Effect of Vitamin E on Serum Aminotransferase and Thioredoxin Levels in Patients with Viral Hepatitis C

Objectives: Oxidative stress induces cellular responses such as cell death, gene activation and cell proliferation, in the liver. Vitamin E (Vit. E) has been found to protect the liver against oxidative stress in animal experiments. Thioredoxin (TRX) is a stress inducible, multifunctional protein, secreted during oxidative stress. This study evaluated effects of Vit. E on serum TRX and aminotransferase levels in hepatitis C virus (HCV) patients, partly non-responsive to initial interferon (IFN), with higher than average level of serum alanine aminotransferase (ALT) after receiving anti-inflammatory drug treatment. Methods: Seventeen HCV patients (male=3; female=14) of age 62±7.65 years receiving anti-inflammatory drug therapy, at least 6 months prior to Vit. E administration, were given d-α´-tocopherol 500?mg/day, orally, for a period of 3 months. ALT, aspartate aminotransferase (AST), TRX and Vit. E were measured at 0, 1, 2 and 3 months and 1 month after end of treatment. As controls, the same patients biochemical data, 3 months from the start of therapy were used. Patients were divided into three categories: total patients “T”, low ALT group “L” (ALT<70?IU/l) and high ALT group “H”(ALT>70?IU/l), respectively.Results: The ALT level was lowered, significantly in group H, in the 1st, 2nd, 3rd and 1-month post therapy, compared to the initial value. But group L showed little or no change in ALT. Post Vit. E therapy, in groups T and H, the TRX level was elevated but remained below initial levels, whereas in group L, TRX level remained significantly lower than the pretreatment value. Groups T and L, showed significant reduction (p<0.05) in serum TRX levels in the 2nd and 3rd month. Group H showed a tendency towards TRX reduction, but not significantly. Serum Vit. E levels increased significantly (p<0.0001) from the 1st to 3rd month in all three T, H and L groups. Conclusion: Oxidative stress induced liver damage is reduced by Vit. E in patients with viral hepatitis C, particularly those with initial ALT levels >70?IU/l. Vit. E treatment causes reduction of oxidative stress markers as TRX and ALT in sera. Therefore, Vit. E can act as a supportive therapy to combat liver damage caused by oxidative stress, in such patients with continuously high levels of ALT even after anti-viral and anti-inflammatory drug therapy.     

Age as a Predictor of Significant Fibrosis Features in HBeAg-Negative Chronic Hepatitis B Virus Infection with Persistently Normal Alanine Aminotransferase

Background

Although alanine aminotransferase (ALT) levels reflect the degree of liver damage, not all patients with chronic hepatitis B virus (HBV) infection exhibit persistently elevated ALT levels. In the present study, we aimed to comprehensively evaluate the characteristics of histological abnormalities in a large population of Chinese patients with chronic HBV and persistently normal ALT levels.

Methods

In total, 2303 consecutive patients who underwent liver biopsy were screened. Of these patients, 273 were categorized as having persistently normal ALT levels (PNALT), whereas 618 were categorized as having persistently or intermittently elevated ALT levels (PIALT). All these patients had at least three ALT values recorded in the year prior to the baseline liver biopsy.

Results

Significant necroinflammation was observed in 9.7% (11/113) patients with PNALT, 23.3% (42/180) patients with PIALT (ALT 1–2× upper limit of normal [ULN]), and 27.8% (42/151) patients with PIALT (ALT > 2× ULN), whereas significant fibrosis was observed in 8.8% (10/113) patients with PNALT, 27.8% (42/151) patients with PIALT (ALT 1–2× ULN), and 21.2% (32/151) patients with PIALT (ALT > 2× ULN). Multiple logistic regression analysis indicated that age parameters were associated with significant histological abnormalities in patients with PNALT. The area under the curve showed that age was associated with significant fibrosis characteristics in patients with hepatitis B extracellular antigen (HBeAg)-negative PNALT.

Conclusion

Significant histological abnormalities are not often observed in Chinese patients with PNALT. Interestingly, age appears to be a predictor of significant fibrosis in patients with HBeAg-negative PNALT.     

Short and Long-Term Effects of Telaprevir on Kidney Function in Patients with Hepatitis C Virus Infection: A Retrospective Cohort Study

Background

Recent reports suggest that telaprevir, a protease inhibitor used to treat hepatitis C infection, is associated with decline in kidney function during therapy, particularly in patients with baseline renal impairment.

