Limitations of liver biopsy and non-invasive diagnostic tests for the diagnosis of nonalcoholic fatty liver disease/nonalcoholic steatohepatitis

It is estimated that 30%of the adult population in Japan is affected by nonalcoholic fatty liver disease(NAFLD).Fatty changes of the liver are generally diagnosed using imaging methods such as abdominal ultrasonography(US)and computed tomography(CT),but the sensitivity of these imaging techniques is low in cases of mild steatosis.Alanine aminotransferase levels may be normal in some of these patients,warranting the necessity to establish a set of parameters useful for detecting NAFLD,and the more severe form of the disease,nonalcoholic steatohepatitis(NASH).Although liver biopsy is currently the gold standard for diagnosing progressive NASH,it has many drawbacks,such as sampling error,cost,and risk of complications.Furthermore,it is not realistic to perform liver biopsies on all NAFLD patients.Diagnosis of NASH using various biomarkers,scoring systems and imaging methods,such as elastography,has recently been attempted.The NAFIC score,calculated from the levels of ferritin,fasting insulin,and typeⅣcollagen 7S,is useful for the diagnosis of NASH,while the NAFLD fibrosis score and the FIB-4 index are useful for excluding NASH in cases of advanced fibrosis.This article reviews the limitations and merits of liver biopsy and noninvasive diagnostic tests in the diagnosis of NAFLD/NASH.     

Role of liver biopsy in nonalcoholic fatty liver disease

Nonalcoholic fatty liver disease (NAFLD), defined as abnormal accumulation (> 5%) of hepatic triglyceride without excess alcohol intake, is the most common form of chronic liver disease in adults and children in the United States. NAFLD encompasses a spectrum of histologic findings including uncomplicated steatosis, steatosis with inflammation and steatohepatitis [nonalcoholic steatohepatitis (NASH)]; the latter can advance to cirrhosis and hepatocellular carcinoma. NASH is currently accepted as the hepatic manifestation of the set of cardiovascular risk factors collectively known as metabolic syndrome. In 1999 a system for histologic grading and staging for NASH was proposed; this was revised by the NASH Clinical Research Network in 2005 for the entire spectrum of lesions in NAFLD, including the lesions and patterns of pediatric NAFLD, and for application in clinical research trials. Diagnosis remains distinct from grade and stage. A recent European proposal separates steatosis from activity to derive a numeric diagnosis of NASH. Even though there have been promising advancements in non-invasive testing, these tests are not yet detailed enough to replace the full range of findings provided by liver biopsy evaluation. Limitations of biopsy are acknowledged, but liver biopsy remains the “gold standard” for diagnosis and determination of amounts of necroinflammatory activity, and location of fibrosis, as well as remodeling of the parenchyma in NASH. This review focuses on the specific histologic lesions of NAFLD and NASH, grading and staging, differential diagnoses to be considered, and the continuing role of the liver biopsy in this important liver disease.     

Histopathology of nonalcoholic fatty liver disease

Histological analysis of liver biopsies remains a standard against which other methods of assessment for the presence and amount of hepatic injury due to nonalcoholic fatty liver disease(NAFLD) are measured.Histological evaluation remains the sole method of distinguishing steatosis from advanced forms of NAFLD,i.e.nonalcoholic steatohepatitis(NASH) and fibrosis.Included in the lesions of NAFLD are steatosis,lobular and portal inflammation,hepatocyte injury in the forms of ballooning and apoptosis,and fibros...     

Non-invasive methods for the diagnosis of nonalcoholic fatty liver disease

Nonalcoholic fatty liver disease(NAFLD) is the commonest chronic liver disease and includes simple steatosis and nonalcoholic steatohepatitis(NASH). Since NASH progresses to cirrhosis more frequently and increases liver-related and cardiovascular disease risk substantially more than simple steatosis, there is a great need to differentiate the two entities. Liver biopsy is the gold standard for the diagnosis of NAFLD but its disadvantages, including the risk of complications and sampling bias, stress the need for developing alternative diagnostic methods. Accordingly, several non-invasive markers have been evaluated for the diagnosis of simple steatosis and NASH, including both serological indices and imaging methods. The present review summarizes the current knowledge on the role of these markers in the diagnosis of NAFLD. Current data suggest that ultrasound and the fibrosis-4 score are probably the most appealing methods for detecting steatosis and for distinguishing NASH from simple steatosis, respectively, because of their low cost and relatively high accuracy. However, currently available methods, both serologic and imaging, cannot obviate the need for liver biopsy for diagnosing NASH due to their substantial false positive and false negative rates. Therefore, the current role of these methods is probably limited in patients who are unwilling or have contraindications for undergoing biopsy.     

