Hepatocellular Carcinoma in Patients with a Sustained Response to Anti-Hepatitis C Therapy

Hepatocellular carcinoma (HCC) is a common, life-threatening complication of longstanding infection with the hepatitis C virus (HCV), likely a consequence of the direct oncogenic activity of the virus cooperating with liver cell inflammation in transforming the liver into a mitogenic and mutagenic environment. The achievement of a sustained virological response (SVR) to interferon-based therapies has been shown to benefit the course of hepatitis C in terms of reduced rates of liver-related complications and mortality from all causes. Interestingly, while achievement of an SVR is associated with a negligible risk of developing clinical decompensation over the years, the risk of HCC is not fully abrogated following HCV clearance, but it remains the dominant complication in all SVR populations. The factors accounting for such a residual risk of HCC in SVR patients are not fully understood, yet the persistence of the subverted architecture of the liver, diabetes and alcohol abuse are likely culprits. In the end, the risk of developing an HCC in SVR patients is attenuated by 75% compared to non-responders or untreated patients, whereas responders who develop an HCC may be stratified in different categories of HCC risk by a score based on the same demographic and liver disease-based variables, such as those that predict liver cancer in viremic patients. All in all, this prevents full understanding of those factors that drive HCC risk once HCV has been eradicated. Here, we critically review current understanding of HCC in SVR patients focusing on factors that predict residual risk of HCC among these patients and providing a glimpse of the expected benefits of new anti-HCV regimens based on direct antiviral agents.     

Genetic Variants in the Apoptosis Gene BCL2L1 Improve Response to Interferon-Based Treatment of Hepatitis C Virus Genotype 3 Infection

Genetic variation upstream of the apoptosis pathway has been associated with outcome of hepatitis C virus (HCV) infection. We investigated genetic polymorphisms in the intrinsic apoptosis pathway to assess their influence on sustained virological response (SVR) to pegylated interferon-α and ribavirin (pegIFN/RBV) treatment of HCV genotypes 1 and 3 infections. We conducted a candidate gene association study in a prospective cohort of 201 chronic HCV-infected individuals undergoing treatment with pegIFN/RBV. Differences between groups were compared in logistic regression adjusted for age, HCV viral load and interleukin 28B genotypes. Four single nucleotide polymorphisms (SNPs) located in the B-cell lymphoma 2-like 1 (BCL2L1) gene were significantly associated with SVR. SVR rates were significantly higher for carriers of the beneficial rs1484994 CC genotypes. In multivariate logistic regression, the rs1484994 SNP combined CC + TC genotypes were associated with a 3.4 higher odds ratio (OR) in SVR for the HCV genotype 3 (p = 0.02). The effect estimate was similar for genotype 1, but the association did not reach statistical significance. In conclusion, anti-apoptotic SNPs in the BCL2L1 gene were predictive of SVR to pegIFN/RBV treatment in HCV genotypes 1 and 3 infected individuals. These SNPs may be used in prediction of SVR, but further studies are needed.     

全球首个抗HCV全口服新药索非布韦

丙型肝炎病毒感染极大地威胁着公众健康,目前的标准治疗方案是聚乙二醇修饰的干扰素-α(PegIFN)与广谱抗病毒药物利巴韦林(RBV)联合,然而这种疗法的持续病毒学应答相对较低,新的研究如直接作用的抗病毒药(DAAs)提供了一种新的治疗策略,以提高疗效和耐受性。索非布韦是首个全口服抗丙型肝炎病毒药物,通过抑制NS5B聚合酶来有效抑制HCV的复制,从而达到治疗效果。本文主要就其化学合成、作用机理、药代动力学特征及临床疗效和安全性做一综述。     

Differential Expression of the Host Lipid Regulators ANGPTL-3 and ANGPTL-4 in HCV Infection and Treatment