Methods

Patients treated with telaprevir in a single healthcare network were retrospectively reviewed. Kidney function was determined at baseline, during therapy, and twelve weeks and twelve months after telaprevir discontinuation. Significant creatinine rise during therapy was defined as an increase in serum creatinine ≥ 0.3mg/dL from baseline during treatment with telaprevir.

Results

Between July 2011 to January 2013,seventy-eight patients began treatment. The majority completed the prescribed twelve weeks of telaprevir therapy; 32% discontinued due to side effects. The average rise in serum creatinine during therapy was 0.22mg/dL (standard deviation 0.22mg/dL). Thirty-one percent experienced a significant creatinine rise during therapy. Decline in estimated glomerular filtration rate (eGFR) was lower in those with baseline eGFR < 90 mL/min/1.73m² compared to the group with baseline eGFR ≥ 90 mL/min/1.73m² (12 vs. 18 mL/min/1.73m², P = 0.047). Serum creatinine fully normalized by twelve weeks after cessation of telaprevir in 83% of patients, however experiencing a significant creatinine rise during telaprevir use was associated with a 6.6mL/min/1.73m² decrease in estimated glomerular filtration rate at twelve months in an adjusted model.

Conclusions

Decline in kidney function during therapy with telaprevir is common and is not associated with baseline eGFR < 90mL/min/1.73m² as previously reported.     

Plasma Sphingolipids as Potential Indicators of Hepatic Necroinflammation in Patients with Chronic Hepatitis C and Normal Alanine Aminotransferase Level

Accurate estimation of hepatic necroinflammation caused by chronic hepatitis C (CHC) is crucial for prediction of prognosis and design of therapeutic strategy, which is particularly true for CHC patients with normal alanine aminotransferase (ALT) level. Recent studies have shown that sphingolipids have a close relationship with hepatitis C virus infection. The present study aimed to identify plasma sphingolipids related to hepatic necroinflammation. We included 120 treatment-naïve CHC patients and 64/120 had normal ALT levels (<40 U/L). CHC patients who underwent liver biopsies were subjected to Scheuer scoring analysis for scope of hepatic inflammation. Plasma sphingolipids were detected by high-performance liquid chromatography tandem mass spectrometry. Our results showed 44 plasma sphingolipids were detected altogether. Of all detected sphingolipids, hexosylceramide (HexCer) (d18∶1/22∶0) and HexCer (d18∶1/24∶0) showed a significant difference among G0/G1, G2, and G3/G4 (P<0.05). For identifying hepatic necroinflammation (G≥2), after adjusting other factors, the odds ratio (OR) of HexCer (d18∶1/22∶0) reached 1.01 (95% confidence interval [CI]: 1.00–1.02). Furthermore, the area under the curve (AUC) of HexCer (d18∶1/22∶0) was 0.7 (P = 0.01) and approached that of ALT (AUC = 0.78). However, in CHC patients with normal ALT, HexCer (d18∶1/22∶0) was an independent factor (OR: 1.02, 95% CI: 1.01–1.03) to identify the hepatic necroinflammation (G≥2). HexCer (d18∶1/22∶0) not only showed the largest AUC (0.78, P = 0.001), but also exhibited the highest specificity of all indicators. These results indicate that plasma HexCer (d18∶1/22∶0) is a potential indicator to distinguish hepatic necroinflammation in CHC patients. For CHC with normal ALT, the ability of HexCer (d18∶1/22∶0) to distinguish hepatic necroinflammation might be superior to conventional serum indicators.     

Altered Functionality of Anti-Bacterial Antibodies in Patients with Chronic Hepatitis C Virus Infection

Background

Using comparative glycoproteomics, we have previously identified a glycoprotein that is altered in both amount and glycosylation as a function of liver cirrhosis. The altered glycoprotein is an agalactosylated (G0) immunoglobulin G molecule (IgG) that recognizes the heterophilic alpha-gal epitope. Since the alpha gal epitope is found on gut enterobacteria, it has been hypothesized that anti-gal antibodies are generated as a result of increased bacterial exposure in patients with liver disease.

Methods

The N-linked glycosylation of anti-gal IgG molecules from patients with fibrosis and cirrhosis was determined and the effector function of anti-bacterial antibodies from over 100 patients examined. In addition, markers of microbial exposure were determined.