Animal models of nonalcoholic fatty liver disease/nonalcoholic steatohepatitis

Nonalcoholic fatty liver disease (NAFLD) is a condition in which excess fat accumulates in the liver of a patient without a history of alcohol abuse. Nonalcoholic steatohepatitis (NASH), a severe form of NAFLD, can progress to liver cirrhosis and hepatocellular carcinoma. NAFLD is regarded as a hepatic manifestation of metabolic syndrome and incidence has been increasing worldwide in line with the increased prevalence of obesity, type 2 diabetes, and hyperlipemia. Animal models of NAFLD/NASH give crucial information, not only in elucidating pathogenesis of NAFLD/NASH but also in examining therapeutic effects of various agents. An ideal model of NAFLD/NASH should correctly reflect both hepatic histopathology and pathophysiology of human NAFLD/NASH. Animal models of NAFLD/NASH are divided into genetic, dietary, and combination models. In this paper, we review commonly used animal models of NAFLD/NASH referring to their advantages and disadvantages.     

Noninvasive biomarkers in pediatric nonalcoholic fatty liver disease

Nonalcoholic fatty liver disease (NAFLD) is the leading cause of chronic liver disease worldwide among children and adolescents. It encompasses a spectrum of disease, from its mildest form of isolated steatosis, to nonalcoholic steatohepatitis (NASH) to liver fibrosis and cirrhosis, or end-stage liver disease. The early diagnosis of pediatric NAFLD is crucial in preventing disease progression and in improving outcomes. Currently, liver biopsy is the gold standard for diagnosing NAFLD. However, given its invasive nature, there has been significant interest in developing noninvasive methods that can be used as accurate alternatives. Here, we review noninvasive biomarkers in pediatric NAFLD, focusing primarily on the diagnostic accuracy of various biomarkers as measured by their area under the receiver operating characteristic, sensitivity, and specificity. We examine two major approaches to noninvasive biomarkers in children with NAFLD. First, the biological approach that quantifies serological biomarkers. This includes the study of individual circulating molecules as biomarkers as well as the use of composite algorithms derived from combinations of biomarkers. The second is a more physical approach that examines data measured through imaging techniques as noninvasive biomarkers for pediatric NAFLD. Each of these approaches was applied to children with NAFLD, NASH, and NAFLD with fibrosis. Finally, we suggest possible areas for future research based on current gaps in knowledge.     

非酒精性脂肪性肝病的病理特点分析

目的 总结10年来福建省漳州地区肝穿刺活检标本中非酒精性脂肪性肝病(NAFLD)的发病率及病理特征.方法 收集65例NAFLD患者的相关资料,肝组织标本常规行苏木精-伊红(HE)、Masson及网状纤维染色,观察其病理特点,并进行分级.结果 在所有3 000例行肝活检的病例中,出现肝细胞脂肪变的病例约占12.7%,而NAFLD的发生率约6.1%.肝组织病理学特征主要为肝细胞大泡性或以大泡为主的脂肪变性,邻近肝细胞不同程度肿胀乃至气球样变,病变越重,肝细胞气球样变越显著,典型Mallory小体未见;纤维化主要位于窦周及中央静脉管壁,脂肪性肝硬化时脂变的肝细胞及小叶内炎细胞均减少.结论 NAFLD是近年来较常见的慢性肝病,肝活检组织学评价是确诊的金标准.     

Noninvasive evaluation of nonalcoholic fatty liver disease: Current evidence and practice

With the increasing number of individuals with diabetes and obesity,nonalcoholic fatty liver disease(NAFLD) is becoming increasingly prevalent,affecting one-quarter of adults worldwide. The spectrum of NAFLD ranges from simple steatosis or nonalcoholic fatty liver(NAFL) to nonalcoholic steatohepatitis(NASH). NAFLD, especially NASH, may progress to fibrosis, leading to cirrhosis and hepatocellular carcinoma. NAFLD can impose a severe economic burden,and patients with NAFLD-related terminal or deteriorative liver diseases have become one of the main groups receiving liver transplantation. The increasing prevalence of NAFLD and the severe outcomes of NASH make it necessary to use effective methods to identify NAFLD. Although recognized as the gold standard, biopsy is limited by its sampling bias, poor acceptability, and severe complications, such as mortality, bleeding, and pain. Therefore, noninvasive methods are urgently needed to avoid biopsy for diagnosing NAFLD. This review discusses the current noninvasive methods for assessing NAFLD,including steatosis, NASH, and NAFLD-related fibrosis, and explores the advantages and disadvantages of measurement tools. In addition, we analyze potential noninvasive biomarkers for tracking disease processes and monitoring treatment effects, and explore effective algorithms consisting of imaging and nonimaging biomarkers for diagnosing advanced fibrosis and reducing unnecessary biopsies in clinical practice.     