Host lipid metabolism reprogramming is essential for hepatitis C virus (HCV) infection and progression to severe liver disease. Direct-acting antivirals (DAAs) achieve a sustained virological response (SVR) in most patients, but virus eradication does not always protect against hepatocellular carcinoma (HCC). Angiopoietin-like protein-3 (ANGPTL-3) and angiopoietin-like protein-4 (ANGPTL-4) regulate the clearance of plasma lipids by inhibiting cellular lipase activity and possess emerging roles in tumourigenesis. We used ELISA and RT-qPCR to investigate ANGPTL-3 and ANGPTL-4 expression in HCV patients with characterised fibrosis throughout the natural history of hepatitis C and in long-term HCV infection in vitro, before and after DAA treatment. ANGPTL-3 was decreased in patients with advanced fibrosis compared to other disease stages, while ANGPTL-4 was progressively increased from acute infection to cirrhosis and HCC, peaking at the advanced fibrosis stage. Only ANGPTL-3 mRNA was down-regulated during early infection in vitro, although both ANGPTLs were increased later. DAA treatment did not alter ANGPTL-3 levels in advanced fibrosis/cirrhosis and in HCV infection in vitro, in contrast to ANGPTL-4. The association between ANGPTLs and fibrosis in HCV infection was underlined by an inverse correlation between the levels of ANGPTLs and serum transforming growth factor- β (TGF-β). Collectively, we demonstrate the pivotal role of advanced fibrosis in defining the expression fate of ANGPTLs in HCV infection and after treatment and propose a role for ANGPTL-3 as a contributor to post-treatment deregulation of lipid metabolism that could predispose certain individuals to HCC development.     

丙型肝炎病毒核心总抗原检测试剂的评价

目的评价国外丙型肝类病毒核心抗原检测试剂盒的质量以及窗口期检测核心总抗原(HCVcAgTrak-C)的意义。方法用该试剂检测8649份自然献血员及566份HCV可疑感染者血清的核心总抗原,并与抗体试剂(Anti-HCV)、游离抗原(HCVfreeAg)试剂、核酸(HCVRNA)试剂进行比较。结果筛出核心总抗原和抗体同时阳性血清152份,抗体阴性而总抗原阳性血清5份。利用RNA试剂检测这5份血清,结果HCVRNA阳性。利用HCVfreeAg试剂检测这9215份血清,发现2份阳性,其中1份为抗原和抗体同时阳性,RIBA检测确证这份血清产生的是核心区抗体。另1份为单独抗原阳性。结论在应用HCV抗体试剂对血液进行筛查的同时使用HCVcAg试剂检测,可以减少因窗口期造成的漏检。     

Hepatocyte Proteome Alterations Induced by Individual and Combinations of Common Free Fatty Acids

Non-alcoholic fatty liver disease is a pathology with a hard-to-detect onset and is estimated to be present in a quarter of the adult human population. To improve our understanding of the development of non-alcoholic fatty liver disease, we treated a human hepatoma cell line model, HepG2, with increasing concentrations of common fatty acids, namely myristic, palmitic and oleic acid. To reproduce more physiologically representative conditions, we also included combinations of these fatty acids and monitored the cellular response with an in-depth proteomics approach and imaging techniques. The two saturated fatty acids initially presented a similar phenotype of a dose-dependent decrease in growth rates and impaired lipid droplet formation. Detailed analysis revealed that the drop in the growth rates was due to delayed cell-cycle progression following myristic acid treatment, whereas palmitic acid led to cellular apoptosis. In contrast, oleic acid, as well as saturated fatty acid mixtures with oleic acid, led to a dose-dependent increase in lipid droplet volume without adverse impacts on cell growth. Comparing the effects of harmful single-fatty-acid treatments and the well-tolerated fatty acid mixes on the cellular proteome, we were able to differentiate between fatty-acid-specific cellular responses and likely common lipotoxic denominators.     

Tamoxifen-induced modification of serum lipoprotein phospholipids in the cockerel

The administration of tamoxifen (Tam), a nonsteroidal antiestrogen, or of a diphenyl-methane derivative of Tam that does not bind to the estrogen receptor (DPPE) of cockerels results in a marked decrease in the concentration of serum lipoprotein constitutents with an apparent alteration in phospholipid composition. To establish the nature of changes in phospholipids, the molecular species of phosphatidylcholine (PC) and sphingomyelin (Sph) were isolated and characterized. Between 9 and 18 hr following the administration of Tam or DPPE, there was a marked decrease in the proportion of molecular species of serum PC containing C16 and C18 fatty acids, but there was an increase in the proportion of molecular species containing C20 and C22 polyunsaturated fatty acids. Fatty acid analyses revealed that this change was due to an increase in arachidonic and docosaxaenoic acids at the expense of oleic and linoleic acids. These proportional changes were due to an absolute decrease in serum of PC molecular species containing palmitic and stearic acids in association with oleic and linoleic acids with very little change in the absolute concentration of molecular species containing arachidonic and docosahexaenoic acids. By contrast, the composition of Sph, which contained palmitic acid as the major fatty acid, was not altered during treatment. It is concluded that the short-term effect of Tam and DPPE on plasma phospholipids of the cockerel is due to a selective conservation of PC containing long chain polyunsaturated fatty acids.     

