS results. Kaplan-Meier and Cox regression analyses were used to assess associations between MASLD and metabolism-related outcomes. RESULTS: Sixteen single-nucleotide polymorphisms (SNPs) were identified at genome-wide significance levels for MASLD and duplicated in the replication cohort. Differences were found after comparing these SNPs with the results of NAFLD-related genetic variants. MASLD cases with high PRS had a multivariate-adjusted hazard ratio of 3.15 (95% confidence interval, 2.54-3.90) for severe liver disease (SLD), and 2.81 (2.60-3.03) for type 2 diabetes mellitus. The high PRS amplified the impact of MASLD on SLD and extrahepatic outcomes. CONCLUSIONS: High PRS of MASLD GWAS amplified the impact of MASLD on SLD and metabolism-related outcomes, thereby refining the process of identification of individuals at high risk of MASLD. Supplementation of this process with relevant genetic backgrounds may lead to more effective MASLD prevention and management.

Exploring the butyrate metabolism-related shared genes in metabolic associated steatohepatitis and ulcerative colitis.

Metabolic-associated steatohepatitis (MASH) and ulcerative colitis (UC) exhibit a complex interconnection with immune dysfunction, dysbiosis of the gut microbiota, and activation of inflammatory pathways. This study aims to identify and validate critical butyrate metabolism-related shared genes between both UC and MASH. Clinical information and gene expression profiles were sourced from the Gene Expression Omnibus (GEO) database. Shared butyrate metabolism-related differentially expressed genes (sBM-DEGs) between UC and MASH were identified via various bioinformatics methods. Functional enrichment analysis was performed, and UC patients were categorized into subtypes using the consensus clustering algorithm based on sBM-DEGs. Key genes within sBM-DEGs were screened through Random Forest, Support Vector Machines-Recursive Feature Elimination, and Light Gradient Boosting. The diagnostic efficacy of these genes was evaluated using receiver operating characteristic (ROC) analysis on independent datasets. Additionally, the expression levels of characteristic genes were validated across multiple independent datasets and human specimens. Forty-nine shared DEGs between UC and MASH were identified, with enrichment analysis highlighting significant involvement in immune, inflammatory, and metabolic pathways. The intersection of butyrate metabolism-related genes with these DEGs produced 10 sBM-DEGs. These genes facilitated the identification of molecular subtypes of UC patients using an unsupervised clustering approach. ANXA5, CD44, and SLC16A1 were pinpointed as hub genes through machine learning algorithms and feature importance rankings. ROC analysis confirmed their diagnostic efficacy in UC and MASH across various datasets. Additionally, the expression levels of these three hub genes showed significant correlations with immune cells. These findings were validated across independent datasets and human specimens, corroborating the bioinformatics analysis results. Integrated bioinformatics identified three significant biomarkers, ANXA5, CD44, and SLC16A1, as DEGs linked to butyrate metabolism. These findings offer new insights into the role of butyrate metabolism in the pathogenesis of UC and MASH, suggesting its potential as a valuable diagnostic biomarker.

Clinical and genetic risk factors for progressive fibrosis in metabolic dysfunction-associated steatotic liver disease.

BACKGROUND: Fibrosis-4 (FIB4) is a recommended noninvasive test to assess hepatic fibrosis among patients with metabolic dysfunction-associated steatotic liver disease (MASLD). Here, we used FIB4 trajectory over time (ie, "slope" of FIB4) as a surrogate marker of liver fibrosis progression and examined if FIB4 slope is associated with clinical and genetic factors among individuals with clinically defined MASLD within the Million Veteran Program Cohort. METHODS: In this retrospective cohort study, FIB4 slopes were estimated through linear regression for participants with clinically defined MASLD and FIB4 <2.67 at baseline. FIB4 slope was correlated with demographic parameters and clinical outcomes using logistic regression and Cox proportional hazard models. FIB4 slope as a quantitative phenotype was used in a genome-wide association analysis in ancestry-specific analysis and multiancestry meta-analysis using METAL. RESULTS: FIB4 slopes, generated from 98,361 subjects with MASLD (16,045 African, 74,320 European, and 7996 Hispanic), showed significant associations with sex, ancestry, and cardiometabolic risk factors (p < 0.05). FIB4 slopes also correlated strongly with hepatic outcomes and were independently associated with time to cirrhosis. Five genetic loci showed genome-wide significant associations (p < 5 × 10-8) with FIB4 slope among European ancestry subjects, including 2 known (PNPLA3 and TM6SF2) and 3 novel loci (TERT 5.1 × 10-11; LINC01088, 3.9 × 10-8; and MRC1, 2.9 × 10-9). CONCLUSIONS: Linear trajectories of FIB4 correlated significantly with time to progression to cirrhosis, with liver-related outcomes among individuals with MASLD and with known and novel genetic loci. FIB4 slope may be useful as a surrogate measure of fibrosis progression.

