Non-alcoholic Fatty Liver Progression Research Articles

The roles of type 2 diabetes and obesity in disease activity and progression of non-alcoholic fatty liver disease/non-alcoholic steatohepatitis

Objective We examined the roles of type 2 diabetes (T2D) and obesity in disease activity and fibrosis progression/regression in patients with non-alcoholic fatty liver disease/non-alcoholic steatohepatitis (NAFLD/NASH). Methods This multi-center, retrospective study included patients with suspected or histologically proven NAFLD/NASH from the NASH Clinical Research Network. Outcomes included disease activity and rate of fibrosis, assessed using liver-biopsy driven measures (NAFLD activity score [NAS] and fibrosis score [FS]). Logistic regression and doubly robu estimation of causal effects tested relationships among T2D, obesity, and NAFLD/NASH. Results The analytical sample included 870 adult patients with baseline biopsy data and 157 patients with multiple biopsy data. Patients with NAFLD/NASH and T2D had significantly higher baseline average NAS (4.52 vs. 4.13; p = 0.009) and FS (2.17 vs. 1.56; p < 0.0001); those with T2D had a significantly greater reduction in average NAS over time (−0.77/year vs. −0.17/year; p = 0.0008). Change in FS over time did not differ significantly by T2D status (−0.23/year vs. −0.04/year; p = 0.34). Baseline NAS, baseline FS, and change in average NAS over time did not differ significantly by obesity status (4.17 vs. 4.47; p = 0.16; 1.73 vs.1.92; p = 0.31; −0.40/year vs. −0.59/year; p = 0.62, respectively). Patients with obesity had a slight increase in FS but those without obesity had a reduction in average FS over time (0.07/year vs. −0.27/year; p = 0.008). Conclusions Patients with NAFLD/NASH and T2D had greater baseline disease activity versus those without T2D, but there was greater regression of disease activity over time among those with T2D. Patients with NAFLD/NASH and obesity had worsening of fibrosis versus those without obesity. NCT00063622

B-003 Decrease in Serum apoE-rich HDL-cholesterol May Be Associated With Cholesterol Accumulation in the Liver of Rats Fed A High-cholesterol Diet

Abstract Background Free cholesterol accumulation in the liver has been suggested to be one of risk factors for the progression of non-alcoholic fatty liver to non-alcoholic steatohepatitis. Therefore, a non-invasive approach to assess hepatic cholesterol accumulation in the early stage without elevation of liver enzymes (AST and ALT) may be useful in clinical setting. In this study, we show that serum apoE-rich HDL-cholesterol level is a possible indicator for predicting hepatic cholesterol accumulation in rats with diet-induced simple liver steatosis. Methods Male Sprague-Dawley rats (8 weeks) were purchased from Charles River Laboratories. Four different diets were obtained from Oriental Yeast; control diet (MF), high-fat diet (HF; 68% MF, 30% palm oil, and 2% cholic acid), high-cholesterol diet (HC; 95.5% MF, 2.5% cholesterol, and 2% cholic acid), and high-fat/high-cholesterol diet (HFC; 81.75% MF, 15% palm oil, 1.25% cholesterol, and 2% cholic acid). The rats were randomly divided into 4 groups and fed as follows. The control and HFC groups received the MF and HFC diets for 8 weeks, respectively. The HF/HC group received the HF diet for the first 4 weeks and the HC diet for the next 4 weeks. The HC/HF group received the HC diet first, then the HF diet. Blood and liver tissue specimens were collected under anesthesia. Total HDL fractions were isolated from whole sera by the polyethylene glycol precipitation method, and applied into a cation-exchange column (HiTrap SPHP, GE healthcare) for determining apoE-rich HDL-cholesterol levels. Correlations between variables were assessed with Spearman’s correlation coefficient (rs). Results Total cholesterol content of the liver was significantly higher in the HFC (11.1-fold, P &amp;lt; 0.001), HF/HC (9.1-fold, P &amp;lt; 0.001), and HC/HF (5.8-fold, P &amp;lt; 0.001) groups than the control group. Hepatic free cholesterol was also higher in the HFC (5.5-fold, P &amp;lt; 0.001), HF/HC (3.9-fold, P &amp;lt; 0.001), and HC/HF (3.3-fold, P &amp;lt; 0.001) groups. Hepatic triglycerides were slightly increased in the HC/HF (1.6-fold, P &amp;lt; 0.01) and HFC (1.5-fold, P &amp;lt; 0.05) groups. Serum apoE-rich HDL-cholesterol was significantly decreased in the HF/HC (0.21-fold, P &amp;lt; 0.001), HC/HF (0.25-fold, P &amp;lt; 0.001) and HFC (0.42-fold, P &amp;lt; 0.01) groups. The HFC group had significantly higher serum ALT (3.0-fold, P &amp;lt; 0.05) and AST (2.3-fold, P &amp;lt; 0.01) levels, compared with the control groups. There was a significant increase in hepatic mRNA expression of TNF-α (24.8-fold, P &amp;lt; 0.01) and TGF-β (3.8-fold, P &amp;lt; 0.001) only in the HFC group, indicating liver inflammation and stellate cell activation. To exclude the effects of hepatocyte injury on apoE-rich HDL metabolism, rats with ALT less than 100 U/mL (n = 16) were included in the following analysis. ApoE-rich HDL-cholesterol levels were significantly and inversely correlated with hepatic total cholesterol (rs = −0.535, P &amp;lt; 0.05) and free cholesterol (rs = −0.532, P &amp;lt; 0.05) levels but not with hepatic triglycerides. Furthermore, the ratio of ALT to apoE-rich HDL-cholesterol was more highly correlated with hepatic total cholesterol (rs = 0.626, P &amp;lt; 0.01) and free cholesterol (rs = 0.632, P &amp;lt; 0.01). There was no such correlation for apoE-poor HDL-cholesterol. Conclusion: The present observations suggest that serum apoE-rich HDL-cholesterol levels may be a useful indicator for cholesterol accumulation in the liver of cholesterol-fed rats.