Results

Surprisingly, the subset of agalactosylated anti-gal antibodies described here, was impaired in their ability to mediate complement mediated lysis and inhibited the complement-mediated destruction of common gut bacteria. In an analysis of serum from more than 100 patients with liver disease, we have shown that those with increased levels of this modified anti-gal antibody had increased levels of markers of bacterial exposure.

Conclusions

Anti-gal antibodies in patients with liver cirrhosis were reduced in their ability to mediate complement mediated lysis of target cells. As bacterial infection is a major complication in patients with cirrhosis and bacterial products such as LPS are thought to play a major role in the development and progression of liver fibrosis, this finding has many clinical implications in the etiology, prognosis and treatment of liver disease.     

Effect of Killer Immunoglobulin-Like Receptors in the Response to Combined Treatment in Patients with Chronic Hepatitis C Virus Infection

Killer immunoglobulin-like receptors (KIRs) are related to the activation and inhibition of NK cells and may play an important role in the innate response against infection with viruses such as hepatitis C virus (HCV). We examined whether the different combinations of KIRs with their HLA class I ligands influenced the response to combined treatment (pegylated alpha interferon and ribavirin) of patients infected by HCV. A total of 186 consecutive patients diagnosed with chronic HCV infection were analyzed. Seventy-seven patients exhibited HCV RNA levels at 6 months posttreatment and were called nonresponders (NR), while 109 cleared viral RNA and were named sustained viral responders (SVR). Patients were typed for HLA-B, HLA-Cw, KIR genes, and HCV genotype. In our study, the frequency of the KIR2DL2 allele was significantly increased in NR (P < 0.001; odds ratio [OR] = 1.95), as was the frequency of the KIR2DL2/KIR2DL2 genotype (P < 0.005; OR = 2.52). In contrast, the frequencies of the KIR2DL3 genotype (P < 0.001) and KIR2DL3/KIR2DL3 genotype (P < 0.05; OR = 0.54) were significantly increased in the SVR. Different combinations of KIR2DL2 and KIR2DL3 alleles with their ligands were analyzed. The frequency of the KIR2DL2/KIR2DL2-HLA-C1C2 genotype was significantly increased in the NR (P < 0.01; OR = 3.15). Additionally, we found a higher frequency of the KIR2DL3/KIR2DL3-HLA-C1C1 genotype in the SVR group (P < 0.05; OR = 0.33). These results were not affected by the HCV genotype. In conclusion, patients who carried the KIR2DL2/KIR2DL2-HLA-C1C2 genotype were less prone to respond to treatment. However, the KIR2DL3/KIR2DL3-HLA-C1C1 genotype clearly correlated with a satisfactory response to treatment, defined by the clearance of HCV RNA.Hepatitis C virus (HCV) infection is a common chronic disease affecting over 170 million people worldwide (48). Around 80% of these individuals evolve to chronic infection, and 10 to 20% of patients develop cirrhosis over a 20-year period. A minority (2%) progresses to hepatocellular carcinoma annually (18). Several host factors including age, body mass index (BMI), gender, fibrosis, cirrhosis, or the absence of cirrhosis and several viral factors including viral genotype and viral load can influence the response to treatment (6, 32, 42). Pegylated alpha interferon (Peg-IFN-α) plus ribavirin (combined therapy) constitute the most effective therapy for the treatment of chronic HCV infection (13). Since this treatment carries serious side effects, it is necessary to identify those patients who can clear HCV infection in order to reduce the period of this aggressive therapy.Natural killer (NK) cells are lymphocytes that play an important role in the host defense against HCV infection (14). NK cell activity is determined by the balance of different signals received and the equilibrium between inhibitory and activating receptors (3). Some receptors are specific for human leukocyte antigen (HLA) class I molecules (4). NK cells check the surface of the surrounding cells, detect the presence of their HLA class I molecules, and then discriminate between healthy, infected, or transformed cells (10). When NK cells contact target cells, the resulting interactions of their receptors produce either activating or inhibitory signals. If the expression of HLA class I molecules on the target cell is absent or reduced, the inhibitory signal is not generated (25).Killer cell immunoglobulin-like receptors (KIRs) are members of a group of regulatory molecules expressed on NK cells and a subset of T cells (30). This family of polymorphic genes is located on chromosome 19 (19q13.4), within the leukocyte receptor complex. The leukocyte receptor complex also encodes a number of genetically and functionally related genes. KIRs with long cytoplasmic tails are inhibitors, based on the presence of immunoreceptor tyrosine-based inhibition motifs in their cytoplasmic domains. KIRs with short tails interact with adaptor molecules such as DAP-12 (DNAX activation protein), which contain immunoreceptor tyrosine-based activation motifs and transmit activating signals (24). Several inhibitory KIRs have been well defined. KIR2DL1 binds the subset of HLA-Cw molecules with lysine at position 80 of the heavy chain (HLA-C2 group). KIR2DL2 and KIR2DL3 bind the subset of HLA-Cw molecules with asparagine at position 80 (HLA-C1 group) (34).Studies that have associated KIR genotypes with diseases have identified mainly viral infections and autoimmune diseases (22, 45). The importance of NK cells in the resolution of viral infections has prompted studies that correlate KIRs and their ligands with outcomes (12). Some studies identified a relationship between KIR genotypes and outcomes with several infectious agents such as human immunodeficiency virus (27, 29), cytomegalovirus (7), hepatitis B virus (28), and HCV (21, 26).Recently, a protective association of the inhibitory receptor KIR2DL3 with HLA-C^Asn80 (HLA-C1) and its effect on the course of HCV infection were described (21). The prevalence of KIR2DL3 and its ligand HLA-C1 is increased in individuals who eliminate HCV spontaneously, in contrast to those who remain chronically infected. The protective effect of KIR2DL3/HLA-C^Asn80 was observed only among individuals who carried both homozygous genes and had received a low HCV exposure dose. Recently, we found that the frequency of HLA-Bw4^I80 ligand and the activating receptor KIR3DS1 was increased healthy in HCV carriers compared to patients who had developed hepatocellular carcinoma (26).The aim of this study was to investigate the influence of KIR genes and KIR-HLA combinations on the response to combined therapy with Peg-IFN-α-2b and ribavirin in a group of patients with HCV infection.     