Fibrosis in nonalcoholic fatty liver disease: Noninvasive assessment using computed tomography volumetry

AIM To evaluate the diagnostic performance of computed tomography(CT) volumetry for discriminating the fibrosis stage in patients with nonalcoholic fatty liver disease(NAFLD).METHODS A total of 38 NAFLD patients were enrolled. On the basis of CT imaging, the volumes of total, left lateral segment(LLS), left medial segment, caudate lobe, and right lobe(RL) of the liver were calculated with a dedicated liver application. The relationship between the volume percentage of each area and fibrosis stage was analyzed using Spearman's rank correlation coefficient. A receiver operating characteristic(ROC) curve analysis was performed to determine the accuracy of CT volumetry for discriminating fibrosis stage.RESULTS The volume percentages of the caudate lobe and the LLS significantly increased with the fibrosis stage(r = 0.815, P 0.001; and r = 0.465, P = 0.003, respectively). Contrarily, the volume percentage of the RL significantly decreased with fibrosis stage(r =-0.563, P 0.001). The volume percentage of the caudate lobe had the best diagnostic accuracy for staging fibrosis, and the area under the ROC curve values for discriminating fibrosis stage were as follows: ≥ F1, 0.896; ≥ F2, 0.929; ≥ F3, 0.955; and ≥ F4, 0.923. The best cut-off for advanced fibrosis(F3-F4) was 4.789%, 85.7% sensitivity and 94.1% specificity.CONCLUSION The volume percentage of the caudate lobe calculated by CT volumetry is a useful diagnostic parameter for staging fibrosis in NAFLD patients.     

86例非酒精性脂肪肝的临床与病理分析

目的 研究非酒精性脂肪肝的临床与病理特征。方法 对86例非酒精性脂肪肝的临床及病理资料进行回顾性分析。结果 在86例非酒精性脂肪肝患者中，肥胖性脂肪肝为42例（48．8％），糖尿病性脂肪肝为11例（12．8％），高脂血症性脂肪肝为27例（31．4％），慢性丙型肝炎伴脂肪肝为6例（70％）；73．3％的患者有非特异性症状；高胆固醇血症者22例（25．6％），高甘油三酯血症者27例（31．4％）；丙氨酸转氨酶升高4例，天冬氨酸转氨酶升高3例；肝活检前B超诊断脂肪肝63例（73．3％）、B超不支持脂肪肝23例（26．7％）；病理检查，Ⅰ级肝细胞脂肪变26例（30．2％），Ⅱ级肝细胞脂肪变41例（47．7％），Ⅲ级肝细胞脂肪变19例（22．1％）。结论 非酒精性脂肪肝主要由肥胖、高脂血症和糖尿病引起，B超是诊断脂肪肝的主要方法，选择性的肝活检有助于进一步明确诊断。     