A wash-free fluorescent probe with a large Stokes shift for the identification of NAFL through tracing the change of lipid droplets.

As one of the important organelles in cells, lipid droplets (LDs) are involved in various physiological processes, especially affecting the occurrence and progression of non-alcoholic fatty liver (NAFL). Therefore, it is of great significance to develop LD-specific probes with excellent biocompatibility, deep penetration and bright fluorescence. Herein, a fluorescent probe LD-HWZ was designed and synthesized based on triphenylamine and the dicyanoisophorone group. It is found that probe LD-HWZ has a large Stokes shift (Δλ = 160 nm in DMSO) and exhibits bright fluorescence in a lipid environment. In addition, biological experiments showed that LD-HWZ can localize in lipid droplets, which can be used to detect the dynamic changes of LDs. Importantly, LD-HWZ has been successfully used to discriminate NAFL tissues from normal livers. The excellent properties of probe LD-HWZ in this work are expected to shed new light on the design of lipid droplet probes for the study of fatty liver diagnosis.

Endogenous Liver Protections Against Lipotoxicity and Oxidative Stress to Avoid the Progression of Non-alcoholic Fatty Liver to more Serious Disease.

Non-alcoholic fatty liver disease (NAFLD) is a metabolic disorder characterized by an ectopic accumulation of lipids in at least 5% of hepatocytes. The first phase of the disease, called hepatic steatosis, progresses over time to chronic conditions, such as steatohepatitis, cirrhosis, and finally, hepatic insufficiency and cancer. The accumulation of free fatty acids in hepatocytes, particularly saturated fatty acids, is a key process in the development and progression of NAFLD. Furthermore, the accumulation of oxidative stress markers in NAFLD is closely linked to lipotoxicity due to impaired lipid metabolism and increased generation of reactive oxygen species (ROS). However, endogenous mechanisms are activated early in the liver to protect against lipotoxicity and oxidative stress, thus preventing liver mass loss and disease progression. Thus, in order to develop appropriate therapies, the purpose of this review is to discuss recent data from the literature regarding the importance of intrinsic mechanisms deployed by the liver in protecting itself against the adverse effects related to chronic lipid accumulation and ROS generation.

Machine learning to predict progression of non-alcoholic fatty liver to non-alcoholic steatohepatitis or fibrosis.

