Interleukin-4 polymorphisms and response to combination therapy in Egyptian chronic hepatitis C patients

In hepatitis C infection, the production of inappropriate cytokines levels may contribute to viral persistence and may affect the response to antiviral therapy. We investigate the effect of IL4 C-590T and C-33T polymorphisms on the response to combination therapy with interferon and ribavirin in chronic HCV patients. These single nucleotide polymorphisms were determined by PCR-RFLP in 235 responder and 210 non-responder to combination therapy. The IL4-590 T/T and -33 T/T genotypes were associated with resistance to the therapy (p<0.001, p=0.001 respectively). Haplotypes T(-590) T(-33) and T(-590) C(-33) were associated with a higher risk in non-responder patients than the responders (p<0.001 for each) while frequency of haplotype C(-590) C(-33) (with all wild alleles) was significantly higher in responders as compared to non-responders (p<0.001). These results suggest that inheritance of the IL4 polymorphisms may be associated with resistance to combined antiviral therapy in Egyptian HCV patients.     

LecT-Hepa facilitates estimating treatment outcome during interferon therapy in chronic hepatitis C patients

Background

A combination treatment of interferon and ribavirin is the standard and the commonly used treatment for chronic hepatitis C (CHC). Developing noninvasive tests like serum indicators that can predict treatment outcome at an early stage of therapy is beneficial for individualized treatment and management of CHC. A glyco-indicator based on the glyco-alteration of serum α1-acid glycoprotein, LecT-Hepa, was discovered by glycomics technologies as a robust indicator of liver fibrosis. Here, we investigated the clinical utility of LecT-Hepa for evaluation of treatment outcome.

Results

Firstly, ninety-seven patients with CHC were used for comparison of LecT-Hepa in serum and plasma. We found no significant difference in the concentrations of LecT-Hepa in serum and plasma. And then, 213 serum specimens from 45 patients who received 48 weeks of treatment with interferon and ribavirin were followed up for 96 weeks, and were used for evaluation of the role of LecT-Hepa. We found that LecT-Hepa might reflect the change in fibrosis regression during the treatment process. Moreover, the change of LecT-Hepa at the first 12 weeks of treatment could already predict the antiviral treatment response, which was more superior to FIB-4 index and aspartate aminotransferase-to-platelet ratio index (APRI) in this study.

Conclusions

These results provide a new perspective that serum glycoprotein could be used as a joint diagnosis indicator for estimation treatment outcome of viral hepatitis at earlier stage of therapy.

Electronic supplementary material

The online version of this article (doi:10.1186/1559-0275-11-44) contains supplementary material, which is available to authorized users.     

Serum tumour necrosis factor-alpha and transforming growth factor-beta levels in chronic hepatitis C patients are immunomodulated by therapy

Our aims were: (i) to characterize serum levels of tumour necrosis factor alpha (TNF-alpha) and transforming growth factor beta (TGF-beta) in non-cirrhotics with hepatitis C; (ii) to correlate levels of theses cytokines with degree of disease at baseline; (iii) to characterize the immunomodulatory effects of therapy with response and (iv) to compare profiles of cytokines in patients treated with pegylated-interferon alpha-2b monotherapy (PMT) vs its combination with ribavirin (PCT1-low dose ribavirin and PCT2-high dose ribavirin). We studied 56 patients that were part of two randomized, controlled, clinical trials. At baseline, high TNF-alpha levels paralleled the degree of inflammation as determined by histology. In PCT2, a significant reduction was seen in levels of TNF-alpha, TGF-beta and fibrosis scores when comparing baseline with follow-up. In sustained responders, regardless of therapy, the histological activity scores were lower at follow-up as compared to baseline. In conclusion, PCT2 is able to constantly reduce and sustain TNF-alpha levels, which is responsible for the sustained decline in liver inflammation as shown by the histological activity index and it is also able to reduce fibrosis as judged both by TGF-beta levels and fibrosis scores.     