丙型肝炎病毒准种在NS2区的变异状况初探

探讨HCV准种在NS2区的基因结构特征及变异状况。利用逆转录－巢式PCR从1份HCV慢性携带者的阳性血清及1份丙肝患者的血清中获得HCV NS2全长cDNA,将其克隆于T载体,各随机挑取5个阳性克隆进行序列测定,结果显示克隆到HCV NS2全长基因,所测克隆在核苷酸水平和氨基酸水平互不相同。该慢性携带者HCV NS2区序列以完整读码框架（ORF）为主,一个于HCV多聚蛋白第835位氨基酸的位置出现终止信号,而该丙型肝炎患者以NS2N端发现终止信号的序列为主,其中三个于第835位氨基酸的位置出现终止信号,一个于第887位氨基酸的位置出现终止信号,仅一个克隆的序列为完整ORF。对ORF完整的序列进行比较,发现丙型肝炎患者氨基酸变异主要集中于N端,蛋白二级结构模拟显示丙肝患者NS2与慢性携带者的优势二级结构类似,研究表明从我们选择的两种感染者的HCV NS2序列看,不同临床类型的HCV病人体内的HCV准种在NS2区存在差异,这种差异可能与病毒存在于机体的状态一定的一致性。     

The Impact of Hepatitis A Virus Infection on Hepatitis C Virus Infection: A Competitive Exclusion Hypothesis

We address the observation that, in some cases, patients infected with the hepatitis C virus (HCV) are cleared of HCV when super-infected with the hepatitis A virus (HAV). We hypothesise that this phenomenon can be explained by the competitive exclusion principle, including the action of the immune system, and show that the inclusion of the immune system explains both the elimination of one virus and the co-existence of both infections for a certain range of parameters. We discuss the potential clinical implications of our findings.     

丙型肝炎病毒准种在NS2区的变异状况初探

探讨HCV准种在NS2区的基因结构特征及变异状况.利用逆转录-巢式PCR从1份HCV慢性携带者的阳性血清及1份丙肝患者的血清中获得HCVNS2全长cDNA,将其克隆于T载体,各随机挑取5个阳性克隆进行序列测定.结果显示克隆到HCVNS2全长基因,所测克隆在核苷酸水平和氨基酸水平互不相同.该慢性携带者HCVNS2区序列以完整读码框架(ORF)为主,一个于HCV多聚蛋白第835位氨基酸的位置出现终止信号,而该丙型肝炎患者以NS2N端发现终止信号的序列为主,其中三个于第835位氨基酸的位置出现终止信号,一个于第887位氨基酸的位置出现终止信号,仅一个克隆的序列为完整ORF.对ORF完整的序列进行比较,发现丙型肝炎患者氨基酸变异主要集中于N端,蛋白二级结构模拟显示丙肝患者NS2与慢性携带者的优势二级结构类似.研究表明从我们选择的两例感染者的HCVNS2序列看,不同临床类型的HCV病人体内的HCV准种在NS2区存在差异,这种差异可能与病毒存在于机体的状态有一定的一致性.     