Association of nonalcoholic fatty liver disease and liver cancer

AIM:To investigate the association between nonalcoholic fatty liver disease(NAFLD) and liver cancer,and NAFLD prevalence in different liver tumors.METHODS:This is a retrospective study of the clinical,laboratory and histological data of 120 patients diagnosed with primary or secondary hepatic neoplasms and treated at a tertiary center where they underwent hepatic resection and/or liver transplantation,with subsequent evaluation of the explant or liver biopsy.The following criteria were used to exclude patients from the study:a history of alcohol abuse,hepatitis B or C infection,no tumor detected in the liver tissue examined by histological analysis,and the presence of chronic autoimmune hepatitis,hemochromatosis,Wilson’s disease,or hepatoblastoma.The occurrence of NAFLD and the association with its known risk factors were studied.The risk factors considered were diabetes mellitus,impaired glucose tolerance,impaired fasting glucose,body mass index,dyslipidemia,and arterial hypertension.Presence of reticulin fibers in the hepatic neoplasms was assessed by histological analysis using slide-mounted specimens stained with either hematoxylin and eosin or Masson’s trichrome and silver impregnation.Analysis of tumor-free liver parenchyma was carried out to determine the association between NAFLD and its histological grade.RESULTS:No difference was found in the association of NAFLD with the general population(34.2% and 30.0% respectively,95%CI:25.8-43.4).Evaluation by cancer type showed that NAFLD was more prevalent in patients with liver metastasis of colorectal cancer than in patients with hepatocellular carcinoma and intrahepatic cholangiocarcinoma(OR = 3.99,95%CI:1.78-8.94,P < 0.001 vs OR = 0.60,95%CI:0.18-2.01,P = 0.406 and OR = 0.70,95%CI:0.18-2.80,P = 0.613,respectively).There was a higher prevalence of liver fibrosis in patients with hepatocellular carcinoma(OR = 3.50,95%CI:1.06-11.57,P = 0.032).Evaluation of the relationship between the presence of NAFLD,nonalcoholic steatohepatitis,and liver fibrosis,and their risk factors,showed no significant statistical association for any of the tumors studied.CONCLUSION:NAFLD is more common in patients with liver metastases caused by colorectal cancer.     

Cytokeratin-18 fragments and biomarkers of the metabolic syndrome in nonalcoholic steatohepatitis

Nonalcoholic fatty liver disease （NAFLD） remains a leading cause of chronic liver disease. In the context of NAFLD, the presence of nonalcoholic steatohepatitis （NASH） portends an adverse prognosis with greater risk of liver fibrosis and cirrhosis. Although liver biopsy is the keystone of patient management in NAFLD, it is also increasingly clear that such evaluation has its limitations. The availability of biochemical markers of NAFLD and NASH has tremendous potential to radically alter management strategies for these conditions, as well as to monitor disease activity. Our article provides an overview of biomarker discovery and selection in the setting of NAFLD and highlights future directions in the field.     

Histopathology of nonalcoholic fatty liver disease/nonalcoholic steatohepatitis

Nonalcoholic fatty liver disease (NAFLD), a hepatic manifestation of metabolic syndrome, is the most common chronic liver disease, and the prevalence is rapidly increasing worldwide. Nonalcoholic steatohepatitis (NASH), the severe form of NAFLD, can progress to liver cirrhosis and hepatocellular carcinoma (HCC). Although noninvasive clinical scores and image-based diagnosis for NAFLD have improved, histopathological evaluation of biopsy specimens remains the gold standard for diagnosing NAFLD/NASH. Steatosis, lobular inflammation, and hepatocellular ballooning are all necessary components for the diagnosis of NASH; fibrosis is also typically observed. Other histopathological abnormalities commonly observed in NASH include hepatocellular glycogenated nuclei, lipogranulomas, and acidophil bodies. The characteristics of pediatric NAFLD/NASH differ from adult NAFLD/NASH. Specifically, steatosis and portal inflammation are more severe in pediatric NAFLD, while intralobular inflammation and perisinusoidal fibrosis are milder. Although interobserver agreement for evaluating the extent of steatosis and fibrosis is high, agreement is low for intralobular and portal inflammation. A recently reported histological variant of HCC, steatohepatitic HCC (SH-HCC), shows features that resemble non-neoplastic steatohepatitis, and is thought to be strongly associated with underlying NASH. In this report, we review the histopathological features of NAFLD/NASH.     

Liver transplantation for nonalcoholic fatty liver disease:New challenges and new opportunities

Nonalcoholic fatty liver disease(NAFLD)is becoming rapidly one of the most common indications for orthotopic liver transplantation in the world.Development of graft steatosis is a significant problem during the posttransplant course,which may happen as a recurrence of pre-existing disease or de novo NAFLD.There are different risk factors that might play a role in development of graft steatosis including post-transplant metabolic syndrome,immune-suppressive medications,genetics and others.There are few studies that assessed the effects of NAFLD on graft and patient survival;most of them were limited by the duration of follow up or by the number of patients.With this review article we will try to shed light on post-liver transplantation NAFLD,significance of the disease,how it develops,risk factors,clinical course and treatment options.     

