Different Stages Of Nonalcoholic Fatty Liver Disease Research Articles

Liver Metabolomics Analysis Revealing Key Metabolites Associated with Different Stages of Nonalcoholic Fatty Liver Disease in Hamsters.

Nonalcoholic fatty liver disease (NAFLD) is not only the top cause of liver diseases but also a hepatic-correlated metabolic syndrome. This study performed untargeted metabolomics analysis of NAFLD hamsters to identify the key metabolites to discriminate different stages of NAFLD. Hamsters were fed a high-fat diet (HFD) to establish the NAFLD model with different stages (six weeks named as the NAFLD1 group and twelve weeks as the NAFLD2 group, respectively). Those liver samples were analyzed by untargeted metabolomics (UM) analysis to investigate metabolic changes and metabolites to discriminate different stages of NAFLD. The significant liver weight gain in NAFLD hamsters was observed, accompanied by significantly increased levels of serum triglyceride (TG), total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), alanine aminotransferase (ALT), and aspartate aminotransferase (AST). Moreover, the levels of TG, LDL-C, ALT, and AST were significantly higher in the NAFLD2 group than in the NAFLD1 group. The UM analysis also revealed the metabolic changes; 27 differently expressed metabolites were detected between the NAFLD2 and NAFLD1 groups. More importantly, the levels of N-methylalanine, allantoin, glucose, and glutamylvaline were found to be significantly different between any two groups (control, NAFLD2 and NAFLD1). Receiver operating characteristic curve (ROC) curve results also showed that these four metabolites are able to distinguish control, NAFLD1 and NAFLD2 groups. This study indicated that the process of NAFLD in hamsters is accompanied by different metabolite changes, and these key differently expressed metabolites may be valuable diagnostic biomarkers and responses to therapeutic interventions.

Effect of Intermittent Fasting on Lipid Profile, Anthropometric and Hepatic Markers in Non-Alcoholic Fatty Liver Disease (NAFLD): A Systematic Review.

The first-line treatment for non-alcoholic fatty liver disease (NAFLD) is lifestyle modification; this should accompany any pharmacological intervention. Intermittent fasting (IF) has shown benefits over metabolic and cardiovascular parameters. Non-religious IF includes Time-Restricted Feeding (TRF), Alternate-Day Fasting (ADF), and 5:2 IF interventions. To evaluate the effects of IF on anthropometric, liver damage, and lipid profile markers in subjects with NAFLD. A bibliographic search was carried out according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines using PubMed and Scopus databases. Five studies involving 470 patients with NAFLD were included. In relation to anthropometric markers, all the articles reported body weight reduction (2.48-7.63%), but only ADF and 5:2 IF reported a body weight reduction >5%; also, all the articles reported fat mass reduction. Concerning hepatic markers, all the articles reported a reduction in hepatic steatosis and alanine aminotransferase activity, but no changes in fat-free mass and high-density lipoprotein cholesterol levels. There were variable results on fibrosis, other liver enzymes, waist circumference and body mass index, as well as the levels of triglycerides, total cholesterol, and low-density lipoprotein cholesterol. Any form of IF could be potentially beneficial for NAFLD treatment and some associated cardiometabolic parameters. However, it is necessary to evaluate the effects and safety of IF in long-term studies involving a higher number of participants with different stages of NAFLD. The effect of IF on NAFLD-associated vascular risk also needs evaluation.

Quasi-LD-Targeted and ONOO--Responsive Fluorescent Probe for Investigating the Interaction of Nonalcoholic Fatty Liver with Drug-Induced Liver Injury.

Hepatic lipid droplets (LDs) and peroxynitrite (ONOO-) levels are closely related to nonalcoholic fatty liver disease (NAFLD). Additionally, some drug-induced liver injury (DILI) is often associated with ONOO-. Here, we constructed and screened the quasi-LDs-targeted and ONOO--responsive fluorescent probe MBDP-Py+ to investigate the interaction of NAFLD with DILI. By monitoring the upregulation of the ONOO- levels and the accumulation of LDs, MBDP-Py+ was more sensitive and efficient than tissue staining and serum markers detection in evaluating the early toxicity of NAFLD and diagnosing the anticancer-DILI. More importantly, the sensitive enhancement of fluorescence signals demonstrated that in different stages of NAFLD, the dominant element of liver injury was different in the NAFLD combined with DILI mice models. As the degree of NAFLD deepens, the synergistic effect of the two will lead to more serious liver damage.