Comparison of catabolism rate of fatty acids to carbon dioxide in mice

The catabolism rates of a medium chain fatty acid (octanoic acid), an even‐numbered fatty acid (palmitic acid), and odd‐numbered fatty acids (pentadecanoic acid and heptadecanoic acid) in mice were compared using stable isotope (¹³C) labeled fatty acids and isotope‐ratio MS (IRMS). The catabolism rates of respective fatty acids were evaluated by the ratio of ¹³C and ¹²C in carbon dioxide expired from mice. The results show that the catabolism rate of octanoic acid is three times faster than that of palmitic acid. This result is in agreement with previous knowledge that medium chain fatty acids are easily beta‐oxidized as compared to long chain fatty acids. The catabolism rates of odd‐numbered fatty acids such as pentadecanoic acid and heptadecanoic acid were significantly lower as compared to those of even‐numbered fatty acids such as palmitic acid. This finding supports our previous report that odd‐numbered fatty acids are easily accumulated into body fat. The high accumulation of odd‐numbered fatty acids in body fat would be a direct result of their low beta‐oxidizability. Practical applications: ¹³C‐labeled fatty acids were administered to mice and the rates of ¹³CO₂ formation were compared among medium chain, even‐numbered, and odd‐numbered fatty acids using IRMS. We found that the catabolism rates of odd‐numbered fatty acids such as pentadecanoic acid and heptadecanoic acid were significantly lower in comparison to those of even‐numbered fatty acids such as palmitic acid. These findings could be valuable for the development of the lipid metabolism field.     

Solidification of unsaturated/saturated fatty acid mixtures and its relationship to chilling sensitivity in plants

Freezing points of mixtures of palmitic and linoleic, or palmitic and linolenic acids, the predominant fatty acids in plants, decrease slowly as the unsaturated fatty acid is increased to 60 mole %. Beyond this per cent the freezing point is depressed quite markedly by each addition of unsaturated fatty acid. Linoleic and linolenic acids have similar effects on the freezing points of the mixtures until about 82 mole % unsaturated fatty acid. Differences of less than 5% in the amount of unsaturated fatty acid have a marked effect on the freezing point of mixtures at the approximate composition of fatty acid in plant membrane lipids.     

2-O-Methylhonokiol Suppresses HCV Replication via TRAF6-Mediated NF-kB Activation

Hepatitis C virus (HCV) is associated with various liver diseases. Chronic HCV infection is characterized by an abnormal host immune response. Therefore, it is speculated that to suppress HCV, a well-regulated host immune response is necessary. 2-O-methylhonokiol was identified by the screening of anti-HCV compounds using Renilla luciferase assay in Huh 7.5/Con 1 genotype 1b replicon cells. Here, we investigated the mechanism by which 2-O-methylhonokiol treatment inhibits HCV replication using real-time PCR. Our data shows that treatment with 2-O-methylhonokiol activated innate immune responses via nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kB) pathway. Additionally, the immunoprecipitation result shows that treatment with 2-O-methylhonokiol augmented tumor necrosis factor receptor (TNFR)-associated factor 6 (TRAF6) by preventing p62 from binding to TRAF6, resulting in reduced autophagy caused by HCV. Finally, we reproduced our data with the conditioned media from 2-O-methylhonokiol-treated cells. These findings strongly suggest that 2-O-methylhonokiol enhances the host immune response and suppresses HCV replication via TRAF6-mediated NF-kB activation.     

