Risk factors and mechanisms of non-alcoholic steatohepatitis

The worldwide proportion of overweight and obese individuals has increased yearly for more than a decade. Along with rates of obesity, the incidence of co-morbid conditions such as type 2 diabetes, cardiovascular disease and liver disease has also increased. The form of liver disease associated with obesity is termed non-alcoholic steatohepatitis (NASH) due to the histological similarities to livers of chronic alcoholics. NASH has been observed in adult as well as pediatric populations; however, the etiology of this disease is still unknown. This review outlines some of the risk factors commonly associated with NASH and describes molecular mechanisms proposed to underlie disease pathogenesis.     

Hepatocellular carcinoma arising in non-alcoholic steatohepatitis

The incidence and significance of hepatocellular carcinoma (HCC) in non-alcoholic steatohepatitis (NASH) has not been previously evaluated in detail. We recently experienced a case of NASH with multicentric HCC in a female patient. At the age of 58 years, the patient was diagnosed with non-insulin-dependent diabetes mellitus, treated by insulin therapy. The patient did not drink alcohol. She was negative for all serological markers of hepatitis B and C virus infection. Because of liver dysfunction, a needle biopsy was performed at the age of 62 years, and pathological findings, such as fatty change, Mallory's body, nuclear glycogen and pericellular fibrosis, suggested a diagnosis of NASH. Subsequently, four nodules were detected in the liver by imaging. Liver biopsies were performed from each nodule. One nodule was pathologically diagnosed as a pseudolymphoma, while three other nodules were moderately differentiated HCC (10 years after the diagnosis of non-alcoholic steatohepatitis), well-differentiated HCC (11 years later) and dysplastic nodule (11 years later), suggesting multicentric occurrence of HCC. This case suggests that HCC could be a late complication of NASH.     

Obesity and steatohepatitis. Histologic aspects

The traditional concept of adipose tissue as a passive reservoir for energy storage is no longer valid because it has been demonstrated that adipose tissue is a complex, essential, and highly active metabolic and endocrine organ that not only responds to afferent signals from traditional hormone systems and the central nervous system (CNS), but also expresses and secretes factors with important endocrine functions. These factors include leptin and other cytokines. Adipose tissue is also a major site for metabolism of sex steroids and glucocorticoids. The important endocrine function of adipose tissue is emphasized by adverse metabolic consequences of both adipose tissue excess and deficiency. Adipose tissue excess, particularly in visceral compartment, is associated with insulin resistance, hyperglycemia, dyslipidemia, hypertension, and prothrombotic and proinflammatory states. Liver is one of the principal targets of lipid-associated damage by mechanisms that involve apoptosis activation by source of tumoral necrosis factor-alpha and caspase activation and liberation of oxygen-reactive species by oxidative stress and enzymatic chains such as P450, CYP2E1, and CYP3A4, resulting in a continuum involving non alcohol-related fatty liver, non-alcoholic steatohepatitis with or without fibrosis, and liver cirrhosis. This work presents an overview of endocrine functions of adipose tissue and its influence on mechanisms of liver damage.     

Pathogenesis and treatment of non-alcoholic steatohepatitis and its fibrosis

The initial presentation of non-alcoholic steatohepatitis (NASH) is hepatic steatosis. The dysfunction of lipid metabolism within hepatocytes caused by genetic factors, diet, and insulin resistance causes lipid accumulation. Lipotoxicity, oxidative stress, mitochondrial dysfunction, and endoplasmic reticulum stress would further contribute to hepatocyte injury and death, leading to inflammation and immune dysfunction in the liver. During the healing process, the accumulation of an excessive amount of fibrosis might occur while healing. During the development of NASH and liver fibrosis, the gut-liver axis, adipose-liver axis, and renin-angiotensin system (RAS) may be dysregulated and impaired. Translocation of bacteria or its end-products entering the liver could activate hepatocytes, Kupffer cells, and hepatic stellate cells, exacerbating hepatic steatosis, inflammation, and fibrosis. Bile acids regulate glucose and lipid metabolism through Farnesoid X receptors in the liver and intestine. Increased adipose tissue-derived non-esterified fatty acids would aggravate hepatic steatosis. Increased leptin also plays a role in hepatic fibrogenesis, and decreased adiponectin may contribute to hepatic insulin resistance. Moreover, dysregulation of peroxisome proliferator-activated receptors in the liver, adipose, and muscle tissues may impair lipid metabolism. In addition, the RAS may contribute to hepatic fatty acid metabolism, inflammation, and fibrosis. The treatment includes lifestyle modification, pharmacological therapy, and non-pharmacological therapy. Currently, weight reduction by lifestyle modification or surgery is the most effective therapy. However, vitamin E, pioglitazone, and obeticholic acid have also been suggested. In this review, we will introduce some new clinical trials and experimental therapies for the treatment of NASH and related fibrosis.     