Prediction of effect of pegylated interferon alpha-2b plus ribavirin combination therapy in patients with chronic hepatitis C infection

Treatment with pegylated interferon alpha-2b (PEGIFN) plus ribavirin (RBV) is standard therapy for patients with chronic hepatitis C. Although the effectiveness, patients with high titres of group Ib hepatitis C virus (HCV) respond poorly compared to other genotypes. At present, we cannot predict the effect in an individual. Previous studies have used traditional statistical analysis by assuming a linear relationship between clinical features, but most phenomena in the clinical situation are not linearly related. The aim of this study is to predict the effect of PEG IFN plus RBV therapy on an individual patient level using an artificial neural network system (ANN). 156 patients with HCV group 1b from multiple centres were treated with PEGIFN (1.5 μg/kg) plus RBV (400-1000 mg) for 48 weeks. Data on the patients' demographics, laboratory tests, PEGIFN, and RBV doses, early viral responses (EVR), and sustained viral responses were collected. Clinical data were randomly divided into training data set and validation data set and analyzed using multiple logistic regression analysis (MLRs) and ANN to predict individual outcomes. The sensitivities of predictive expression were 0.45 for the MLRs models and 0.82 for the ANNs and specificities were 0.55 for the MLR and 0.88 for the ANN. Non-linear relation analysis showed that EVR, serum creatinine, initial dose of Ribavirin, gender and age were important predictive factors, suggesting non-linearly related to outcome. In conclusion, ANN was more accurate than MLRs in predicting the outcome of PEGIFN plus RBV therapy in patients with group 1b HCV.     

Serum levels of Th17 associated cytokines in chronic hepatitis C virus infection

The Th17-mediated immune response was investigated in patients chronically infected with hepatitis C virus (HCV) by determining the serum levels of the cytokines involved in the induction of the Th17 response (TGF-β and IL-6), the cytokines produced by Th17 cells (IL-17A, IL-17F and IL-22) and the cytokines whose production is stimulated by Th17 lymphocytes (IL-8 and GM-CSF). We investigated the relationships among the levels of these cytokines by assessing clinical findings, liver histology and viremia. Sixty untreated patients and 28 healthy individuals were included in the study. Cytokine levels were determined using ELISA. Differences between HCV and control groups were identified in the median levels of IL-17F (controls=172.4pg/mL; HCV=96.8pg/mL, p<0.001) and IL-8 (controls=30.1pg/mL; HCV=18.1pg/mL, p<0.05). IL-6 levels were higher in patients presenting moderate liver necroinflammation than in patients with mild or no liver necroinflammation (p<0.05). IL-17F levels were increased in patients that had increased ALT levels. Additionally, a strong positive correlation was observed between IL-17F and IL-22 levels in the two groups investigated, and the IL-17F/IL-22 ratio was lower in the patients infected with HCV (p<0.0001). Patients with low HCV viral loads had higher median levels of IL-8 (32.5pg/mL) than did patients with high HCV loads (16.7pg/mL, p<0.05). These results suggest that in chronic hepatitis C infection, IL-17F and IL-8 could be associated with the control of liver injury and infection, respectively.     

Serum levels of trace elements and iron-deficiency anemia in adult Vietnamese

This study was aimed at assessing the serum levels of vitamin A, copper, zinc, selenium, and iron among adult Vietnamese with and without iron-deficiency anemia. Blood was collected from adult Vietnamese living in the midland of northern Vietnam. One hundred twenty-three subjects in the age range 20–60 yr were included in the study. Anemia, where the concentration of hemoglobin in whole blood is less than 120 g/L in females and 130 g/L in males, was found in 30% (37/123) of the study population. The levels of vitamin A and selenium in the sera of anemic subjects (n=37) were significantly lower than that in nonanemic group (n=86). On the other hand, no significant differences were observed in the concentrations of copper and zinc between the two groups. This study was the first to show serum levels of trace elements in adult Vietnamese, providing useful baseline information for further studies.     