BackgroundNon‐alcoholic fatty liver (NAFL) can progress to the severe subtype non‐alcoholic steatohepatitis (NASH) and/or fibrosis, which are associated with increased morbidity, mortality, and healthcare costs. Current machine learning studies detect NASH; however, this study is unique in predicting the progression of NAFL patients to NASH or fibrosis.AimTo utilize clinical information from NAFL‐diagnosed patients to predict the likelihood of progression to NASH or fibrosis.MethodsData were collected from electronic health records of patients receiving a first‐time NAFL diagnosis. A gradient boosted machine learning algorithm (XGBoost) as well as logistic regression (LR) and multi‐layer perceptron (MLP) models were developed. A five‐fold cross‐validation grid search was utilized for hyperparameter optimization of variables, including maximum tree depth, learning rate, and number of estimators. Predictions of patients likely to progress to NASH or fibrosis within 4 years of initial NAFL diagnosis were made using demographic features, vital signs, and laboratory measurements.ResultsThe XGBoost algorithm achieved area under the receiver operating characteristic (AUROC) values of 0.79 for prediction of progression to NASH and 0.87 for fibrosis on both hold‐out and external validation test sets. The XGBoost algorithm outperformed the LR and MLP models for both NASH and fibrosis prediction on all metrics.ConclusionIt is possible to accurately identify newly diagnosed NAFL patients at high risk of progression to NASH or fibrosis. Early identification of these patients may allow for increased clinical monitoring, more aggressive preventative measures to slow the progression of NAFL and fibrosis, and efficient clinical trial enrollment.

Impaired Hepatic Mitochondrial Capacity in Nonalcoholic Steatohepatitis Associated With Type 2 Diabetes.

Individuals with type 2 diabetes are at higher risk of progression of nonalcoholic fatty liver (steatosis) to steatohepatitis (NASH), fibrosis, and cirrhosis. The hepatic metabolism of obese individuals adapts by upregulation of mitochondrial capacity, which may be lost during the progression of steatosis. However, the role of type 2 diabetes with regard to hepatic mitochondrial function in NASH remains unclear. We therefore examined obese individuals with histologically proven NASH without (OBE) (n = 30; BMI 52 ± 9 kg/m2) or with type 2 diabetes (T2D) (n = 15; 51 ± 7 kg/m2) as well as healthy individuals without liver disease (CON) (n = 14; 25 ± 2 kg/m2). Insulin sensitivity was measured by hyperinsulinemic-euglycemic clamps with d-[6,6-2H2]glucose. Liver biopsies were used for assessing mitochondrial capacity by high-resolution respirometry and protein expression. T2D and OBE had comparable hepatic fat content, lobular inflammation, and fibrosis. Oxidative capacity in liver tissue normalized for citrate synthase activity was 59% greater in OBE than in CON, whereas T2D presented with 33% lower complex II-linked oxidative capacity than OBE and higher H2O2 production than CON. Interestingly, those with NASH and hepatic fibrosis score ≥1 had lower oxidative capacity and antioxidant defense than those without fibrosis. Loss of hepatic mitochondrial adaptation characterizes NASH and type 2 diabetes or hepatic fibrosis and may thereby favor accelerated disease progression.

The methyltransferase METTL3 negatively regulates nonalcoholic steatohepatitis (NASH) progression

Nonalcoholic steatohepatitis (NASH) is a key step in the progression of nonalcoholic fatty liver (NAFL) to cirrhosis. However, the molecular mechanisms of the NAFL-to-NASH transition are largely unknown. Here, we identify methyltransferase like 3 (METTL3) as a key negative regulator of NASH pathogenesis. Hepatocyte-specific deletion of Mettl3 drives NAFL-to-NASH progression by increasing CD36-mediated hepatic free fatty acid uptake and CCL2-induced inflammation, which is due to increased chromatin accessibility in the promoter region of Cd36 and Ccl2. Antibody blockade of CD36 and CCL2 ameliorates NASH progression in hepatic Mettl3 knockout mice. Hepatic overexpression of Mettl3 protects against NASH progression by inhibiting the expression of CD36 and CCL2. Mechanistically, METTL3 directly binds to the promoters of the Cd36 and Ccl2 genes and recruits HDAC1/2 to induce deacetylation of H3K9 and H3K27 in their promoters, thus suppressing Cd36 and Ccl2 transcription. Furthermore, METTL3 is translocated from the nucleus to the cytosol in NASH, which is associated with CDK9-mediated phosphorylation of METTL3. Our data reveal a mechanism by which METTL3 negatively regulates hepatic Cd36 and Ccl2 gene transcription via a histone modification pathway for protection against NASH progression.