Impaired GK-GKRP interaction rather than direct GK activation worsens lipid profiles and contributes to long-term complications: a Mendelian randomization study.

BACKGROUND: Glucokinase (GK) plays a key role in glucose metabolism. In the liver, GK is regulated by GK regulatory protein (GKRP) with nuclear sequestration at low plasma glucose level. Some GK activators (GKAs) disrupt GK-GKRP interaction which increases hepatic cytoplasmic GK level. Excess hepatic GK activity may exceed the capacity of glycogen synthesis with excess triglyceride formation. It remains uncertain whether hypertriglyceridemia associated with some GKAs in previous clinical trials was due to direct GK activation or impaired GK-GKRP interaction. METHODS: Using publicly available genome-wide association study summary statistics, we selected independent genetic variants of GCKR and GCK associated with fasting plasma glucose (FPG) as instrumental variables, to mimic the effects of impaired GK-GKRP interaction and direct GK activation, respectively. We applied two-sample Mendelian Randomization (MR) framework to assess their causal associations with lipid-related traits, risks of metabolic dysfunction-associated steatotic liver disease (MASLD) and cardiovascular diseases. We verified these findings in one-sample MR analysis using individual-level statistics from the Hong Kong Diabetes Register (HKDR). RESULTS: Genetically-proxied impaired GK-GKRP interaction increased plasma triglycerides, low-density lipoprotein cholesterol and apolipoprotein B levels with increased odds ratio (OR) of 14.6 (95% CI 4.57-46.4) per 1 mmol/L lower FPG for MASLD and OR of 2.92 (95% CI 1.78-4.81) for coronary artery disease (CAD). Genetically-proxied GK activation was associated with decreased risk of CAD (OR 0.69, 95% CI 0.54-0.88) and not with dyslipidemia. One-sample MR validation in HKDR showed consistent results. CONCLUSIONS: Impaired GK-GKRP interaction, rather than direct GK activation, may worsen lipid profiles and increase risks of MASLD and CAD. Development of future GKAs should avoid interfering with GK-GKRP interaction.

Liver microRNA transcriptome reveals miR-182 as link between type 2 diabetes and fatty liver disease in obesity.

Background: The development of obesity-associated comorbidities such as type 2 diabetes (T2D) and hepatic steatosis has been linked to selected microRNAs in individual studies; however, an unbiased genome-wide approach to map T2D induced changes in the miRNAs landscape in human liver samples, and a subsequent robust identification and validation of target genes are still missing. Methods: Liver biopsies from age- and gender-matched obese individuals with (n=20) or without (n=20) T2D were used for microRNA microarray analysis. The candidate microRNA and target genes were validated in 85 human liver samples, and subsequently mechanistically characterized in hepatic cells as well as by dietary interventions and hepatic overexpression in mice. Results: Here, we present the human hepatic microRNA transcriptome of type 2 diabetes in liver biopsies and use a novel seed prediction tool to robustly identify microRNA target genes, which were then validated in a unique cohort of 85 human livers. Subsequent mouse studies identified a distinct signature of T2D-associated miRNAs, partly conserved in both species. Of those, human-murine miR-182-5 p was the most associated with whole-body glucose homeostasis and hepatic lipid metabolism. Its target gene LRP6 was consistently lower expressed in livers of obese T2D humans and mice as well as under conditions of miR-182-5 p overexpression. Weight loss in obese mice decreased hepatic miR-182-5 p and restored Lrp6 expression and other miR-182-5 p target genes. Hepatic overexpression of miR-182-5 p in mice rapidly decreased LRP6 protein levels and increased liver triglycerides and fasting insulin under obesogenic conditions after only seven days. Conclusions: By mapping the hepatic miRNA-transcriptome of type 2 diabetic obese subjects, validating conserved miRNAs in diet-induced mice, and establishing a novel miRNA prediction tool, we provide a robust and unique resource that will pave the way for future studies in the field. As proof of concept, we revealed that the repression of LRP6 by miR-182-5 p, which promotes lipogenesis and impairs glucose homeostasis, provides a novel mechanistic link between T2D and non-alcoholic fatty liver disease, and demonstrate in vivo that miR-182-5 p can serve as a future drug target for the treatment of obesity-driven hepatic steatosis. Funding: This work was supported by research funding from the Deutsche Forschungsgemeinschaft (KI 1887/2-1, KI 1887/2-2, KI 1887/3-1 and CRC-TR296), the European Research Council (ERC, CoG Yoyo LepReSens no. 101002247; PTP), the Helmholtz Association (Initiative and Networking Fund International Helmholtz Research School for Diabetes; MB) and the German Center for Diabetes Research (DZD Next Grant 82DZD09D1G).