Iron and Proinflammatory Cytokines in Chronic Hepatitis C Virus Infection

Steatohepatitis is a common finding in chronic hepatitis C virus (HCV) infection. As in other forms of steatohepatitis, oxidative damage may play an outstanding role. However, there are conflicting results relative to the role of iron on hepatic lipogenesis. Proinflammatory cytokines up-regulate ferritin expression, probably reflecting a defensive mechanism against increased oxidative stress, capable to open haem ring and release reactive iron. On the contrary, some adipokines, such as adiponectin, are associated with low ferritin levels. The aim of this study is to analyse the relationships of the amount of liver steatosis with serum iron, transferrin and ferritin as well as with proinflammatory cytokines, such as tumour necrosis factor (TNF)-α and interleukin (IL)-6, and adiponectin levels. We included 82 HCV infected patients and assessed the amount of liver fat by histomorphometry and its relationships with serum iron, ferritin and transferrin, adiponectin and TNF-α and IL-6. Liver steatosis was observed in 67 patients out of 82; in the remaining 15 patients, no steatosis at all was found. Patients with steatosis showed significantly higher serum ferritin levels than patients without steatosis (Z?=?2.14; p?=?0.032). When patients were classified in quartiles according to the intensity of steatosis, we observed that both TNF-α (KW?=?10.6; p?=?0.014) and IL-6 (KW?=?15.2; p?=?0.002) were significantly different among the four groups. Patients with more intense steatosis (highest quartile) showed the highest TNF-α and IL-6 values. Patients with severe hepatitis had higher levels of serum iron than patients with mild to moderate hepatitis. Serum iron also showed a correlation with the proportion of fibrosis (ρ?=?0.30; p?=?0.007). Serum iron levels are related with biochemical and histological parameters derived from liver inflammation in HCV-associated liver disease. Serum ferritin is higher among those with intense steatosis and also shows a (non-significant) trend to be associated with the more severe forms of hepatitis.     

Patterns of Hepatitis C Virus RNA Levels during Acute Infection: The InC3 Study

Background

Understanding the patterns of HCV RNA levels during acute hepatitis C virus (HCV) infection provides insights into immunopathogenesis and is important for vaccine design. This study evaluated patterns of HCV RNA levels and associated factors among individuals with acute infection.

Methods

Data were from an international collaboration of nine prospective cohorts of acute HCV (InC³ Study). Participants with well-characterized acute HCV infection (detected within three months post-infection and interval between the peak and subsequent HCV RNA levels≤120 days) were categorised by a priori-defined patterns of HCV RNA levels: i) spontaneous clearance, ii) partial viral control with persistence (≥1 log IU/mL decline in HCV RNA levels following peak) and iii) viral plateau with persistence (increase or <1 log IU/mL decline in HCV RNA levels following peak). Factors associated with HCV RNA patterns were assessed using multinomial logistic regression.

Results

Among 643 individuals with acute HCV, 162 with well-characterized acute HCV were identified: spontaneous clearance (32%), partial viral control with persistence (27%), and viral plateau with persistence (41%). HCV RNA levels reached a high viraemic phase within two months following infection, with higher levels in the spontaneous clearance and partial viral control groups, compared to the viral plateau group (median: 6.0, 6.2, 5.3 log IU/mL, respectively; P=0.018). In the two groups with persistence, Interferon lambda 3 (IFNL3) CC genotype was independently associated with partial viral control compared to viral plateau (adjusted odds ratio [AOR]: 2.75; 95%CI: 1.08, 7.02). In the two groups with viral control, female sex was independently associated with spontaneous clearance compared to partial viral control (AOR: 2.86; 95%CI: 1.04, 7.83).

Conclusions

Among individuals with acute HCV, a spectrum of HCV RNA patterns is evident. IFNL3 CC genotype is associated with initial viral control, while female sex is associated with ultimate spontaneous clearance.     