Highly Selective Fluorescent Probe Design for Visualizing Hepatic Hydrogen Sulfide in the Pathological Progression of Nonalcoholic Fatty Liver.

Hydrogen sulfide (H2S), emerging as an important gaseous signal, has attracted more and more attention for its key role in chronic fatty liver diseases. However, lacking tools for H2S-specific in situ detection, the changes of endogenous hepatic H2S levels in the pathological progression of chronic liver diseases are still unclear. To this end, we adopted a strategy of combining molecular probe design and nanofunctionalization to develop a highly selective near-infrared (NIR) fluorescent probe, which allows in vivo real-time monitoring of hepatic H2S levels in the process of nonalcoholic fatty liver disease (NAFLD). As a proof of strategy demonstration, we first designed NIR molecular probes for H2S sensing through chemical design and probe screening and then loaded molecular probes into mesoporous silicon nanomaterials (MSNs) with surface encapsulation using poly(ethylene glycol) to construct a highly selective probe MSN@CSN@PEG, with significantly improved selectivity and photostability. Moreover, MSN@CSN@PEG exhibited high selectivity and sensitivity for endogenous H2S in cells and tumors in vivo, eliminating the interference of a high concentration of biothiols and sulfhydryl proteins. Furthermore, the probe was applied to in situ intravital imaging and systematic assessment of hepatic H2S levels in different stages of NAFLD for the first time, which may offer a promising tool for the future study of fatty liver diseases and other chronic liver diseases.

Identification of Autophagy-Related Genes in the Progression from Non-Alcoholic Fatty Liver to Non-Alcoholic Steatohepatitis.

BackgroundNon-alcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver disease worldwide. Autophagy plays a vital role in NAFLD development and progression. We aimed to establish a novel autophagy-related gene (ARG) signature as a therapeutic target in NAFLD patients based on high-throughput sequencing data.MethodsARGs obtained from the HAMdb and high-sequencing data obtained from the Gene Expression Omnibus (GEO) database were analyzed to identify differentially expressed ARGs (DEARGs) between normal and NASH tissues. Then, gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were performed to explore potential biological and pathological functions of DEARGs. The protein–protein interaction (PPI) network of the DEARGs was established through the STRING website, and visualized by Cytoscape. In addition, hub genes were validated by an independent dataset GSE89632. Finally, we performed Gene Set Variation Analysis (GSVA) pathway-related analysis to identify the pivotal signaling pathways and genes for the progression of non-alcoholic fatty liver (NAFL) to non-alcoholic steatohepatitis (NASH).ResultsA total of 76 DEARGs were identified in the GSE126848 dataset, of which 45 genes were upregulated and 31 genes were downregulated. GO analysis showed that the biological functions of DEARGs focused primarily on autophagy, cellular response to external stimulus, fibroblast proliferation, late endosome, and ubiquitin protein ligase binding. KEGG pathway analysis showed that these DEARGs were mainly involved in the apoptosis, PI3K-Akt signaling pathway, and estrogen signaling pathway. Among DEGs, 9 most closely related genes were identified from the PPI network. Furthermore, NOS3, IGF1, VAMP8, FOS, and HMOX1 were verified in the GSE89632 dataset. At last, the MAPK signal pathway was identified as important pathway, and JUN was identified as a key gene involved in the progression from NAFL to NASH.ConclusionThis study may provide credible molecular biomarkers in terms of screening and diagnosis for NAFLD. Meanwhile, it also serves as a basis for exploring the molecular mechanisms underlying the progression of NAFL to NASH.