Ferroptosis in non-alcoholic liver disease: Molecular mechanisms and therapeutic implications.

Ferroptosis refers to a novel modality of regulated cell death characterized by excessive iron accumulation and overwhelming lipid peroxidation, which takes an important part in multiple pathological processes associated with cell death. Considering the crucial roles of the liver in iron and lipid metabolism and its predisposition to oxidative insults, more and more studies have been conducted to explore the relationship between ferroptosis and various liver disorders, including non-alcoholic fatty liver disease (NAFLD). With increased morbidity and high mortality rates, NAFLD has currently emerged as a global public health issue. However, the etiology of NAFLD is not fully understood. In recent years, an accumulating body of evidence have suggested that ferroptosis plays a pivotal role in the pathogenesis of NAFLD, but the precise mechanisms underlying how ferroptosis affects NAFLD still remain obscure. Here, we summarize the molecular mechanisms of ferroptosis and its complicated regulation systems, delineate the different effects that ferroptosis exerts in different stages of NAFLD, and discuss some potential effective therapies targeting ferroptosis for NAFLD treatment, which putatively points out a novel direction for NAFLD treatment.

Hepatic and serum branched-chain fatty acid profile in patients with nonalcoholic fatty liver disease: A case-control study.

Alterations in the hepatic lipidome are a crucial factor involved in the pathophysiology of nonalcoholic fatty liver disease (NAFLD). The aim of this study was to evaluate the serum and hepatic profile of branched-chain fatty acids (BCFAs) in patients with different stages of NAFLD. This was a case-control study performed in 27 patients without NAFLD, 49 patients with nonalcoholic fatty liver,and 17 patients with nonalcoholic steatohepatitis, defined by liver biopsies. Serum and hepatic levels of BCFAs were analyzed by gas chromatography-mass spectrometry. The hepatic expression of genes involved in the endogenous synthesis of BCFAs was analyzed byreal-time quantitative polymerase chain reaction (RT-qPCR). A significant increase in hepatic BCFAs was found in subjects with NAFLD compared with those without NAFLD; no differences were observed in serum BCFAs between study groups. Trimethyl BCFAs, iso-BCFAs, and anteiso-BCFAs were increased in subjects with NAFLD (either nonalcoholic fatty liver or nonalcoholic steatohepatitis) compared with those without NAFLD. Correlation analysis showed a relationship between hepatic BCFAs and the histopathological diagnosis of NAFLD, as well as other histological and biochemical parameters related to this disease. Gene expression analysis in liver showed that the mRNA levels of BCAT1, BCAT2, and BCKDHA were upregulated in patients with NAFLD. These results suggest that the increased production of liver BCFAs might be related to NAFLD development and progression.

S100 Proteins in Fatty Liver Disease and Hepatocellular Carcinoma.

Non-alcoholic fatty liver disease (NAFLD) is a highly prevalent and slow progressing hepatic pathology characterized by different stages of increasing severity which can ultimately give rise to the development of hepatocellular carcinoma (HCC). Besides drastic lifestyle changes, few drugs are effective to some extent alleviate NAFLD and HCC remains a poorly curable cancer. Among the deregulated molecular mechanisms promoting NAFLD and HCC, several members of the S100 proteins family appear to play an important role in the development of hepatic steatosis, non-alcoholic steatohepatitis (NASH) and HCC. Specific members of this Ca2+-binding protein family are indeed significantly overexpressed in either parenchymal or non-parenchymal liver cells, where they exert pleiotropic pathological functions driving NAFLD/NASH to severe stages and/or cancer development. The aberrant activity of S100 specific isoforms has also been reported to drive malignancy in liver cancers. Herein, we discuss the implication of several key members of this family, e.g., S100A4, S100A6, S100A8, S100A9 and S100A11, in NAFLD and HCC, with a particular focus on their intracellular versus extracellular functions in different hepatic cell types. Their clinical relevance as non-invasive diagnostic/prognostic biomarkers for the different stages of NAFLD and HCC, or their pharmacological targeting for therapeutic purpose, is further debated.