维生素D_3与非酒精性脂肪性肝炎的研究进展

非酒精性脂肪性肝炎(NASH)是非酒精性脂肪性肝病(NAFLD)的最重要形式,可发展为肝硬化和肝癌。在啮齿类动物中发现维生素D3(VD3)具有抗氧化、抗凋亡、抗感染和抗肝纤维化的作用。补充VD3可部分改善NASH的进展。VD3有望成为临床治疗NASH的有效药物。     

内毒素诱发大鼠非酒精性脂肪性肝炎和胰岛素抵抗

目的:观察多次皮下注射小剂量内毒素是否能诱导大鼠发生非酒精性脂肪性肝炎(NASH)与胰岛素抵抗(IR)。方法:将24只大鼠随机分为4组:Ⅰ组为4周正常组;Ⅱ组为4周内毒素组;Ⅲ组为9周正常组;Ⅳ组为9周内毒素组;测定血浆丙氨酸氨基转移酶(ALT)、白细胞介素-10(IL-10)、脂联素(APN)、肿瘤坏死因子-α(TNF-α)、内毒素(ET)、空腹胰岛素(FINS)、空腹血糖(FPG)水平,计算胰岛素抵抗指数(IRI),取肝组织进行HE染色和苏丹Ⅳ染色,以确定是否发生了非酒精性脂肪性肝炎与胰岛素抵抗。结果:4周内毒素组与4周正常组相比,血浆ALT、TNF-α、FINS、ET有升高趋势,IL-10有下降趋势,但均无显著差异(P0.05)。9周内毒素组与9周正常组比较,血浆TNF-α、FINS、FPG、ET以及IRI明显升高,IL-10明显降低且均有显著差异(P0.05);但ALT仅有升高趋势而无显著差异(P0.05),APN亦无明显差异(P0.05)。结论:多次皮下注射小剂量内毒素可使大鼠发生非酒精性脂肪性肝炎与胰岛素抵抗,这提示非酒精性脂肪性肝炎与胰岛素抵抗的发生可能与内毒素密切相关。     

Unusual warm autoimmune hemolytic anemia in non-alcoholic steatohepatitis

Warm autoimmune hemolytic anemia (WAIHA), a rare disease (0.2-1 per 100,000 population), ranges from an indolent form with mild hemolysis to a life-threatening condition that necessitates transfusion of incompatible red cells. WAIHA can be either idiopathic or secondary to medications or to a lymphoproliferative disorder. We report a case of profound hemolytic anemia in a liver-transplant eligible patient who was diagnosed with cirrhosis secondary to non-alcoholic steatohepatitis (NASH). The patient initially was treated with red cell transfusion, iv immunoglobulin, and steroids. He developed acute renal failure that required dialysis. Subsequent management included plasmapheresis and rituximab therapy. The patient developed hepatorenal syndrome and died from progressive hepatic failure. To our knowledge, this is the first report of an association between NASH and WAIHA.     