Intervention by picroside II on FFAs induced lipid accumulation and lipotoxicity in HepG2 cells

BackgroundAccumulation of free fatty acids (FFAs) in hepatocytes is a hallmark of liver dysfunction and non-alcoholic fatty liver disease (NAFLD). Excessive deposition of FFAs alters lipid metabolism pathways increasing the oxidative stress and mitochondrial dysfunction. Attenuating hepatic lipid accumulation, oxidative stress, and improving mitochondrial function could provide potential targets in preventing progression of non-alcoholic fatty liver (NAFL) to non-alcoholic steatohepatitis (NASH). Earlier studies with Picrorhiza kurroa extract have shown reduction in hepatic damage and fatty acid infiltration in several experimental models and also clinically in viral hepatitis. Thus, the effect of P. kurroa's phytoactive, picroside II, needed mechanistic investigation in appropriate in vitro liver cell model. Objective(s)To study the effect of picroside II on FFAs accumulation, oxidative stress and mitochondrial function with silibinin as a positive control in in vitro NAFLD model. Materials and methodsHepG2 cells were incubated with FFAs-1000μM in presence and absence of Picroside II-10 μM for 20 hours. ResultsHepG2 cells incubated with FFAs-1000μM lead to increased lipid accumulation. Picroside II-10μM attenuated FFAs-induced lipid accumulation (33%), loss of mitochondrial membrane potential (ΔΨm), ATP depletion, and production of reactive oxygen species (ROS). A concomitant increase in cytochrome C at transcription and protein levels was observed. An increase in expression of MnSOD, catalase, and higher levels of tGSH and GSH:GSSG ratios underlie the ROS salvaging activity of picroside II. ConclusionPicroside II significantly attenuated FFAs-induced-lipotoxicity. The reduction in ROS, increased antioxidant enzymes, and improvement in mitochondrial function underlie the mechanisms of action of picroside II. These findings suggest a need to develop an investigational drug profile of picroside II for NAFLD as a therapeutic strategy. This could be evaluated through the fast-track path of reverse pharmacology.

The Alterations of Mitochondrial Function during NAFLD Progression-An Independent Effect of Mitochondrial ROS Production.

The progression of non-alcoholic fatty liver (NAFL) into non-alcoholic steatohepatitis implicates multiple mechanisms, chief of which is mitochondrial dysfunction. However, the sequence of events underlying mitochondrial failure are still poorly clarified. In this work, male C57BL/6J mice were fed with a high-fat plus high-sucrose diet for 16, 20, 22, and 24 weeks to induce NAFL. Up to the 20th week, an early mitochondrial remodeling with increased OXPHOS subunits levels and higher mitochondrial respiration occurred. Interestingly, a progressive loss of mitochondrial respiration along “Western diet” feeding was identified, accompanied by higher susceptibility to mitochondrial permeability transition pore opening. Importantly, our findings prove that mitochondrial alterations and subsequent impairment are independent of an excessive mitochondrial reactive oxygen species (ROS) generation, which was found to be progressively diminished along with disease progression. Instead, increased peroxisomal abundance and peroxisomal fatty acid oxidation-related pathway suggest that peroxisomes may contribute to hepatic ROS generation and oxidative damage, which may accelerate hepatic injury and disease progression. We show here for the first time the sequential events of mitochondrial alterations involved in non-alcoholic fatty liver disease (NAFLD) progression and demonstrate that mitochondrial ROS are not one of the first hits that cause NAFLD progression.

NLRP3 deficiency did not attenuate NASH development under high fat calorie diet plus high fructose and glucose in drinking water