Liver-directed gamma interferon gene delivery in chronic hepatitis C

Gamma interferon (IFN-gamma) has been shown to inhibit replication of subgenomic and genomic hepatitis C virus (HCV) RNAs in vitro and to noncytolytically suppress hepatitis B virus (HBV) replication in vivo. IFN-gamma is also known for its immunomodulatory effects and as a marker of a successful cellular immune response to HCV. Therapeutic expression of IFN-gamma in the liver may therefore facilitate resolution of chronic hepatitis C, an infection that is rarely resolved spontaneously. To analyze immunomodulatory and antiviral effects of liver-specific IFN-gamma expression in vivo, we intravenously injected two persistently HCV-infected chimpanzees twice with a recombinant, replication-deficient HBV vector and subsequently with a recombinant adenoviral vector. These vectors expressed human IFN-gamma under control of HBV- and liver-specific promoters, respectively. Gene transfer resulted in a transient increase of intrahepatic IFN-gamma mRNA, without increase in serum alanine aminotransferase levels. Ex vivo analysis of peripheral blood lymphocytes demonstrated enhanced CD16 expression on T cells and upregulation of the liver-homing marker CXCR3. Moreover, an increased frequency of HCV-specific T cells was detected ex vivo in the peripheral blood and in vitro in liver biopsy-derived, antigen-nonspecifically expanded T-cell lines. None of these immunologic effects were observed in the third chimpanzee injected with an HBV control vector. Despite these immunologic effects of the experimental vector, however, IFN-gamma gene transfer did not result in a significant and long-lasting decrease of HCV titers. In conclusion, liver-directed IFN-gamma gene delivery resulted in HCV-specific and nonspecific activation of cellular immune responses but did not result in effective control of HCV replication.     

Serum vitamin D levels are not predictive of the progression of chronic liver disease in hepatitis C patients with advanced fibrosis

In animal models and human cross-sectional studies, vitamin D deficiency has been associated with liver disease progression. Vitamin D supplementation has been suggested as a treatment to prevent disease progression. We sought to evaluate the role of vitamin D levels in predicting chronic liver disease development. We conducted a nested case-control study of vitamin D levels in subjects with (cases) and without (controls) liver histologic progression or clinical decompensation over the course of the HALT-C Trial. Vitamin D levels were measured at 4 points over 45 months. 129 cases and 129 aged-matched controls were included. No difference in baseline vitamin D levels were found between cases and controls. (44.8 ng/mL vs. 44.0 ng/mL, P = 0.74). Vitamin D levels declined in cases and controls over time (P = 0.0005), however, there was no difference in the level of decline (P = 0.37). Among study subjects with diabetes mellitius, baseline vitamin D levels were higher in cases, 49.9 ng/mL, than controls, 36.3 ng/mL. (P = 0.03) In addition, baseline vitamin D levels were higher in black case subjects, 32.7 ng/mL, than in black control subjects, 25.2 ng/mL (P = 0.08) No difference in vitamin D levels was found between patients with and without progression of hepatitis C-associated liver disease over 4 years. Our data do not suggest any role for vitamin D supplementation in patients with advanced chronic hepatitis C and raise the possibility that higher vitamin D levels may be associated with disease progression.     

Elevated levels of interleukin-8 in serum are associated with hepatitis C virus infection and resistance to interferon therapy

Hepatitis C virus (HCV), a major cause of liver disease worldwide, is frequently resistant to the antiviral alpha interferon (IFN). We have recently found that the HCV NS5A protein induces expression of the proinflammatory chemokine IL-8 to partially inhibit the antiviral actions of IFN in vitro. To extend these observations, in the present study we examined the relationship between levels of IL-8 in serum, HCV infection, and biochemical response to IFN therapy. Levels of IL-8 were significantly elevated in 132 HCV-infected patients compared to levels in 32 normal healthy subjects and were also significantly higher in patients who did not respond to IFN therapy than in patients who did respond to therapy. This study suggests that HCV-induced changes in levels of chemokine and cytokine expression may be involved in HCV antiviral resistance, persistence, and pathogenesis.     