Mouse models in non-alcoholic fatty liver disease and steatohepatitis research

Non-alcoholic fatty liver disease (NAFLD) represents a histological spectrum of liver disease associated with obesity, diabetes and insulin resistance that extends from isolated steatosis to steatohepatitis and cirrhosis. As well as being a potential cause of progressive liver disease in its own right, steatosis has been shown to be an important cofactor in the pathogenesis of many other liver diseases. Animal models of NAFLD may be divided into two broad categories: those caused by genetic mutation and those with an acquired phenotype produced by dietary or pharmacological manipulation. The literature contains numerous different mouse models that exhibit histological evidence of hepatic steatosis or, more variably, steatohepatitis; however, few replicate the entire human phenotype. The genetic leptin-deficient (ob/ob) or leptin-resistant (db/db) mouse and the dietary methionine/choline-deficient model are used in the majority of published research. More recently, targeted gene disruption and the use of supra-nutritional diets to induce NAFLD have gained greater prominence as researchers have attempted to bridge the phenotype gap between the available models and the human disease. Using the physiological processes that underlie the pathogenesis and progression of NAFLD as a framework, we review the literature describing currently available mouse models of NAFLD, highlight the strengths and weaknesses of established models and describe the key findings that have furthered the understanding of disease pathogenesis.     

The scaffold-dependent function of RIPK1 in experimental non-alcoholic steatohepatitis

Journal of Molecular Medicine -     

Noninvasive assessment of fibrosis in non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH)

The term "non-alcoholic fatty liver disease" (NAFLD) encompasses a wide range of pathological conditions ranging from accumulation of fat (fatty liver) to various degrees of inflammation and fibrosis (NASH), and finally to cryptogenic cirrhosis and its clinical sequelae (HCC, liver decompensation). The progression from one stage to the next can be triggered by genetic and environmental factors alone and also through their interaction. Fatty liver is known to follow a benign course, whereas the presence of inflammation, ballooning degeneration, and fibrosis, which are typical features of NASH, can lead to cirrhosis. Despite the serious risks associated with NASH, there are few tools for monitoring the progression of the disease and identification of high-risk patients. The aim of this article is to review the pros and cons of some noninvasive methods for assessing liver fibrosis in NASH.     

Linagliptin alleviates hepatic steatosis and inflammation in a mouse model of non-alcoholic steatohepatitis

Non-alcoholic steatohepatitis (NASH) is a primary cause of cirrhosis and hepatocellular carcinoma. Dipeptidyl peptidase (DPP)-4 inhibitors are established therapies for type 2 diabetes and although DPP-4 inhibitors can reduce hepatic steatosis, their impact on local inflammation and fibrosis in NASH remains unknown. Using two different experimental treatment regimens (4- and 2-week treatments) in streptozotocin-treated neonatal mice on a high-fat diet, we show that the DPP-4 inhibitor linagliptin (10 and 30 mg/kg) significantly attenuated the NAS score from 4.9 ± 0.6 to 3.7 ± 0.4 and 3.6 ± 0.3, respectively, in the 4-week study. In the 2-week study, linagliptin 10 mg/kg significantly reduced NAS score from 4.1 ± 0.4 to 2.4 ± 0.4. Telmisartan was used as a positive control in both studies and lowered NAS score to 1.9 ± 0.7 and 1.4 ± 0.3, respectively. Due to streptozotocin treatment, elevated glucose levels were unchanged by either drug treatment. Further, linagliptin 10 mg/kg significantly reduced mRNA levels of SOCS-3 (from 1.68 ± 0.2 to 0.83 ± 0.08), IFN-γ (from 4.0 ± 0.5 to 2.3 ± 0.3), and TNF-α (from 5.7 ± 0.5 to 2.13 ± 0.3). The latter observation was confirmed by immunohistochemistry of TNF-α in liver specimens. In addition, using microautoradiography, we showed that the distribution of radiolabeled linagliptin was heterogeneous with the highest density associated with interlobular bile ducts and portal tracts (acini). In conclusion, these studies confirm that linagliptin has high exposure in hepatic tissue and has both anti-inflammatory and anti-steatotic activity in NASH.     