NOD-like receptor protein 3 (NLRP3) promotes the inflammatory response during progression of nonalcoholic fatty liver (NAFL) to nonalcoholic steatohepatitis (NASH). This study aimed to further delineate the role of NLRP3 in NASH development by abolishing its expression in mice. A high-fat and calorie diet plus high fructose and glucose in drinking water (HFCD-HF/G) was used to establish NASH in both wild-type (WT) and NLRP3 knock-out (KO) mice. Hepatocellular injury, hepatic steatosis and fibrosis, as well as inflammatory response and insulin resistance in the liver and epidydimal white adipose tissue (eWAT) were determined. Elevated body weight, liver weight and serum alanine transaminase level, increased hepatic triglyceride accumulation and collagen deposition, and worsened systemic insulin resistance were observed in Nlrp3−/− mice compared to WT mice under HFCD-HF/G feeding. Upregulated hepatic transcription of tumor necrosis factor-α (TNF-α) and monocyte chemotactic protein-1 (MCP-1), and enhanced infiltration of inducible nitric oxide synthase-positive (iNOS+) M1 macrophages were also documented in HFCD-HF/G-fed Nlrp3−/− mice in comparison to HFCD-HF/G-fed WT mice. Moreover, transcription of TNF-α and MCP-1 and infiltration of iNOS+ M1 macrophages were increased in the liver of Nlrp3−/− mice under control diet. NLRP3 deficiency did not attenuate, but instead aggravated NASH development under HFCD-HF/G feeding. The worsened extent of NASH might be attributed to enhanced hepatic MCP-1 expression and M1 macrophage infiltration in Nlrp3−/− mice. Our study points to additional caution when NLRP3 blockade is considered as a therapeutic strategy in the treatment of human NASH.

A network pharmacology approach: Inhibition of the NF-κB signaling pathway contributes to the NASH preventative effect of an Oroxylum indicum seed extract in oleic acid-stimulated HepG2 cells and high-fat diet-fed rats

BackgroundThe incidence of nonalcoholic fatty liver disease (NAFLD), especially nonalcoholic steatohepatitis (NASH), has significantly increased in recent years and has become an important public health issue. However, no U.S. Food and Drug Administration (FDA)-approved first-line drug is currently available for the treatment of NAFLD and NASH; therefore, research on new drugs is currently a hot topic. Oroxylum indicum (Linn.) Kurz is extensively distributed in South China and South Asia and has many biological activities. However, its effects on NAFLD or even NASH and the corresponding mechanisms are still not clear. PurposeTo investigate the effect and mechanism of O. indicum seed extract (OISE) on preventing anti-inflammatory action in the progression from simple nonalcoholic fatty liver (NAFL) to NASH. MethodsA network pharmacology method to construct ingredient-target networks and the protein-protein interaction (PPI) network of OISE in NASH were constructed for topological analyses and hub-target screening. Enrichment analyses were performed to identify the critical biological processes and signaling pathways. Simultaneously, in vitro and in vivo experiments investigated the effect and mechanism of OISE, baicalein, and chrysin on inflammation by biochemical indicator detection, luciferase reporters, pathological staining, and immunoblotting in oleic acid-stimulated HepG2 cells or in high-fat diet-fed rats. ResultsThe network pharmacology showed that OISE prevented the development and progression of NAFL into NASH through various pathways and targets and that the nuclear factor NF-κB (NF-κB) pathway regulated by baicalein and chrysin played an important role in the treatment of NASH. In in vitro experiments, we further showed that OISE and its ingredients, namely, baicalein and chrysin, all improved the inflammatory status in oleic acid-stimulated HepG2 cells, inhibited the nuclear transcriptional activities of NF-κB, increased the IκB level, and decreased the phosphorylation level of NF-κB. Furthermore, in a high-fat diet-induced NASH model in rats, we also showed that OISE prevented the development and progression of NASH by inhibiting the nuclear transcriptional activity of NF-κB. ConclusionOISE suppressed inflammatory responses and prevented the development and progression of NAFL into NASH through inhibition of the nuclear transcriptional activity of NF-κB. OISE may be used to treat NAFLD through many functions, including an increase in insulin sensitivity, a decrease in lipid accumulation in the liver, suppression of inflammation, and clearance of free radicals.

MicroRNAs involved in the progression of non-alcoholic fatty liver

MicroRNAs involved in the progression of non-alcoholic fatty liver

Helicobacter pylori and non-alcoholic fatty liver disease.