Chronic Hepatitis C Virus Infection Is Associated with the Development of Rheumatoid Arthritis: A Nationwide Population-Based Study in Taiwan

ObjectiveThe association between chronic hepatitis virus infection and rheumatoid arthritis (RA) remains debatable. This nationwide population-based cohort study assessed the risk of RA among patients with a chronic infection of hepatitis B and/or C virus.ResultsAfter adjusting for covariates, chronic HCV infection alone was significantly associated with an increased risk for RA (hazard ratio (HR)  = 2.03, 95% confidence interval (CI)  = 1.27–3.22). The increased risk for RA among participants with chronic HCV infection remained significant after restricting the analysis to those who were prescribed disease-modifying anti-rheumatic drugs. The corresponding HR for the overall sample was 1.89 (95% CI  = 1.15–3.11). However, HBV carriers did not appear to be at a significantly higher risk for RA.ConclusionOur data imply that chronic HCV infection is associated with RA development.     

Effect of Retreatment with Interferon Alone or Interferon plus Ribavirin on Hepatitis C Virus Quasispecies Diversification in Nonresponder Patients with Chronic Hepatitis C

Alpha interferon (IFN-alpha) treatment is effective on a long-term basis in only 15 to 25% of patients with chronic hepatitis C. The results of recent trials indicate that response rates can be significantly increased when IFN-alpha is given in combination with ribavirin. However, a large number of patients do not respond even to combination therapy. Nonresponsiveness to IFN is characterized by evolution of the hepatitis C virus (HCV) quasispecies. Little is known about the changes occurring within the HCV genomes when nonresponder patients are retreated with IFN or with IFN plus ribavirin. In the present study we have examined the genetic divergence of HCV quasispecies during unsuccessful retreatment with IFN or IFN plus ribavirin. Fifteen nonresponder patients with HCV-1 (4 patients with HCV-1a and 11 patients with HCV-1b) infection were studied while being retreated for 2 months (phase 1) with IFN-alpha (6 MU given three times a week), followed by IFN plus ribavirin or IFN alone for an additional 6 months (phase 2). HCV quasispecies diversification in the E2 hypervariable region-1 (HVR1) and in the putative NS5A IFN sensitivity determining region (ISDR) were analyzed for phase 1 and phase 2 by using the heteroduplex tracking assay and clonal frequency analysis techniques. A major finding of this study was the relatively rapid evolution of the HCV quasispecies observed in both treatment groups during the early phase 1 compared to the late phase 2 of treatment. The rate of quasispecies diversification in HVR1 was significantly higher during phase 1 versus phase 2 both in patients who received IFN plus ribavirin (P = 0.017) and in patients who received IFN alone (P = 0. 05). A trend toward higher rates of quasispecies evolution in the ISDR was also observed during phase 1 in both groups, although the results did not reach statistical significance. However, the NS5A quasispecies appeared to be rather homogeneous and stable in most nonresponder patients, suggesting the presence of a single well-fit major variant, resistant to antiviral treatment, in agreement with published data which have identified an IFN sensitivity determinant region within the NS5A. During the entire 8 months of retreatment, there was no difference in the rate of fixation of mutation between patients who received combination therapy and patients who were treated with IFN alone, suggesting that ribavirin had no major effects on the evolution of the HCV quasispecies after the initial 2 months of IFN therapy.     

A Serum MicroRNA Signature Is Associated with the Immune Control of Chronic Hepatitis B Virus Infection

Background and Aims

The virus/host interplay mediates liver pathology in chronic HBV infection. MiRNAs play a pivotal role in virus/host interactions and are detected in both serum and HBsAg-particles, but studies of their dynamics during chronic infection and antiviral therapy are missing. We studied serum miRNAs during different phases of chronic HBV infection and antiviral treatment.

Methods

MiRNAs were profiled by miRCURY-LNA-Universal-RT-miRNA-PCR (Exiqon-A/S) and qPCR-panels-I/II-739-miRNA-assays and single-RT-q-PCRs. Two cohorts of well-characterized HBsAg-carriers were studied (median follow-up 34–52 months): a) training-panel (141 sera) and HBsAg-particles (32 samples) from 61 HBsAg-carriers and b) validation-panel (136 sera) from 84 carriers.