乙酰半胱氨酸纳米活性炭缓释微囊对大鼠非酒精性脂肪性肝炎抗氧化作用的影响

目的 探讨乙酰半胱氨酸纳米活性碳缓释微囊（ACNAC）对非酒精性脂肪性肝炎（NASH）大鼠肝脏抗氧化能力的影响.方法 通过高脂饮食12周诱导建立NASH大鼠动物模型,分别给予ACNAC高、中、低每日不同剂量（800 mg/kg、400 mg/kg和200 mg/kg）与易善复（0.692 mg/kg）、乙酰半胱氨酸（400 mg/kg）连续灌胃8周后,检测血清生化指标、肝匀浆丙二醛（MDA）含量、总超氧化物歧化酶（SOD）活性、还原型谷胱甘肽（GSH）、谷胱甘肽过氧化物酶（GSH-Px）活性及肝脏病理指标.结果 ACNAC中剂量组与高脂组相比,血清ALT、AST明显下降（P＜0.01 、P ＜0.05）,且优于易善复组（P＜0.05）.血清TCHOL 、TG、LDL-C和CR检测结果差异虽无统计学意义,但其数值上ACNAC明显低于高脂组,且优于易善复组和NAC组.ACNAC高、中、低三组与高脂组相比,SOD活力明显升高（P ＜0.01） 、GSH含量明显升高（P＜0.01）,ACNAC中剂量组与高脂组比较GSH-PX活性明显升高（P＜0.05）但略低于易善复组（P＜0.01））.结论 相关剂量的乙酰半胱氨酸纳米活性炭缓释微囊可增强非酒精性脂肪性肝炎大鼠的抗氧化能力.     

Screening of non-alcoholic steatohepatitis (NASH)-related datasets and identification of NASH-related genes

Non-alcoholic steatohepatitis (NASH) is a common liver disease in the western world. The mechanisms behind NASH formation are poorly understood, but there may be multiple targets considering the disease’s multifactorial nature. To explore the genes related to the pathogenesis of NASH, we downloaded clinical data and gene expression of NASH patients from the Gene Expression Omnibus database (GEO). We identified 281 genes with a common expression in two NASH-related datasets ({"type":"entrez-geo","attrs":{"text":"GSE89632","term_id":"89632"}}GSE89632 and {"type":"entrez-geo","attrs":{"text":"GSE83452","term_id":"83452"}}GSE83452), suggesting that they may be related to NASH. Further study showed that Angptl4, Foxo1, and Ttc39B might be essential for NASH progression, and these have been poorly studied. Therefore, we explored their roles in NASH. Our data show that these genes participate in the development of NASH through lipid metabolism. This suggests that the three genes can be used as therapeutic targets in NASH.     

红景天苷对非酒精性脂肪性肝炎大鼠肝组织氧化应激的抑制作用

 目的：观察红景天苷（salidroside ,SDS）对大鼠非酒精性脂肪性肝炎（NASH）肝组织氧化应激的影响。方法：以高脂高胆固醇饮食14周诱导建立大鼠NASH模型,药物干预组在造模第8周末时给予SDS 300 mg·kg-1·d-1体重灌胃连续6周,在14周末检测大鼠血清丙氨酸氨基转移酶(ALT)、天冬氨酸氨基转移酶(AST)、总胆固醇（TC）和甘油三酯（TG）,以及肝组织TC、TG、丙二醛（MDA）、超氧化物歧化酶（SOD）和谷胱甘肽（GSH）含量的变化,ELISA检测8-异前列腺素F2α(8-iso-PGF2α)的水平变化, HE染色观察肝组织病理学变化,免疫组化观察8-羟基脱氧鸟苷酸（8-OHdG）表达的变化。结果：在14 周末, NASH模型组大鼠肝组织病理学检查显示肝组织呈中重度脂肪变性并同时伴有炎症细胞浸润;与正常对照组相比,NASH模型组大鼠血清ALT、AST、TG和TC,以及肝TG、TC和MDA的含量显著上升,而肝SOD和GSH的含量显著下降,8-iso-PGF2α的水平显著上升,免疫组化显示8-OHdG表达的阳性细胞数显著增多。与模型组相比,SDS药物干预组大鼠ALT、AST、TG、TC、MDA和8-iso-PGF2α的含量均显著下降,而肝SOD和GSH的含量显著上升,肝病理学变化得到显著改善,8-OHdG表达的阳性细胞数明显减少。结论：SDS对高脂高胆固醇饮食诱导的NASH有较好的抑制效果,其机制可能与SDS的抗氧化作用有关。     

A murine model for non-alcoholic steatohepatitis showing evidence of association between diabetes and hepatocellular carcinoma