Modulatory Roles of AHR, FFAR2, FXR, and TGR5 Gene Expression in Metabolic-Associated Fatty Liver Disease and COVID-19 Outcomes.

Metabolic-associated fatty liver disease (MAFLD) is a risk factor for severe COVID-19. This study explores the potential influence of gut hormone receptor and immune response gene expression on COVID-19 outcomes in MAFLD patients. METHODS: We investigated gene expression levels of AHR, FFAR2, FXR, and TGR5 in patients with MAFLD and COVID-19 compared to controls. We examined associations between gene expression and clinical outcomes. RESULTS: COVID-19 patients displayed altered AHR expression, potentially impacting immune response and recovery. Downregulated AHR in patients with MAFLD correlated with increased coagulation parameters. Elevated FFAR2 expression in patients with MAFLD was linked to specific immune cell populations and hospital stay duration. A significantly lower FXR expression was observed in both MAFLD and severe COVID-19. CONCLUSION: Our findings suggest potential modulatory roles for AHR, FFAR2, and FXR in COVID-19 and MAFLD.

Genetic Background Strongly Influences the Impact of Carrying the Thr92Ala-DIO2 Polymorphism in the Male Mouse.

About half of the world population carries at least one allele of the Ala92-DIO2, which slows down the activity of the type 2 deiodinase (D2), the enzyme that activates T4 to T3. Carrying the Ala92-DIO2 allele has been associated with increased body mass index and insulin resistance, but this has not been reproduced in all populations. To test if the genetic background affects the impact of this polymorphism, here we studied the genetically distant C57Bl/6J (B6) and FVB/N (FVB) mice carrying the Ala92-Dio2 allele as compared to control mice carrying the Thr92-Dio2 allele. Whereas B6-Ala92-Dio2 and B6-Thr92-Dio2 mice-fed chow or high-fat diet-behaved metabolically similar in studies using indirect calorimetry, glucose- and insulin tolerance tests, and measuring white adipose tissue (WAT) weight and liver steatosis, major differences were observed between FVB-Ala92-Dio2 and FVB-Thr92-Dio2 mice: carrying the Ala92-Dio2 allele (on a chow diet) resulted in hypercholesterolemia, smaller WAT pads, hepatomegaly, steatosis, and transcriptome changes in the interscapular brown adipose tissue (iBAT) typical of ER stress and apoptosis. Acclimatization at thermoneutrality (30 °C) eliminated most of the metabolic phenotype, indicating that impaired adaptive (BAT) thermogenesis can be involved. In conclusion, the metabolic impact of carrying the Ala92-Dio2 allele depends greatly on the genetic background of the mouse, varying from no phenotype in B6 mice to a major phenotype in FVB mice. These results will help the planning of future clinical trials studying the Thr92Ala-DIO2 polymorphism and may explain why some clinical studies performed in different populations across the globe have obtained inconsistent results.

Liver epigenomic signature associated with chronic oxidative stress in a mouse model of glutathione deficiency.