Helicobacter pylori is the most prevalent infection worldwide, while non-alcoholic fatty liver disease emerged as the most frequent liver disease. The common occurrence can be either by chance or due to certain pathogenetic factors. Epidemiologic studies revealed that the risk of non-alcoholic liver disease is increased in patients infected with Helicobacter pylori. DNA fragments of Helicobacter pylori were rarely identified in human samples of liver carcinoma and fatty liver. Helicobacter pylori could influence the development of non-alcoholic fatty liver either by hormonal (ghrelin? gastrin? insulin?), or by effect of pro-inflammatory cytokines (interleukin 1 and 8, tumor necrosis factor ɑ, interferon ɣ) and by changes of gut microbiome as well. Probiotic supplementation could improve some clinical parameters of non-alcoholic fatty liver disease and eradication rates of Helicobacter pylori. Regimens used for eradication can be safely administered, although non-alcoholic fatty liver increases the risk of drug-induced liver damage. Controlled studies of the effect of eradication on the development and progression of non-alcoholic fatty liver are warranted.

MRI of atherosclerosis and fatty liver disease in cholesterol fed rabbits

BackgroundThe globally rising obesity epidemic is associated with a broad spectrum of diseases including atherosclerosis and non-alcoholic fatty liver (NAFL) disease. In the past, research focused on the vasculature or liver, but chronic systemic effects and inter-organ communication may promote the development of NAFL. Here, we investigated the impact of confined vascular endothelial injury, which produces highly inflamed aortic plaques that are susceptible to rupture, on the progression of NAFL in cholesterol fed rabbits.MethodsAortic atherosclerotic inflammation (plaque Gd-enhancement), plaque size (vessel wall area), and composition, were measured with in vivo magnetic resonance imaging (MRI) in rabbits fed normal chow or a 1% cholesterol-enriched atherogenic diet. Liver fat was quantified with magnetic resonance spectroscopy (MRS) over 3 months. Blood biomarkers were monitored in the animals, with follow-up by histology.ResultsCholesterol-fed rabbits with and without injury developed hypercholesterolemia, NAFL, and atherosclerotic plaques in the aorta. Compared with rabbits fed cholesterol diet alone, rabbits with injury and cholesterol diets exhibited larger, and more highly inflamed plaques by MRI (P < 0.05) and aggravated liver steatosis by MRS (P < 0.05). Moreover, after sacrifice, damaged (ballooning) hepatocytes and extensive liver fibrosis were observed by histology. Elevated plasma gamma-glutamyl transferase (GGT; P = 0.014) and the ratio of liver enzymes aspartate and alanine aminotransferases (AST/ALT; P = 0.033) indicated the progression of steatosis to non-alcoholic steatohepatitis (NASH).ConclusionsLocalized regions of highly inflamed aortic atherosclerotic plaques in cholesterol-fed rabbits may contribute to progression of fatty liver disease to NASH with fibrosis.

Effect of lipid-induced macrophage M1/M2 polarization on lipid metabolism in hepatocytes

Objective: This study aims to explore the effect of lipid-induced macrophage M1/M2 polarization on lipid metabolism in hepatocytes. Methods: RAW264.7 macrophages were incubated with different kinds of fatty acids including saturated fatty acids-palmitic acid (PA), monounsaturated fatty acids-oleic acid (OA) and polyunsaturated fatty acids-docosahexaenoic acid (DHA), and cell culture supernatants were collected to prepare conditioned medium (CM). Hepatocytes were isolated by in situ perfusion of the liver with collagenase in mice, and a macrophage-hepatocyte CM co-culture system was established. Macrophage M1/M2 phenotype markers were detected by Real-time PCR. Lipid synthesis and decomposition related mRNA and protein expressions in hepatocytes were detected by Real-time PCR and Western Blot. Lipid depositions in hepatocytes were detected by oil red O staining. An analysis of variance was used for comparison of means between multiple groups. Results: Compared with control groups, PA polarized macrophages to a M1 phenotype (expression of TNF-α and IL-6 significantly increased, F≥22.68, P < 0.01), OA polarized macrophages to a M1/M2 mixed phenotype (expression of IL-6, Mrc2 and IL-10 increased F≥4.94, P < 0.05) and DHA polarized macrophages to a M2 phenotype (expression of Mrc2 and IL-10 significantly increased, F≥4.94, P < 0.01). CM-PA significantly increased lipid synthesis related genes, including SREBP1C, ACC1 mRNA expression (F≥5.66, P < 0.01) and FASN, ACC1 protein expression (F≥38.34, P < 0.05) in hepatocytes, and decreased lipid decomposition gene ACOX1 protein expression (F=154.48, P < 0.01). CM-OA affected several lipid metabolism genes expression. CM-DHA significantly increased CPT1A mRNA expression (F = 10.30, P < 0.01) and ACOX1, CPT1A protein expression (F≥47.06, P < 0.05), and decreased SREBP1C, ACC1 protein expression (F≥65.84, P < 0.05) in hepatocytes. Massive lipid droplets were deposited in hepatocytes in CM-PA treated hepatocytes, and a few amount of lipid droplets were deposited in CM-DHA treated hepatocytes. Conclusion: Different fatty acids affect the balance of lipid metabolism in hepatocytes and liver by inducing macrophage M1 / M2 polarization, thus promoting or delaying the progression of non-alcoholic fatty liver disease.