Identification of Shared Gene Signatures in Different Stages of Nonalcoholic Fatty Liver Disease Using Integrated Microarray Datasets

Background: Nonalcoholic fatty liver disease (NAFLD) is the most common type of chronic liver disease worldwide. Left untreated, it can be a risk factor for developing cirrhosis or hepatocellular carcinoma (HCC). Although experts have made many efforts to find the underlying mechanisms of NAFLD, they remain a mystery. Objectives: This study aimed to distinguish common gene signatures and pathways in the human liver during NAFLD progression through systems biology. Methods: In this study, the researchers selected three microarray datasets, GSE48452, GSE63067, and GSE89632, from the NCBI GEO database to explore differentially expressed genes (DEGs) among healthy controls, simple steatosis, and nonalcoholic steatohepatitis (NASH) patients. Furthermore, protein-protein interaction (PPI) networks and pathway enrichment analyses were used to detect common genes and biological pathways in different stages of NAFLD. Results: The current study included 45 healthy participants, 36 simple steatosis patients, and 46 NASH patients. Common genes for NAFLD progression were Chi3L1, ICAM1, MT1A, MT1H, ABCB11, ACOT1, CYP2C9, HSP90B1, and CPB2, which are involved in inflammation and oxidative stress pathways. Conclusions: The present study investigated the shared vital genes and pathways between different stages of NAFLD, which may facilitate understanding NAFLD mechanisms and identifying potential therapeutic targets in this disease.

Identification of novel targets in adipose tissue involved in non-alcoholic fatty liver disease progression.

Obesity is a major risk factor for the development of Nonalcoholic fatty liver disease (NAFLD). We hypothesize that a dysfunctional subcutaneous white adipose tissue (scWAT) may lead to an accumulation of ectopic fat in the liver. Our aim was to investigate the molecular mechanisms involved in the causative role of scWAT in NALFD progression. We performed a RNA-sequencing analysis in a discovery cohort (n=45) to identify genes in scWAT correlated with fatty liver index, a qualitative marker of liver steatosis. We then validated those targets in a second cohort (n=47) of obese patients who had liver biopsies available. Finally, we obtained scWAT mesenchymal stem cells (MSCs) from 13 obese patients at different stages of NAFLD and established in vitro models of human MSC (hMSC)-derived adipocytes. We observed impaired adipogenesis in hMSC-derived adipocytes as liver steatosis increased, suggesting that an impaired adipogenic capacity is a critical event in the development of NAFLD. Four genes showed a differential expression pattern in both scWAT and hMSC-derived adipocytes, where their expression paralleled steatosis degree: SOCS3, DUSP1, SIK1, and GADD45B. We propose these genes as key players in NAFLD progression. They could eventually constitute potential new targets for future therapies against liver steatosis.

Effects of Bariatric Endoscopy on Non-Alcoholic Fatty Liver Disease: A Comprehensive Systematic Review and Meta-Analysis.