Oxidative stress is intimately involved in the pathogenesis of fatty liver disease (FLD). A major factor contributing to oxidative stress is the depletion of the ubiquitous antioxidant glutathione (GSH). Unexpectedly, chronic GSH deficiency renders glutamate-cysteine ligase modifier subunit (Gclm)-null mice protected from fatty liver injuries. Epigenetic regulation serves as an important cellular mechanism in modulating gene expression and disease outcome in FLD, although it is not well understood how systemic redox imbalance modifies the liver epigenome. In the current study, utilizing the Gclm-null mouse model, we aimed to elucidate redox-associated epigenomic changes and their implications in liver stress response. We performed high-throughput array-based DNA methylation profiling (MeDIP array) in 22,327 gene promoter regions (from -1300 bp to +500 bp of the Transcription Start Sites) in the liver and peripheral blood cells. Results from the MeDIP array demonstrate that, although global methylation enrichment in gene promoters did not change, low GSH resulted in prevalent demethylation at the individual promoter level. Such an effect likely attributed to a declined availability of the methyl donor S-adenosyl methionine (SAM) in Gclm-null liver. Functional enrichment analysis of liver target genes is suggestive of a potential role of epigenetic mechanisms in promoting cellular survival and lipid homeostasis in Gclm-null liver. In comparison with the liver tissue, MeDIP array in peripheral blood cells revealed a panel of 19 gene promoters that are candidate circulating biomarkers for hepatic epigenomic changes associated with chronic GSH deficiency. Collectively, our results provided new insights into the in vivo interplay between liver redox state and DNA methylation status. The current study laid the groundwork for future epigenetic/epigenomic investigations in experimental settings or human populations under conditions of liver oxidative stress induced by environmental or dietary challenges.

Distinct Gut Microbial Signature and Host Genetic Variants in Association with Liver Fibrosis Severity in Patients with MASLD.

Gut microbiota might affect the severity and progression of metabolic dysfunction-associated steatotic liver disease (MASLD). We aimed to characterize gut dysbiosis and clinical parameters regarding fibrosis stages assessed by magnetic resonance elastography. This study included 156 patients with MASLD, stratified into no/mild fibrosis (F0-F1) and moderate/severe fibrosis (F2-F4). Fecal specimens were sequenced targeting the V4 region of the 16S rRNA gene and analyzed using bioinformatics. The genotyping of PNPLA3, TM6SF2, and HSD17B13 was assessed by allelic discrimination assays. Our data showed that gut microbial profiles between groups significantly differed in beta-diversity but not in alpha-diversity indices. Enriched Fusobacterium and Escherichia_Shigella, and depleted Lachnospira were found in the F2-F4 group versus the F0-F1 group. Compared to F0-F1, the F2-F4 group had elevated plasma surrogate markers of gut epithelial permeability and bacterial translocation. The bacterial genera, PNPLA3 polymorphisms, old age, and diabetes were independently associated with advanced fibrosis in multivariable analyses. Using the Random Forest classifier, the gut microbial signature of three genera could differentiate the groups with high diagnostic accuracy (AUC of 0.93). These results indicated that the imbalance of enriched pathogenic genera and decreased beneficial bacteria, in association with several clinical and genetic factors, were potential contributors to the pathogenesis and progression of MASLD.

Interplay of Mediterranean-diet adherence, genetic factors, and metabolic dysfunction-associated steatotic liver disease risk in Korea.

BACKGROUNDS: Metabolic dysfunction-associated steatotic liver disease (MASLD) has gained attention owing to its severe complications. This study aimed to explore the interaction between Mediterranean-diet (MD) adherence, genetic factors, and MASLD risk in a Korean population. METHODS: In total, 33,133 individuals aged 40 years and older from the Korean Genome and Epidemiology Study (KoGES) were analyzed. Participants were assessed for MASLD based on criteria and MD adherence measured by the Korean version of the Mediterranean-Diet Adherence Screener (K-MEDAS). Individuals were categorized into two groups based on their MD adherence: high adherence (K-MEDAS > 6) and low adherence (K-MEDAS < 5). Single nucleotide polymorphism (SNP) genotypes were obtained using the Korea Biobank array. Logistic regression was used to examine the single-marker variants for genetic associations with MASLD prevalence. RESULTS: Individuals were categorized into MASLD (10,018 [30.2%]) and non-MASLD (23,115 [69.8%]) groups. A significant interaction was observed between the rs780094 glucokinase regulatory protein (GCKR) gene and K-MEDAS on MASLD (p < 10 - 2 ). Of individuals with K-MEDAS > 6, those carrying the minor allele (C) of the GCKR gene rs780094 exhibited a lower risk of MASLD compared to those without the allele (odds ratio [OR] = 0.88 [0.85-0.91], p-value = 5.54e-13). CONCLUSION: The study identified a significant interaction involving the rs780094 variant near the GCKR gene, with carriers of the minor allele exhibiting a lower MASLD risk among those adhering well to the MD. Dietary habits influence the MASLD risk associated with the rs780094 allele, emphasizing the need for personalized nutrition recommendations.