Non-alcoholic fatty liver disease and lifestyle modifications, focusing on physical activity.

Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver disease, and the prevalence of non-alcoholic steatohepatitis (NASH) with fibrosis is increasing as the population with NAFLD ages. To date, lifestyle modifications including weight loss, increased physical activity, and dietary changes remain the treatment of choice for NAFLD because there are no approved effective pharmacologic agents. Increased physical activity has therapeutic effects on NAFLD by reducing hepatic fat independent of weight reduction. Indeed, even minimal physical activity below the recommended threshold may have a beneficial impact on NAFLD. Aerobic activity and resistance training have similar effects on NAFLD. Universal recommendations for the optimal intensity and dose of physical activity have not been established. Therefore, physical activity should be tailored based on a patient’s clinical characteristics, comorbidities, and fitness capacity. Physical activity also prevents the development of NAFLD and may represent a valuable strategy for reducing the public health burden. However, there are insufficient data supporting the effects of physical activity on the progression of non-alcoholic fatty liver to NASH with advanced fibrosis, and on extrahepatic disease-related morbidity and mortality. In this paper, we review the role of physical activity in the management of NAFLD.

Gut microbia dysbiosis in non-alcoholic fatty liver disease

Non-alcoholic fatty liver disease (NAFLD) represents the spectrum of fatty liver that ranges from simple steatosis to steatohepatitis leading to fibrosis and cirrhosis. NAFLD is often considered the hepatic manifestation of metabolic syndrome and it is closely associated with diabetes, insulin resistance and obesity. Prevalence of NAFLD ranges from 10–35% and the differences in prevalence are due to differences in diagnostic methods used to diagnose NAFLD, as well as differences in NAFLD prevalence amongst various ethnic groups. However, the progression of non-alcoholic fatty liver (NAFL) to non-alcoholic steatohepatits (NASH) is seen in 3–5% of the population, and this subgroup is at a high risk of progression to advanced fibrosis including cirrhosis and hepatocellular carcinoma (HCC) (1,2).

Telomeres, NAFLD and Chronic Liver Disease

Telomeres consist of repeat DNA sequences located at the terminal portion of chromosomes that shorten during mitosis, protecting the tips of chromosomes. During chronic degenerative conditions associated with high cell replication rate, progressive telomere attrition is accentuated, favoring senescence and genomic instability. Several lines of evidence suggest that this process is involved in liver disease progression: (a) telomere shortening and alterations in the expression of proteins protecting the telomere are associated with cirrhosis and hepatocellular carcinoma; (b) advanced liver damage is a feature of a spectrum of genetic diseases impairing telomere function, and inactivating germline mutations in the telomerase complex (including human Telomerase Reverse Transcriptase (hTERT) and human Telomerase RNA Component (hTERC)) are enriched in cirrhotic patients independently of the etiology; and (c) experimental models suggest that telomerase protects from liver fibrosis progression. Conversely, reactivation of telomerase occurs during hepatocarcinogenesis, allowing the immortalization of the neoplastic clone. The role of telomere attrition may be particularly relevant in the progression of nonalcoholic fatty liver, an emerging cause of advanced liver disease. Modulation of telomerase or shelterins may be exploited to prevent liver disease progression, and to define specific treatments for different stages of liver disease.