Background and objectiveEndoscopic bariatric and metabolic therapies (EBMTs) are emerging minimally invasive therapeutic options for obesity and its related complications, including non-alcoholic fatty liver disease (NAFLD). This study aimed to evaluate the effects of EBMTs on NALFD in patients with obesity.MethodsFour databases were searched until Nov 2021. Randomized controlled trials (RCTs) and observational studies reporting liver-related outcomes following Food and Drug Administration (FDA)-approved and non-FDA-approved EBMTs were included. Liver parameters, metabolic parameters, and weight loss were evaluated. Risk of bias was assessed using the “risk of bias” tool in the Cochrane Collaboration for RCTs and the Methodological Index for Non-Randomized Studies criteria for observational studies.ResultsThirty-three studies with 1710 individuals were included. Regarding the effects of EBMTs on liver fibrosis, a significant decline of NAFLD Fibrosis Score, but not transient elastography-detected liver stiffness or Fibrosis-4 Index, was observed. EBMTs significantly improved liver steatosis (control attenuation parameter and Hepatic Steatosis Index), NAFLD Activity Score, and Homeostasis Model Assessment of Insulin Resistance. EBMTs reduced serum levels of alanine transaminase, aspartate aminotransferase, and gamma-glutamyl transpeptidase considerably. Moreover, EBMTs had reducing effects on the serum levels of triglycerides and total cholesterol as well as body weight.ConclusionsOur meta-analysis suggested that EBMTs could ameliorate NAFLD based on the evidence of improved liver steatosis, liver function, and insulin resistance. Large-scale, prospective, long-term studies are warranted to clarify the role of EBMTs in patients with different stages of NAFLD.

The Role and Mechanisms of Selenium Supplementation on Fatty Liver-Associated Disorder.

Non-alcoholic fatty liver disease (NAFLD) is the most frequent chronic liver disease without effective therapy. Selenium, as an essential trace element for humans, is notable for its antioxidant properties. The previous study shows that selenium levels in NAFLD patients are lower than normal ones. Selenium supplementation can effectively alleviate metabolic disorders by relieving anti-oxidative stress and anti-inflammatory regulation. However, the correlation between selenium and NAFLD has not been fully clarified. Herein, we review the current studies on selenium in regulating the different stages of NAFLD and summarize relevant clinical trials to highlight the potential roles of selenium in NAFLD treatment.

Phytochemicals targeting NAFLD through modulating the dual function of forkhead box O1 (FOXO1) transcription factor signaling pathways.

Literature evidence reveals that natural compounds are potential candidates for ameliorating obesity-associated non-alcoholic fatty liver disease (NAFLD) by targeting forkhead box O1 (FOXO1) transcription factor. FOXO1 has a dual and complex role in regulating both increase and decrease in lipid accumulation in hepatocytes and adipose tissues (AT) at different stages of NAFLD. In insulin resistance (IR), it is constitutively expressed, resulting in increased hepatic glucose output and lipid metabolism irregularity. The studies on different phytochemicals indicate that dysregulation of FOXO1 causes disturbance in cellular nutrients homeostasis, and the natural entities have an enduring impact on the mitigation of these abnormalities. The current review communicates and evaluates certain phytochemicals through different search engines, targeting FOXO1 and its downstream cellular pathways to find lead compounds as potential therapeutic agents for treating NAFLD and related metabolic disorders. The findings of this review confirm that polyphenols, flavonoids, alkaloids, terpenoids, and anthocyanins are capable of modulating FOXO1 and associated signaling pathways, and they are potential therapeutic agents for NAFLD and related complications. HIGHLIGHTS: • FOXO1 has the potential to be targeted by novel drugs from natural sources for the treatment of NAFLD and obesity. • FOXO1 regulates cellular autophagy, inflammation, oxidative stress, and lipogenesis through alternative mechanisms. • Phytochemicals treat NAFLD by acting on FOXO1 or SREBP1c and PPARγ transcription factor signaling pathways.

Serum proteome profiling reveals differentially expressed proteins between subjects with metabolically healthy obesity and nonalcoholic fatty liver disease

Nonalcoholic fatty liver disease (NAFLD) is the leading risk factor for common chronic liver disease and is often regarded as a prevalent metabolic disorder tightly associated with obesity. However, the existence of metabolically healthy obesity (MHO) indicates that some important factors may participate in protecting individuals with MHO free of NAFLD, even with excessive adiposity. To explore factors independent of obesity that may be involved in the occurrence of NAFLD, we performed an iTRAQ-based proteomic study to identify proteins differentially expressed in serum between NAFLD and MHO subjects. Compared with the MHO group, ten proteins were upregulated and five were downregulated significantly in the NAFLD group. Gene Ontology analysis indicated significant changes in the immune response and triglyceride metabolism-related pathways between MHO and NAFLD. We further validated three candidates markedly dysregulated in NAFLD by Western blotting and ELISA, including two upregulated proteins (afamin and apolipoprotein H) and one downregulated protein (apolipoprotein C-1). Detection of serum apolipoprotein H levels in a large-scale cohort with MHO and different stages of NAFLD indicated that apolipoprotein H may be a potential blood biomarker for distinguishing NAFLD from MHO and an independent risk factor for predicting NAFLD.