Distribution of fatty acids during germination of soybean seeds

Gas chromatographic determination of the fatty acids in the seeds of soybean (Glycine max) showed mainly linoleic, oleic and palmitic acids with linoleic acid being the major component. Changes in the distribution of fatty acids were measured during germination in the cotyledons and roots. A decrease in palmitic and oleic acids was observed in the cotyledons from 6 to 12 days, while linoleic acid increased during the same period. In roots also, the major fatty acid was linoleic acid, while palmitic and linolenic acids were higher in roots in comparison with the cotyledons. During the 3–12 days of germination period, no major changes in the distribution pattern of fatty acids were observed in the roots. The possible significance of these changes is discussed.     

Lipid Composition of Garlic

Neutral, glyco- and phospholipids of garlic were resolved into their component fractions by thin layer chromatography. Neutral lipids contained considerable quantities of monoglycerides (18.5%), diglycerides (14.2%), sterols (16.3%) and triglycerides (41.5%) respectively. The phospholipid fraction was rich in phosphatidyl choline (23.5%), phosphatidyl ethanolamine (17.9%), lysophosphatidyl choline (11.8%) and lysophosphatidyl ethanolamine (8.2%). Digalactosyl diglyceride (10.1%), sterol glycoside (15.6%), cerebrosides (8.1%), acylsterol glycoside (38.6%) and monogalactosyl diglyceride (22.5%) were the major components of the glycolipids of garlic. Lauric, myristic, palmitic and linoleic acids constituted the major fatty acids of monoglycerides, diglycerides and free fatty acid fractions whereas palmitic, linoleic and linolenic acids were the major fatty acids of triglycerides. Palmitic and linoleic acids were the major fatty acids of garlic phospholipids. Except the acylsterol glycoside fraction glycolipids were rich in lauric, palmitic, linoleic and linolenic acids; palmitic acid was the only major fatty acid of acylsterol glycosides.     

Lipids of cultured hepatoma cells: IV. Effect of serum and lipid upon cellular and media neutral lipids

Minimal deviation hepatoma 7288C cells were cultured in a modified Swim's medium supplemented with decreasing levels of serum, lipid-free serum, lipid-free serum plus fatty acids, and other additives. Cellular and media neutral lipid classes were quantitated, the fatty acids of triglycerides and sterol esters analyzed, and the carbon number distribution of triglycerides determined. Cellular triglyceride biosynthesis virtually was inhibited when the medium was supplemented with bovine serum alone. This inhibition was not observed when the medium was supplemented with fetal calf serum alone or mixtures of fetal calf serum and bovine serum. Cells cultivated on medium supplemented with lipid-free serum plus palmitic or linoleic acids had much lower levels of free and esterified cholesterol. The fatty acid composition of cellular triglycerides and cholesterol esters differed dramatically from the corresponding media lipid classes. Except when linoleic acid was added to the medium, changes in the media serum and lipid levels had only marginal effects upon the fatty acid composition of cellular triglycerides and cholesterol esters. These data, in conjunction with earlier data that showed the media neutral lipid levels did not decrease during cell growth, indicate that these hepatoma cells utilize little or no serum triglycerides and cholesterol esters. Linoleic acid added to the medium dramatically reduced the level of 18∶1 acids in cellular triglycerides and cholesterol esters. Palmitic acid added to the medium did not change the fatty acid compositions significantly. Comparison of experimentally determined and calculated triglyceride carbon number percentages indicated a random distribution of fatty acids in this glyceride. The fatty acid composition of cellular triglycerides was similar to the composition of the cholesterol esters. The lack of characteristic and distinguishable compositions of these two classes that occur in most normal tissues suggests a loss of specificity in the lipid metabolism of this neoplasm at the class level.     

The Role of Chemokines in Hepatitis C Virus-Mediated Liver Disease

The hepatitis C virus (HCV) is a global health problem affecting more than 170 million people. A chronic HCV infection is associated with liver fibrosis, liver cirrhosis and hepatocellular carcinoma. To enable viral persistence, HCV has developed mechanisms to modulate both innate and adaptive immunity. The recruitment of antiviral immune cells in the liver is mainly dependent on the release of specific chemokines. Thus, the modulation of their expression could represent an efficient viral escape mechanism to hamper specific immune cell migration to the liver during the acute phase of the infection. HCV-mediated changes in hepatic immune cell chemotaxis during the chronic phase of the infection are significantly affecting antiviral immunity and tissue damage and thus influence survival of both the host and the virus. This review summarizes our current understanding of the HCV-mediated modulation of chemokine expression and of its impact on the development of liver disease. A profound knowledge of the strategies used by HCV to interfere with the host’s immune response and the pro-fibrotic and pro-carcinogenic activities of HCV is essential to be able to design effective immunotherapies against HCV and HCV-mediated liver diseases.     