Do iron homeostasis biomarkers mediate the associations of liability to type 2 diabetes and glycemic traits in liver steatosis and cirrhosis: a two-step Mendelian randomization study.

BACKGROUND: Previous studies, including Mendelian randomization (MR), have demonstrated type 2 diabetes (T2D) and glycemic traits are associated with increased risk of metabolic dysfunction-associated steatotic liver disease (MASLD). However, few studies have explored the underlying pathway, such as the role of iron homeostasis. METHODS: We used a two-step MR approach to investigate the associations of genetic liability to T2D, glycemic traits, iron biomarkers, and liver diseases. We analyzed summary statistics from various genome-wide association studies of T2D (n = 933,970), glycemic traits (n ≤ 209,605), iron biomarkers (n ≤ 246,139), MASLD (n ≤ 972,707), and related biomarkers (alanine aminotransferase (ALT) and proton density fat fraction (PDFF)). Our primary analysis was based on inverse-variance weighting, followed by several sensitivity analyses. We also conducted mediation analyses and explored the role of liver iron in post hoc analysis. RESULTS: Genetic liability to T2D and elevated fasting insulin (FI) likely increased risk of liver steatosis (ORliability to T2D: 1.14 per doubling in the prevalence, 95% CI: 1.10, 1.19; ORFI: 3.31 per log pmol/l, 95% CI: 1.92, 5.72) and related biomarkers. Liability to T2D also likely increased the risk of developing liver cirrhosis. Genetically elevated ferritin, serum iron, and liver iron were associated with higher risk of liver steatosis (ORferritin: 1.25 per SD, 95% CI 1.07, 1.46; ORliver iron: 1.15 per SD, 95% CI: 1.05, 1.26) and liver cirrhosis (ORserum iron: 1.31, 95% CI: 1.06, 1.63; ORliver iron: 1.34, 95% CI: 1.07, 1.68). Ferritin partially mediated the association between FI and liver steatosis (proportion mediated: 7%, 95% CI: 2-12%). CONCLUSIONS: Our study provides credible evidence on the causal role of T2D and elevated insulin in liver steatosis and cirrhosis risk and indicates ferritin may play a mediating role in this association.

PNPLA3 is a triglyceride lipase that mobilizes polyunsaturated fatty acids to facilitate hepatic secretion of large-sized very low-density lipoprotein.

The I148M variant of PNPLA3 is closely associated with hepatic steatosis. Recent evidence indicates that the I148M mutant functions as an inhibitor of PNPLA2/ATGL-mediated lipolysis, leaving the role of wild-type PNPLA3 undefined. Despite showing a triglyceride hydrolase activity in vitro, PNPLA3 has yet to be established as a lipase in vivo. Here, we show that PNPLA3 preferentially hydrolyzes polyunsaturated triglycerides, mobilizing polyunsaturated fatty acids for phospholipid desaturation and enhancing hepatic secretion of triglyceride-rich lipoproteins. Under lipogenic conditions, mice with liver-specific knockout or acute knockdown of PNPLA3 exhibit aggravated liver steatosis and reduced plasma VLDL-triglyceride levels. Similarly, I148M-knockin mice show decreased hepatic triglyceride secretion during lipogenic stimulation. Our results highlight a specific context whereby the wild-type PNPLA3 facilitates the balance between hepatic triglyceride storage and secretion, and suggest the potential contribution of a loss-of-function by the I148M variant to the development of fatty liver disease in humans.