Long non-coding RNA in Non-alcoholic fatty liver disease.

Nonalcoholic fatty liver disease (NAFLD) is a chronic disease of worldwide impact. The disease process begins with steatosis, i.e., fat accumulation in the liver, and proceeds to nonalcoholic steatohepatitis (NASH). Liver biopsy is the gold standard for NASH diagnosis, but the procedure is invasive, expensive, error prone and poses considerable risk. Unfortunately, there are currently no precise FDA-approved therapies for NAFLD, the only options being lifestyle change and symptomatic treatment. Recently, much research has focused on the identification of molecular mechanisms that could be translated into novel diagnostics and therapeutics. With the advent of high throughput genomics and transcriptomics, noncoding RNAs, including long non-coding RNAs (lncRNAs) have been identified as key players of NAFLD pathogenesis and have accordingly attracted much attention as potential diagnostics and therapeutics. In this chapter, we reviewed various lncRNAs and their functions at different stages of NAFLD. We also highlighted how these unique molecules can be developed as stage-specific non-invasive diagnostic biomarkers for NAFLD.

Understanding the Role of the Gut Microbiome and Microbial Metabolites in Non-Alcoholic Fatty Liver Disease: Current Evidence and Perspectives.

Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver disease worldwide. NAFLD begins as a relatively benign hepatic steatosis which can evolve to non-alcoholic steatohepatitis (NASH); the risk of cirrhosis and hepatocellular carcinoma (HCC) increases when fibrosis is present. NAFLD represents a complex process implicating numerous factors—genetic, metabolic, and dietary—intertwined in a multi-hit etiopathogenetic model. Recent data have highlighted the role of gut dysbiosis, which may render the bowel more permeable, leading to increased free fatty acid absorption, bacterial migration, and a parallel release of toxic bacterial products, lipopolysaccharide (LPS), and proinflammatory cytokines that initiate and sustain inflammation. Although gut dysbiosis is present in each disease stage, there is currently no single microbial signature to distinguish or predict which patients will evolve from NAFLD to NASH and HCC. Using 16S rRNA sequencing, the majority of patients with NAFLD/NASH exhibit increased numbers of Bacteroidetes and differences in the presence of Firmicutes, resulting in a decreased F/B ratio in most studies. They also present an increased proportion of species belonging to Clostridium, Anaerobacter, Streptococcus, Escherichia, and Lactobacillus, whereas Oscillibacter, Flavonifaractor, Odoribacter, and Alistipes spp. are less prominent. In comparison to healthy controls, patients with NASH show a higher abundance of Proteobacteria, Enterobacteriaceae, and Escherichia spp., while Faecalibacterium prausnitzii and Akkermansia muciniphila are diminished. Children with NAFLD/NASH have a decreased proportion of Oscillospira spp. accompanied by an elevated proportion of Dorea, Blautia, Prevotella copri, and Ruminococcus spp. Gut microbiota composition may vary between population groups and different stages of NAFLD, making any conclusive or causative claims about gut microbiota profiles in NAFLD patients challenging. Moreover, various metabolites may be involved in the pathogenesis of NAFLD, such as short-chain fatty acids, lipopolysaccharide, bile acids, choline and trimethylamine-N-oxide, and ammonia. In this review, we summarize the role of the gut microbiome and metabolites in NAFLD pathogenesis, and we discuss potential preventive and therapeutic interventions related to the gut microbiome, such as the administration of probiotics, prebiotics, synbiotics, antibiotics, and bacteriophages, as well as the contribution of bariatric surgery and fecal microbiota transplantation in the therapeutic armamentarium against NAFLD. Larger and longer-term prospective studies, including well-defined cohorts as well as a multi-omics approach, are required to better identify the associations between the gut microbiome, microbial metabolites, and NAFLD occurrence and progression.