Results

Thirty-one miRNAs were differentially expressed in inactive-carriers (IC) and chronic-hepatitis-B (CHB) with the largest difference for miR-122-5p, miR-99a-5p and miR-192-5p (liver-specific-miRNAs), over-expressed in both sera and HBsAg-particles of CHB (ANOVA/U-test p-values: <0.000001/0.000001; <0.000001/0.000003; <0.000001/0.000005, respectively) and significantly down-regulated during- and after-treatment in sustained-virological-responders (SVR). MiRNA-profiles of IC and SVR clustered in the heatmap. Liver-miRNAs were combined with miR-335, miR-126 and miR-320a (internal controls) to build a MiR-B-Index with 100% sensitivity, 83.3% and 92.5% specificity (−1.7 cut-off) in both training and validation cohorts to identify IC. MiR-B-Index (−5.72, −20.43/14.38) correlated with ALT (49, 10/2056 U/l, ρ = −0.497, p<0.001), HBV-DNA (4.58, undetectable/>8.3 Log₁₀ IU/mL, ρ = −0.732, p<0.001) and HBsAg (3.40, 0.11/5.49 Log₁₀ IU/mL, ρ = −0.883, p<0.001). At multivariate analysis HBV-DNA (p = 0.002), HBsAg (p<0.001) and infection-phase (p<0.001), but not ALT (p = 0.360) correlated with MiR-B-Index. In SVR to Peg-IFN/NUCs MiR-B-Index improved during-therapy and post-treatment reaching IC-like values (5.32, −1.65/10.91 vs 6.68, 0.54/9.53, p = 0.324) beckoning sustained HBV-immune-control earlier than HBsAg-decline.

Conclusions

Serum miRNA profile change dynamically during the different phases of chronic HBV infection. We identified a miRNA signature associated with both natural-occurring and therapy-induced immune control of HBV infection. The MiR-B-Index might be a useful biomarker for the early identification of the sustained switch from CHB to inactive HBV-infection in patients treated with antivirals.     

Normocaloric Low Cholesterol Diet Modulates Th17/Treg Balance in Patients with Chronic Hepatitis C Virus Infection

Hepatitis C virus (HCV) infection is associated with hepatic and extrahepatic manifestations, including immunological disorders. Chronic Hepatitis C (CHC) is often characterized by cholesterol and lipid metabolism alterations, leading to hepatic steatosis. Cholesterol metabolism, in fact, is crucial for the viral life cycle. Recent works described that a higher dietary cholesterol intake is associated with the progression of HCV-related liver disease. CHC patients have increased levels of T helper 17 (Th17)-cells, a lymphocytic population involved in the pathogenesis of liver inflammation and autoimmune hepatitis. The balance between Th17 and regulatory T (Treg) cells is crucial for chronic inflammation and autoimmunity. Th17-cell differentiation is deeply influenced by the activation LXRs, nuclear receptors modulating cholesterol homeostasis. Moreover, HCV may affect these nuclear receptors, and cholesterol metabolism, through both direct and indirect mechanisms. On these bases, we hypothesized that modulation of cholesterol levels through Normocaloric Low Cholesterol Diet (NLCD) may represent an innovative strategy to reduce the progression of HCV infection, through the modulation of peripheral Th17/Treg balance. To this end, we performed a pilot study to investigate whether a Normocaloric Low Cholesterol Diet may be able to modulate Th17/Treg balance in patients affected by chronic HCV infection. After 30 days of NLCD CHC patients showed a significant reduction in Th17 cells frequency, which correlated with strong reduction of IL-17 and IL-22 serum levels. At the same time, we appreciated an increase in the percentage of Treg cells, thus improving Treg/Th17balance. Moreover, we observed an increased expression of LXRs and their target genes: SREBP-1c and ABCA-1. In conclusion, NLCD finely regulates Th17/Treg balance, improving immune system response in CHC patients. This study could pave the way for new treatments of CHC patients, suggesting that change in lifestyle could support the management of these patients, promoting well-being and possibly hindering disease progression.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT02038387","term_id":"NCT02038387"}}NCT02038387     

Differential Effect of Viral Hepatitis Infection on Mortality among Korean Maintenance Dialysis Patients: A Prospective Multicenter Cohort Study