Hepatocyte-specific loss of DDB1 attenuates hepatic steatosis but aggravates liver inflammation and fibrosis in MASH.

BACKGROUND: MASH is a common clinical disease that can lead to advanced liver conditions, but no approved pharmacotherapies are available due to an incomplete understanding of its pathogenesis. Damaged DNA binding protein 1 (DDB1) participates in lipid metabolism. Nevertheless, the function of DDB1 in MASH is unclear. METHODS: Clinical liver samples were obtained from patients with MASH and control individuals by liver biopsy. Hepatocyte-specific Ddb1-knockout mice and liver Hmgb1 knockdown mice were fed with a methionine-and choline-deficient diet to induce MASH. RESULTS: We found that the expression of DDB1 in the liver was significantly decreased in MASH models. Hepatocyte-specific ablation of DDB1 markedly alleviated methionine-and choline-deficient diet-induced liver steatosis but unexpectedly exacerbated inflammation and fibrosis. Mechanistically, DDB1 deficiency attenuated hepatic steatosis by downregulating the expression of lipid synthesis and uptake genes. We identified high-mobility group box 1 as a key candidate target for DDB1-mediated liver injury. DDB1 deficiency upregulated the expression and extracellular release of high-mobility group box 1, which further increased macrophage infiltration and activated HSCs, ultimately leading to the exacerbation of liver inflammation and fibrosis. CONCLUSIONS: These data demonstrate the independent regulation of hepatic steatosis and injury in MASH. These findings have considerable clinical implications for the development of therapeutic strategies for MASH.

Mechanisms of hepatic steatosis in chickens: integrated analysis of the host genome, molecular phenomics and gut microbiome.

Hepatic steatosis is the initial manifestation of abnormal liver functions and often leads to liver diseases such as nonalcoholic fatty liver disease in humans and fatty liver syndrome in animals. In this study, we conducted a comprehensive analysis of a large chicken population consisting of 705 adult hens by combining host genome resequencing; liver transcriptome, proteome, and metabolome analysis; and microbial 16S ribosomal RNA gene sequencing of each gut segment. The results showed the heritability (h2 = 0.25) and duodenal microbiability (m2 = 0.26) of hepatic steatosis were relatively high, indicating a large effect of host genetics and duodenal microbiota on chicken hepatic steatosis. Individuals with hepatic steatosis had low microbiota diversity and a decreased genetic potential to process triglyceride output from hepatocytes, fatty acid ß-oxidation activity, and resistance to fatty acid peroxidation. Furthermore, we revealed a molecular network linking host genomic variants (GGA6: 5.59-5.69 Mb), hepatic gene/protein expression (PEMT, phosphatidyl-ethanolamine N-methyltransferase), metabolite abundances (folate, S-adenosylmethionine, homocysteine, phosphatidyl-ethanolamine, and phosphatidylcholine), and duodenal microbes (genus Lactobacillus) to hepatic steatosis, which could provide new insights into the regulatory mechanism of fatty liver development.

Increased capacity to maintain glucose homeostasis in a transgenic mouse expressing human but not mouse growth hormone with developing high-fat diet-related insulin resistance, hepatic steatosis and adipose dysfunction.

The objective was to assess the potential differential effects of human versus mouse growth hormone in vivo, given that human unlike mouse growth hormone can bind prolactin as well as the growth hormone receptor. To this end, a transgenic CD-1 mouse expressing human but not mouse growth hormone was generated, and the phenotypes of male mice fed with a regular chow or high-fat diet were assessed. Pancreas and epididymal white adipose tissue gene expression and/or related function were targeted as the pancreas responds to both prolactin and growth hormone receptor signaling, and catabolic effects like lipolytic activity are more directly attributable to growth hormone and growth hormone receptor signaling. The resulting human growth hormone-expressing mice are smaller than wild-type CD-1 mice, despite higher body fat and larger adipocytes, but both mouse types grow at the same rate with similar bone densities. Unlike wild-type mice, there was no significant delay in glucose clearance in human growth hormone-expressing mice when assessed at 8 versus 24 weeks on a high-fat diet. However, both mouse types showed signs of hepatic steatosis that correlated with elevated prolactin but not growth hormone RNA levels. The larger adipocytes in human growth hormone-expressing mice were associated with modified leptin (higher) and adiponectin (lower) RNA levels. Thus, while limited to observations in the male, the human growth hormone-expressing mice exhibit signs of growth hormone insufficiency and adipocyte dysfunction as well as an initial resistance to the negative effects of high-fat diet on glucose clearance.