Ursodeoxycholic Acid Treatment Restores Gut Microbiota and Alleviates Liver Inflammation in Non-Alcoholic Steatohepatitic Mouse Model.

Gut microbiota dysbiosis plays an important role in the progression of non-alcoholic fatty liver disease (NAFLD), and no approved drugs are available for NAFLD treatment. In this study, we aimed to explore the dynamic changes of gut microbiota at the different stages of NAFLD and determine whether ursodeoxycholic acid (UDCA) could improve liver histopathological features of non-alcoholic steatohepatitis (NASH) mice induced by a high-fat high-cholesterol (HFHC) diet and its impact on gut microbiota. 6-week-old male C57BL/6 mice were fed with a HFHC or normal diet for 12, 18, and 24 weeks, respectively, to simulate the different stages of NAFLD. 16s ribosomal RNA genes from mice fecal samples at the different time points were sequenced to evaluate the dynamic changes of the gut microbiota. Then, C57BL/6 mice were fed with a HFHC diet for 24 weeks to establish the NASH model. Different doses of UDCA were administered intragastrically for additional 4 weeks. Normal diet–fed mice were taken as control. Serum samples, liver, and intestine tissues were harvested for biochemical tests and histopathological examinations. 16s ribosomal RNA genes from mice fecal samples were sequenced to assess the structural changes of gut microbiota. HFHC diet–fed mice developed simple steatosis, steatohepatitis, and fibrosis at 12, 18, and 24 weeks, respectively. The profile of gut microbiota dynamically changed with the different stages of NAFLD. NASH mice had significantly higher abundance of Fecalibaculum, Coriobacteriaceae_UCG-002, and Enterorhabdus, and lower abundance of norank_f_Muribaculaceae, Bacteroides, and Alistipes, which were partially restored by UDCA treatment. UDCA treatment significantly attenuated hepatic inflammation of NASH mice as indicated by the sum of ballooning and lobular inflammation of the NALFD activity score (3.2 ± 0.8 vs 1.8 ± 0.8, p = 0.029), and partially restored gut microbiota dysbiosis, and increased the expression of Claudin-1 and ZO-1 in the intestine, but did not activate the suppressed Farnesoid X receptor signal pathway. Conclusions: The gut microbiota dynamically changes with the different stages of NAFLD. UDCA treatment (120 mg/kg) could partially restore gut microbiota, repair gut barrier integrity, and attenuate hepatic inflammation in the NASH mouse model.

Сучасні підходи до діагностики та скринінгу метаболічних порушень у хворих із неалкогольною жировою хворобою печінки

Неалкогольна жирова хвороба печінки (НАЖХП) є найпоширенішою патологією серед усіх хронічних захворювань печінки, що призводить до погіршення якості життя, інвалідизації і смерті. Згідно із сучасними уявленнями, НАЖХП належить до одного з компонентів метаболічного синдрому, що підкреслює її взаємозв’язок із цукровим діабетом типу 2 та серцево-судинними захворюваннями. В цій статті узагальнено підходи щодо сучасних методів інструментально-лабораторної діагностики різних стадій НАЖХП. Також акцентовано увагу на проведенні скринінгу метаболічних порушень та оцінці серцево-судинного ризику в цієї когорти хворих, оскільки НАЖХП є раннім предиктором розвитку зазначених патологічних станів.

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.

Changes in Key Mitochondrial Lipids Accompany Mitochondrial Dysfunction and Oxidative Stress in NAFLD.