Hepatocellular carcinoma (HCC) is the third most common cause of cancer-related deaths. In addition to hepatitis viral infections, several cohort studies have shown that diabetes mellitus is a risk factor of HCC, making the incidence alarming high. However, it has not been demonstrated directly how diabetes develops to HCC, because of its difficulty to follow changes of liver histology in diabetic populations. Here, we report that non-alcoholic steatohepatitis (NASH) is pivotal to link diabetes with HCC by establishing a novel, reproducible NASH–HCC model in mice. Neonatal male mice exposed to low-dose streptozotocin (STZ) developed liver steatosis with diabetes 1 week after feeding high-fat diet (HFD). Continuous HFD decreased hepatic fat deposit whilst increased lobular inflammation with foam cell-like macrophages, showing NASH pathology. In parallel with decreased phagocytosis of macrophages, fibroblasts accumulated to form “chicken-wired” fibrosis. All mice developed multiple HCC later. Female mice treated with STZ–HFD and male mice treated with STZ alone showed diabetes but never developed HCC by the absence of NASH-based fibrosis. Thus, the present study provides the evidence in novel mouse model that NASH-based fibrosis is an essential histological process for diabetic populations to accelerate the development of HCC.     

Hepatocellular carcinoma with silent and cirrhotic non-alcoholic steatohepatitis, accompanying ectopic liver tissue attached to gallbladder

A 70-year-old man had been obese since youth. He had been treated for hypertension and diabetes mellitus. An abdominal ultrasound showed a mass in the liver. He was admitted to St Marianna University School of Medicine Hospital for further evaluation. There was no history of alcohol use, and hepatitis viral markers and autoantibodies were all negative. Several imaging studies showed overt hepatocellular carcinoma (HCC). Transcatheter arterial embolization was performed, followed by surgical resection. Histopathological examination revealed moderately differentiated HCC. The non-tumor areas had pseudolobules in a diffuse pattern similar to alcoholic cirrhosis. The histological findings in the ectopic liver tissue attached to the gallbladder, which was also resected during surgery, were that there was no cirrhosis, but fine fibrosis with inflammatory cell infiltration of sinusoids. These findings were consistent with non-alcoholic steatohepatitis (NASH). There was probably a progression of similar findings that had developed into cirrhosis. These findings confirmed a diagnosis of HCC, cirrhosis, and underlying NASH in this patient. The present case is important for investigation of the development into cirrhosis and carcinogenesis of NASH. The present case demonstrates the importance of evaluating obese patients with fatty liver for underlying NASH and ongoing follow up for development of cirrhosis and HCC.     

非酒精性脂肪性肝炎的ACE-ID与AT1R-A1166C多态性研究

目的:研究血管紧张素转化酶(ACE)基因ID多态性和血管紧张素Ⅱ1型受体(AT1R)基因A1166C多态性与日本高知地区非酒精性脂肪性肝炎(NASH)易感性的相关性,从基因学角度探讨NASH的发生发展机制,为NASH的预防、诊断和治疗提供一定的理论依据。方法:应用聚合酶链反应-限制性片段长度多态性(PCR-PFLP)方法对日本高知地区104例NASH患者和150例正常人ACE-ID基因和AT1R-A1166C基因多态性进行分析。结果:NASH病例组D等位基因频率显著高于正常对照组(P0.01);NASH病例组的DD基因型频率显著高于正常对照组(P0.01);NASH病例组A等位基因频率与正常对照组相比差异无显著(P(0.05)。NASH病例组AA基因型频率与正常对照组相比差异无显著(P(0.05)。结论:ACE-ID基因位于内含子16上D等位基因与DD基因型与NASH的发生发展显著相关,可能是促进NASH发生的原因之一;AT1R-A1166C基因多态性与NASH的发生发展尚未显示相关性。     

CEACAM1 loss links inflammation to insulin resistance in obesity and non-alcoholic steatohepatitis (NASH)

Mounting epidemiological evidence points to an association between metabolic syndrome and non-alcoholic steatohepatitis (NASH), an increasingly recognized new epidemic. NASH pathologies include hepatocellular ballooning, lobular inflammation, hepatocellular injury, apoptosis, and hepatic fibrosis. We will review the relationship between insulin resistance and inflammation in visceral obesity and NASH in an attempt to shed more light on the pathogenesis of these major metabolic diseases. Moreover, we will identify loss of the carcinoembryonic antigen-related cell adhesion molecule 1 as a unifying mechanism linking the immunological and metabolic abnormalities in NASH.