Liver matrin-3 protects mice against hepatic steatosis and stress response via constitutive androstane receptor.

OBJECTIVE: The prevalence of metabolic dysfunction-associated steatotic liver disease (MASLD) continues to rise with the increasing obesity epidemic. Rezdiffra as an activator of a thyroid hormone receptor-beta is the only Food and Drug Administration approved therapy. As such, there is a critical need to improve our understanding of gene expression regulation and signaling transduction in MASLD to develop new therapies. Matrin-3 is a DNA- and RNA-binding protein involved in the pathogenesis of human diseases. Here we examined its previously uncharacterized role in limiting hepatic steatosis and stress response via the constitutive androstane receptor (CAR). METHODS: Matrin-3 floxed and liver-specific knockout mice were fed either a chow diet or 60 kcal% high-fat diet (HFD) for up to 16 weeks. The mice were euthanized for different analysis including liver histology, lipid levels, and gene expression. Bulk RNA-seq, bulk ATAC-seq, and single-nucleus Multiome were used to examine changes of transcriptome and chromatin accessibility in the liver. Integrative bioinformatics analysis of our data and publicly available datasets and different biochemical assays were performed to identify underlying the molecular mechanisms mediating matrin-3's effects. Liver-tropic adeno-associated virus was used to restore the expression of CAR for lipid, acute phase genes, and histological analysis. RESULTS: Matrin-3 expression is induced in the steatotic livers of mice. Liver-specific matrin-3 deletion exacerbated HFD-induced steatosis, acute phase response, and inflammation in the liver of female mice. The transcriptome and chromatin accessibility were re-programmed in the liver of these mice with signatures indicating that CAR signaling is dysregulated. Mechanistically, matrin-3 interacts with CAR mRNA, and matrin-3 deficiency promotes CAR mRNA degradation. Consequently, matrin-3 deletion impaired CAR signaling by reducing CAR expression. Matrin-3 levels positively correlate with CAR expression in human livers. Ces2a and Il1r1 were identified as new target genes of CAR. Interestingly, we found that CAR discords with the expression of its target genes including Cyp2b10 and Ces2a in response to HFD, indicating CAR signaling is dysregulated by HFD despite increased CAR expression. Dysregulated CAR signaling upon matrin-3 deficiency reduced Ces2a and de-repressed Il1r1 expression. CAR restoration partially abrogated the dysregulated gene expression, exacerbated hepatic steatosis, acute phase response, and inflammation in liver-specific matrin-3 knockout mice fed a HFD. CONCLUSIONS: Our findings demonstrate that matrin-3 is a key upstream regulator maintaining CAR signaling upon metabolic stress, and the matrin-3-CAR axis limits hepatic steatosis and stress response signaling that may give insights for therapeutic intervention.

Suppression of STK39 weakens the MASLD/MASH process by protecting the intestinal barrier.

STK39 is reportedly a critical negative regulator of intestinal barrier. Pharmacological targeting of STK39 is expected to protect the intestinal barrier and thereby weaken metabolic dysfunction-associated steatohepatitis (MASH); Proximal colon biopsy tissues from patients with metabolic dysfunction-associated steatotic liver disease (MASLD) and those without MASLD were analyzed for STK39 expression. Wildtype (WT) mice and systemic STK39 gene knockout (STK39-/-) male mice were fed a normal diet or a high-fat methionine-choline deficient diet (HFMCD) for 8 weeks. The MASH mice were grouped and treated with ZT-1a (a STK39 inhibitor) or vehicle intraperitoneal injection during the procedure of HFMCD induction. Liver and intestinal tissues were collected for further examination; Colon tissues from patients with MASLD exhibited higher levels of STK39 than those from subjects without MASLD. Knockout of STK39 diminished CD68+ Kupffer cells and α-SMA+ hepatic stellate cells infiltration in mouse MASH model. Treatment with ZT-1a also prevented severe steatohepatitis in a mouse MASH model, including milder histological and pathological manifestations (lobular inflammation and fibrosis) in the liver. Interestingly, Inhibition of STK39 had minimal effects on hepatic lipid metabolism. The reduced liver injury observed in mice with STK39 inhibition was linked to significant decreases in mucosal inflammation, tight junction disruption and intestinal epithelial permeability to bacterial endotoxins; Collectively, we have revealed that inhibiting STK39 prevents the progression of MASH by protecting the intestinal epithelial barrier.