Promising diagnostic biomarkers of nonalcoholic fatty liver disease and nonalcoholic steatohepatitis: From clinical proteomics to microbiome

Fatty liver has been present in the lives of patients and physicians for almost two centuries. Vast knowledge has been generated regarding its etiology and consequences, although a long path seeking novel and innovative diagnostic biomarkers for nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH) is still envisioned. On the one hand, proteomics and lipidomics have emerged as potential noninvasive resources for NAFLD diagnosis. In contrast, metabolomics has been able to distinguish between NAFLD and NASH, even detecting degrees of fibrosis. On the other hand, genetic and epigenetic markers have been useful in monitoring disease progression, eventually functioning as target therapies. Other markers involved in immune dysregulation, oxidative stress, and inflammation are involved in the instauration and evolution of the disease. Finally, the fascinating gut microbiome is significantly involved in NAFLD and NASH. This review presents state-of-the-art biomarkers related to NAFLD and NASH and new promises that could eventually be positioned as diagnostic resources for this disease. As is evident, despite great advances in studying these biomarkers, there is still a long path before they translate into clinical benefits.     

Effects of pentoxifylline on nonalcoholic fatty liver disease: A meta-analysis

AIM:To evaluate the effects of pentoxifylline therapy in patients with nonalcoholic fatty liver disease(NAFLD).METHODS:We searched PubMed,Medline,Google Scholar,Embase,Web of Science,the Cochrane Library and the Chinese Biomedicine Database for all relevant controlled trials of pentoxifylline in patients with NAFLD from 1997 to July 2013.Five studies(3 randomized,double-blind,placebo-controlled trials and 2 prospective cohort studies with concurrent controls)were included in this meta-analysis.Statistical analysis was performed using RevMan 5.0 software.RESULTS:Five randomized trials of 147 patients with NAFLD/nonalcoholic steatohepatitis(NASH)were included.The results showed that compared to placebo,pentoxifylline therapy resulted in a signi?cant decrease in body weight(P=0.04),alanine aminotransferase (P<0.00001),aspartate transaminase(P=0.0006),glucose(P=0.0008)and tumor necrosis factor-α(P=0.007),but did not significantly affect body mass index(P=0.28),total cholesterol(P=0.80),triglyceride(P=0.98),alkaline phosphatase(P=0.29),γ-glutamyl transferase(P=0.39)and interleukin-6(P=0.38).With regard to histological changes,pentoxifylline only reduced the NAFLD activity score(P<0.00001)and improved lobular inflammation(P<0.0001).Improvements in steatosis grade(P=0.11),ballooning(P=0.10)and fibrosis(P=0.50)were not obvious.CONCLUSION:Pentoxifylline therapy results in weight loss,improved liver function and histological changes in patients with NAFLD/NASH.Therefore,pentoxifylline may be a new treatment option for NAFLD.     

臭氧对高脂喂养诱导新西兰兔非酒精性脂肪性肝炎的防治作用

目的:研究臭氧对非酒精性脂肪性肝病(nonalcoholic fatty liver disease,NAFLD)炎症水平的防治作用及安全性评估.方法:24只♂新西兰兔分为3组:模型组10只,臭氧组4只,普拉固加阿司匹林组10只.实验时间12wk,每周称量1次体质量.超声检测第2周、第5周、第8周及第12周的颈总动脉和腹主动脉内膜-中层厚度(intimal-medial thickness,IMT).病理观察颈总动脉、主动脉、肝、心及肾脏病理变化.检测血清生化、血脂、肌酐和尿酸水平.ELISA测定血清8-OHdG、TRX、4-HNE、8-iso-PGF2a、LEP、ADPN、FFA、ET、IL-6、TNF-α、MDA、MCP-1、hs-CRP、NOS、NO、GSH、reduced glutathione、GSH-Px的含量.结果:实验动物超声检测内膜增厚,提示模型成功,臭氧组体质量明显轻于其他组.各组主动脉内膜下脂质沉积斑块大小有统计学意义(P=0.037,P<0.05),而脂质沉积厚度各组无统计学意义.臭氧组肝脏气球样变性程度明显轻于模型组和普拉固加阿司匹林组(P=0.04,P<0.05),但肝脂肪变性程度3组间无统计学意义.病理心脏脂质沉积及肾脏肾小管上皮细胞空泡变性程度均无统计学意义.各组血清生化指标均无统计学差异(P>0.05).氧化应激、脂质过氧化、炎症水平相关等18个血清细胞因子检测,与模型组相比,臭氧组除了ET、8-OHdG、GSH、GSH-Px及FFA无统计学意义,其余13个因子均有统计学意义(P<0.05),其中LEP、ADPN、IL-6、TNF-α、MCP-1、hs-CRP、NOS、TRX、MDA、4-HNE、8-iso-PGF2a水平增高,NO、reduced glutathione水平降低,但普拉固组加阿司匹林组上述所有因子与模型组间均无统计学意义.结论:臭氧减轻非酒精性脂肪性肝炎(nonalcoholic steatohepatitis,NASH)炎症的同时可改善颈总动脉、腹主动脉粥样硬化斑块面积的趋势,未发现臭氧作为抗氧剂对肝脏、肾脏和心脏造成的病理可见损伤.臭氧全面提高了血清中众多涉及氧化应激、脂质过氧化、炎症细胞因子水平,其作用机制值得进一步探讨.关键词:非酒精性脂肪性肝病;非酒精性脂肪性肝炎;动脉粥样硬化;臭氧     