The role of infection with hepatitis B virus (HBV) and hepatitis C virus (HCV) in terms of survival among dialysis patients remains incompletely understood. In the present multicenter prospective cohort study, we investigated the prevalences of HBV and HCV infection among 3,321 patients receiving maintenance dialysis in Korea, and assessed the impacts of these infections on survival. All included patients underwent hepatitis B antigen (HBsAg) and HCV antibody (Ab) testing, which revealed that 236 patients (7.1%) were HBsAg-positive, and 123 patients (3.7%) were HCV Ab-positive. HBsAg-positive and HCV Ab-positive patients were matched to hepatitis virus-negative patients using a propensity score at a ratio of 1:2. The prevalences of HBV and HCV infection did not significantly differ according to dialysis modality. Linear-by-linear association analysis revealed that hepatitis B prevalence significantly increased with increasing dialysis vintage (p = 0.001), and hepatitis C prevalence tended to be higher with increasing dialysis vintage (p = 0.074). We compared the survival of HBsAg-positive and HCV Ab-positive patients to that of hepatitis virus-negative patients. After propensity score matching, cumulative survival did not differ between HBsAg-positive and HBsAg-negative patients (p = 0.37), while HCV Ab-positive patients showed significantly lower survival than HCV Ab-negative patients (p = 0.03). The main conclusions of the present study are that HBV infection prevalence increased with longer dialysis vintage, and that both HBV and HCV infections were most prevalent among patients with the longest dialysis vintage. Additionally, HCV infection among maintenance dialysis patients is associated with an increased risk of mortality.     

Serum Osteopontin Predicts Degree of Hepatic Fibrosis and Serves as a Biomarker in Patients with Hepatitis C Virus Infection

Background & Aims

Osteopontin (OPN) is a matricellular protein that upregulates during pathogenesis of hepatic fibrosis. The present study was aimed to evaluate whether serum OPN could be used as a biomarker to assess the degree of hepatic fibrosis in patients with hepatitis C virus (HCV) infection.

Methods

Needle biopsy was performed on HCV patients and scored as zero fibrosis (F0), mild fibrosis (F1), moderate fibrosis (F2), severe fibrosis (F3) and liver cirrhosis (F4) based on Masson’s trichrome and α-smooth muscle actin (α-SMA) staining. Serum OPN levels were measured using ELISA and correlated with the degree of fibrosis. Furthermore, the OPN values were correlated and evaluated with platelets count, serum hyaluronic acid (HA), and collagen type IV and subjected to receiver operating characteristic (ROC) curve analysis.

Results

Serum OPN levels were remarkably increased from F0 through F4 in a progressive manner and the differences were significant (P < 0.001) between each group. The data were highly correlated with the degree of hepatic fibrosis. The ROC curve analysis depicted that serum OPN is an independent risk factor and an excellent biomarker and a prognostic index in HCV patients.

Conclusions

The results of the present study indicate that serum OPN levels reflect the degree of hepatic fibrosis and could be used as a biomarker to assess the stage of fibrosis in HCV patients which would help to reduce the number of liver biopsies. Furthermore, serum OPN serves as a prognostic index towards the progression of hepatic fibrosis to cirrhosis and hepatocellular carcinoma.     

Angiopoietin-2 Serum Levels Improve Noninvasive Fibrosis Staging in Chronic Hepatitis C: A Fibrogenic-Angiogenic Link

Aims

Accurate liver fibrosis staging is crucial for the management of chronic hepatitis C (CHC). The invasiveness and cost burden of liver biopsy have driven the search for new noninvasive biomarkers of fibrosis. Based on the link between serum angiopoietin-1 and 2 levels and CHC progression, we aimed to determine the value of these angiogenic factors as noninvasive biomarkers of liver fibrosis.

Methods

Serum levels of angiopoietin-1 and -2 were measured by ELISA in 108 CHC patients who underwent pretreatment liver biopsy. The correlation between angiopoietins and clinical and demographic variables with liver fibrosis was analyzed by univariate regression. Significant factors were then subjected to multivariate analysis, from which we constructed a novel noninvasive liver fibrosis index (AngioScore), whose performance was validated in an independent series of 71 CHC patients. The accuracy of this model was compared with other documented fibrosis algorithms by De Long test.

Results

Angiopoietins correlated significantly with hepatic fibrosis; however, only angiopoietin-2 was retained in the final model, which also included age, platelets, AST, INR, and GGT. The model was validated and behaved considerably better than other fibrosis indices in discriminating all, significant, moderate and severe liver fibrosis (0.886, 0.920, 0.923). Using clinically relevant cutoffs, we classified CHC patients by discarding significant fibrosis and diagnosing moderate and severe fibrosis with greater accuracy, sensitivity, and specificity.

Conclusions

Our novel noninvasive liver fibrosis model, based on serum angiopoietin-2 levels, outperforms other indices and should help substantially in managing CHC and monitoring long-term follow-up prognosis.