Hepatocyte-specific Selenoi deficiency predisposes mice to hepatic steatosis and obesity.

Selenoprotein I (Selenoi) is highly expressed in liver and plays a key role in lipid metabolism as a phosphatidylethanolamine (PE) synthase. However, the precise function of Selenoi in the liver remains elusive. In the study, we generated hepatocyte-specific Selenoi conditional knockout (cKO) mice on a high-fat diet to identify the physiological function of Selenoi. The cKO group exhibited a significant increase in body weight, with a 15.6% and 13.7% increase in fat accumulation in white adipose tissue (WAT) and the liver, respectively. Downregulation of the lipolysis-related protein (p-Hsl) and upregulation of the adipogenesis-related protein (Fasn) were observed in the liver of cKO mice. The cKO group also showed decreased oxygen consumption (VO2), carbon dioxide production (VCO2), and energy expenditure (p < .05). Moreover, various metabolites of the steroid hormone synthesis pathway were affected in the liver of cKO mice. A potential cascade of Selenoi-phosphatidylethanolamine-steroid hormone synthesis might serve as a core mechanism that links hepatocyte-specific Selenoi cKO to biochemical and molecular reactions. In conclusion, we revealed that Selenoi inhibits body fat accumulation and hepatic steatosis and elevates energy consumption; this protein could also be considered a therapeutic target for such related diseases.

Phospholipid metabolism-related genotypes of PLA2R1 and CERS4 contribute to nonobese MASLD.

BACKGROUND: Abnormal phospholipid metabolism is linked to metabolic dysfunction-associated steatotic liver disease (MASLD) development and progression. We aimed to clarify whether genetic variants of phospholipid metabolism modify these relationships. METHODS: This case-control study consecutively recruited 600 patients who underwent MRI-based proton density fat fraction examination (240 participants with serum metabonomics analysis, 128 biopsy-proven cases) as 3 groups: healthy control, nonobese MASLD, and obese MASLD, (n = 200 cases each). Ten variants of phospholipid metabolism-related genes [phospholipase A2 Group VII rs1805018, rs76863441, rs1421378, and rs1051931; phospholipase A2 receptor 1 (PLA2R1) rs35771982, rs3828323, and rs3749117; paraoxonase-1 rs662 and rs854560; and ceramide synthase 4 (CERS4) rs17160348)] were genotyped using SNaPshot. RESULTS: The T-allele of CERS4 rs17160348 was associated with a higher risk of both obese and nonobese MASLD (OR: 1.95, 95% CI: 1.20-3.15; OR: 1.76, 95% CI: 1.08-2.86, respectively). PLA2R1 rs35771982-allele is a risk factor for nonobese MASLD (OR: 1.66, 95% CI: 1.11-1.24), moderate-to-severe steatosis (OR: 3.24, 95% CI: 1.96-6.22), and steatohepatitis (OR: 2.61, 95% CI: 1.15-3.87), while the paraoxonase-1 rs854560 T-allele (OR: 0.50, 95% CI: 0.26-0.97) and PLA2R1 rs3749117 C-allele (OR: 1.70, 95% CI: 1.14-2.52) are closely related to obese MASLD. After adjusting for sphingomyelin level, the effect of the PLA2R1 rs35771982CC allele on MASLD was attenuated. Furthermore, similar effects on the association between the CERS4 rs17160348 C allele and MASLD were observed for phosphatidylcholine, phosphatidic acid, sphingomyelin, and phosphatidylinositol. CONCLUSIONS: The mutations in PLA2R1 rs35771982 and CERS4 rs17160348 presented detrimental impact on the risk of occurrence and disease severity in nonobese MASLD through altered phospholipid metabolism.