Rapid separation of triacylglycerol positional isomers binding two saturated Fatty acids using octacocyl silylation column

The rapid separation of a triacylglycerol positional isomer (TAG-PI) pair was examined via high-performance liquid chromatography-atmospheric pressure chemical ionization tandem mass spectrometry using an octacocyl silylation (C28) column. A TAG-PI pair binding two palmitic acids and one fatty acid that structurally differs from palmitic acid was separated at 10°C and 15°C using acetone as the mobile phase. However, the TAG-PI pair binding two unsaturated fatty acids and one saturated fatty acid was not separated by the C28 column. The results indicate that the structures of the two palmitic acids (saturated fatty acids) and the other fatty acid at the α or β position in TAG play an important role in the separation of the TAG-PI pair, and that the structure of the fatty acid needs to be considerably different from that of the palmitic acid, specifically in terms of the chain length or the location of the double bond.     

The Emerging Role of Non-Coding RNAs in the Regulation of Virus Replication and Resultant Cellular Pathologies

Non-coding RNAs, particularly lncRNAs and miRNAs, have recently been shown to regulate different steps in viral infections and induction of immune responses against viruses. Expressions of several host and viral lncRNAs have been found to be altered during viral infection. These lncRNAs can exert antiviral function via inhibition of viral infection or stimulation of antiviral immune response. Some other lncRNAs can promote viral replication or suppress antiviral responses. The current review summarizes the interaction between ncRNAs and herpes simplex virus, cytomegalovirus, and Epstein–Barr infections. The data presented in this review helps identify viral-related regulators and proposes novel strategies for the prevention and treatment of viral infection.     

Hepatocellular Carcinoma in Chronic Viral Hepatitis: Where Do We Stand?

Hepatocellular carcinoma (HCC) is one of the major causes of cancer-related death. Although the burden of alcohol- and NASH-related HCC is growing, chronic viral hepatitis (HBV and HCV) remains a major cause of HCC development worldwide. The pathophysiology of viral-related HCC includes liver inflammation, oxidative stress, and deregulation of cell signaling pathways. HBV is particularly oncogenic because, contrary to HCV, integrates in the cell DNA and persists despite virological suppression by nucleotide analogues. Surveillance by six-month ultrasound is recommended in patients with cirrhosis and in “high-risk” patients with chronic HBV infection. Antiviral therapy reduces the risks of development and recurrence of HCC; however, patients with advanced chronic liver disease remain at risk of HCC despite virological suppression/cure and should therefore continue surveillance. Multiple scores have been developed in patients with chronic hepatitis B to predict the risk of HCC development and may be used to stratify individual patient’s risk. In patients with HCV-related liver disease who achieve sustained virological response by direct acting antivirals, there is a strong need for markers/scores to predict long-term risk of HCC. In this review, we discuss the most recent advances regarding viral-related HCC.     

The variation of triglyceride structure with fatty acid composition in pig adipose tissue

Forty-five triglyceride samples with a wide range of fatty acid compositions were selected from a large number of pig adipose tissue samples (inner and outer back fats and perenephric fat) available from nutritional experiments. These samples were subjected to stereospecific analysis to determine the changes occurring in the positional distribution of the component fatty acids. The oleic acid content of the triglycerides was taken as the standard of comparison and as this increased, the proportions of the other unsaturated fatty acids also increased in a linear manner and the concentrations of the saturated components decreased proportionately. In position 1, the palmitic acid concentration remained constant while the stearic acid concentration decreased linearly and the concentrations of the unsaturated fatty acids increased. In position 2 the stearic acid concentration remained almost constant while the palmitic acid concentration decreased linearly in response to increases in the concentrations of the unsaturated acids. The least change occurred in position 3 where there were slight decreases in the concentrations of saturated acids as the concentrations of unsaturated acids increased. The precise quantitative relationships depended on the tissue examined. Constant proportions of the available myristic and palmitoleic acids were found in all three positions and constant proportions of the available stearic and oleic acids were found in position 1. These results are discussed in relation to possible pathways of triglyceride biosynthesis in pig adipose tissues.