Effects of probiotics on nonalcoholic fatty liver disease:A meta-analysis

AIM:To investigate the relationship between the gutliver axis and nonalcoholic fatty liver disease(NAFLD),we performed a meta-analysis to evaluate the effects of probiotic therapy in NAFLD.METHODS:We searched PubMed,Medline,Embase,Web of Science,the Cochrane Library and Chinese Biomedicine Database for all relevant randomized controlled trials on probiotics in patients with NAFLD/nonalcoholic steatohepatitis(NASH).A statistical analysis was performed using RevMan 5.0 software.RESULTS:Four randomized trials involving 134 NAFLD/NASH patients were included.The results showed that probiotic therapy signifcantly decreased alanine aminotransferase(ALT),aspartate transaminase(AST),total-cholesterol(T-chol),high density lipoprotein(HDL),tumor necrosis factor(TNF)-αand homeostasis model assessment of insulin resistance(HOMAIR)[ALT:weighted mean difference(WMD)-23.71,95%CI:-33.46--13.95,P<0.00001;AST:WMD-19.77,95%CI:-32.55--7.00,P=0.002;T-chol:WMD-0.28,95%CI:-0.55--0.01,P=0.04;HDL:WMD-0.09,95%CI:-0.16-0.01,P=0.03;TNF-α:WMD-0.32,95%CI:-0.48--0.17,P<0.0001;HOMA-IR:WMD-0.46,95%CI:-0.73--0.19,P=0.0008].However,the use of probiotics was not associated with changes in body mass index(BMI),glucose(GLU)and low density lipoprotein(LDL)(BMI:WMD 0.05,95%CI:-0.18-0.29,P=0.64;GLU:WMD 0.05,95%CI:-0.25-0.35,P=0.76;LDL:WMD-0.38,95%CI:-0.78-0.02,P=0.06).CONCLUSION:Probiotic therapies can reduce liver aminotransferases,total-cholesterol,TNF-αand improve insulin resistance in NAFLD patients.Modulation of the gut microbiota represents a new treatment for NAFLD.     

The characteristics and natural history of Japanese patients with nonalcoholic fatty liver disease.

AIMS: The aim of our study was to elucidate the characteristics and natural history of Japanese nonalcoholic fatty liver disease (NAFLD). PATIENTS AND METHODS: Two hundred and forty-seven patients were diagnosed as having biopsy-proven NAFLD at Tokyo Women's Medical University or an affiliated hospital from 1990 to June 2004. Biopsies were scored for the severity of steatosis, necro-inflammation, and fibrosis according to modified Brunt criteria. We assessed the clinicopathological features and natural history of NAFLD in patients stratified by the stage of their fibrosis. Univariate and multivariate logistic analyses were performed, and the diagnostic ability was assessed by the area under the receiver operating characteristic curve. RESULTS: Clinicopathological features: The median age of the patients was 53 years, with a range from 10 to 89 years. There were 130 males and 117 females. Histologically, 46 patients were classified as F3 (bridging fibrosis), and 43 patients had F4 (cirrhosis). Females and older patients were more common in the F3-4 patients. Most of the F3-4 patients showed mild elevation of transaminases with significant deterioration of liver function tests compared with F0-2 patients. Ten patients were simultaneously diagnosed as having cirrhotic NASH and hepatocellular carcinoma (HCC). Natural history: During follow-up (median 44 months) of the F3-4 patients, 10 patients developed liver-related morbidity and five patients developed HCC. In the F3-4 patients, the 5-year cumulative incidence of HCC was 20%. Eight patients died (two of liver failure, four of HCC and two of other carcinomas). Serum markers for detecting F3-4: Serum hyaluronic acid levels can accurately evaluate NAFLD patients with F3-4. CONCLUSIONS: The most important consequence of NAFLD patients with advanced fibrosis was HCC. Regular screening for this complication is extremely important.