Nonalcoholic fatty liver disease (NAFLD) is a dysmetabolic hepatic damage of increasing severity: simple fat accumulation (steatosis), nonalcoholic steatohepatitis (NASH), and hepatic fibrosis. Oxidative stress is considered an important factor in producing hepatocyte injury associated with NAFLD progression. Studies also suggest a link between the accumulation of specific hepatic lipid species, mitochondrial dysfunction, and the progression of NAFLD. However, it is unclear whether mitochondrial lipid modifications are involved in NAFLD progression. To gain insight into the relationship between mitochondrial lipids and disease progression through different stages of NAFLD, we performed lipidomic analyses on mouse livers at different stages of western diet-induced NAFLD, with or without hepatic fibrosis. After organelle separation, we studied separately the mitochondrial and the “nonmitochondrial” hepatic lipidomes. We identified 719 lipid species from 16 lipid families. Remarkably, the western diet triggered time-dependent changes in the mitochondrial lipidome, whereas the “nonmitochondrial” lipidome showed little difference with levels of hepatic steatosis or the presence of fibrosis. In mitochondria, the changes in the lipidome preceded hepatic fibrosis. In particular, two critical phospholipids, phosphatidic acid (PA) and cardiolipin (CL), displayed opposite responses in mitochondria. Decrease in CL and increase in PA were concurrent with an increase of coenzyme Q. Electron paramagnetic resonance spectroscopy superoxide spin trapping and Cu2+ measurement showed the progressive increase in oxidative stress in the liver. Overall, these results suggest mitochondrial lipid modifications could act as an early event in mitochondrial dysfunction and NAFLD progression.

Bioengineered Multicellular Liver Microtissues for Modeling Advanced Hepatic Fibrosis Driven Through Non-Alcoholic Fatty Liver Disease.

Despite considerable efforts in modeling liver disease in vitro, it remains difficult to recapitulate the pathogenesis of the advanced phases of non-alcoholic fatty liver disease (NAFLD) with inflammation and fibrosis. Here, a liver-on-a-chip platform with bioengineered multicellular liver microtissues is developed, composed of four major types of liver cells (hepatocytes, endothelial cells, Kupffer cells, and stellate cells) to implement a human hepatic fibrosis model driven by NAFLD: i) lipid accumulation in hepatocytes (steatosis), ii) neovascularization by endothelial cells, iii) inflammation by activated Kupffer cells (steatohepatitis), and iv) extracellular matrix deposition by activated stellate cells (fibrosis). In this model, the presence of stellate cells in the liver-on-a-chip model with fat supplementation showed elevated inflammatory responses and fibrosis marker up-regulation. Compared to transforming growth factor-beta-induced hepatic fibrosis models, this model includes the native pathological and chronological steps of NAFLD which shows i) higher fibrotic phenotypes, ii) increased expression of fibrosis markers, and iii) efficient drug transport and metabolism. Taken together, the proposed platform will enable a better understanding of the mechanisms underlying fibrosis progression in NAFLD as well as the identification of new drugs for the different stages of NAFLD.

Computational Hypothesis: How Intra-Hepatic Functional Heterogeneity May Influence the Cascading Progression of Free Fatty Acid-Induced Non-Alcoholic Fatty Liver Disease (NAFLD).

Non-Alcoholic Fatty Liver Disease (NAFLD) is the most common type of chronic liver disease in developed nations, affecting around 25% of the population. Elucidating the factors causing NAFLD in individual patients to progress in different rates and to different degrees of severity, is a matter of active medical research. Here, we aim to provide evidence that the intra-hepatic heterogeneity of rheological, metabolic and tissue-regenerating capacities plays a central role in disease progression. We developed a generic mathematical model that constitutes the liver as ensemble of small liver units differing in their capacities to metabolize potentially cytotoxic free fatty acids (FFAs) and to repair FFA-induced cell damage. Transition from simple steatosis to more severe forms of NAFLD is described as self-amplifying process of cascading liver failure, which, to stop, depends essentially on the distribution of functional capacities across the liver. Model simulations provided the following insights: (1) A persistently high plasma level of FFAs is sufficient to drive the liver through different stages of NAFLD; (2) Presence of NAFLD amplifies the deleterious impact of additional tissue-damaging hits; and (3) Coexistence of non-steatotic and highly steatotic regions is indicative for the later occurrence of severe NAFLD stages.