Immunomodulators in combined therapy of patients with chronic hepatitis

The efficacy of the use of triple schemes in combined therapy of chronic virus hepatitis was estimated and its safety was monitored. The problems of therapy of mixed hepatitis in drug addicts are discussed. Immunotropic agents, increasing the efficacy of the standard therapy of chronic affections of the liver, are suggested to be used as the third remedy in the combined therapy.     

Histological change after interferon therapy in chronic hepatitis C in view of iron deposition in the liver

We examined the efficacy of interferon (IFN) therapy for chronic hepatitis C (CHC) in view of the change of liver histology and iron staining before and after IFN therapy. Enrolled in this study were 109 patients with CHC who completed IFN treatment and were followed for at least 1 yr after the end of IFN therapy. Serum iron, unsaturated-iron-binding capacity (UIBC), and total-iron-binding capacity (TIBC) were assessed before IFN therapy. Knodell’s histological activity index (HAI) score and iron staining were examined in 55 patients in whom liver biopsy was performed at two points: before and 1 yr after IFN therapy. Serum iron levels before IFN therapy did not correlate with the response to IFN. The HAI score significantly decreased after IFN therapy in complete responders (p<0.01) and biochemical responders (p<0.01). Three factors in the HAI, periportal necrosis, intralobular necrosis, and portal inflammation, but not fibrosis, were significantly decreased in complete responders (p<0.01) and biochemical responders (p<0.01). Of 55 patients, 23 (41.8%) were positive for iron staining before IFN therapy and 14 of 55 (25.5%) after IFN therapy. The positive rate for iron staining tended to decrease after IFN therapy, not correlating to the response to IFN, but the change was not statistically significant. In conclusion, the histological improvement by IFN therapy was mostly seen in necroinflammatory changes but not in fibrosis at least 1 yr after IFN, and iron staining tended to decrease after IFN therapy.     

Long-term changes in trace elements in patients undergoing chronic hemodialysis

Although the connection between aluminum intoxication and dialysis dementia was identified in the 1980s, understanding of trace element imbalances in hemodialysis patients is as yet incomplete. Recent application of newer inductively coupled plasma-mass spectrometry (ICP/MS) techniques has resulted in renewed study of this population. We used ICP/MS to evaluate serum concentrations of Cu, Se, Zn, Mn, and Ni in a relatively large population of hemodialysis patients compared with healthy age-matched controls. Comparison were also done by duration of hemodialysis treatment to see whether length of treatment correlates with severity of imbalance. Patients had significantly lower concentrations of the three elements Se, Zn, and Mn. Patients had significantly higher concentrations of Ni, and there was a positive correlation between duration and severity of imbalance for this one element. There was no difference in Cu concentrations between patients and controls. Our findings confirm relative Ni excess and deficiencies of Se, Zn, and Mn in hemodialysis patients, documenting the value of ICP/MS in research work on trace element imbalances as well as clinical monitoring of individual patients.     

Serum alpha-fetoprotein predicts treatment outcome in chronic hepatitis C patients regardless of HCV genotype

We examined the association between serum alpha-fetoprotein (AFP) level and sustained virological response (SVR) in 93 chronic hepatitis C patients. The SVR rate was much higher among patients with serum AFP levels below rather than above the median value (5.7 ng/ml) (58.7% and 19.2%, respectively; P<0.0001). Serum AFP should be added to the list of factors predictive of treatment response in chronic hepatitis C.     

Genetic variants associated with the progression of hepatocellular carcinoma in hepatitis C Egyptian patients

Background

Hepatocellular carcinoma (HCC) associated to infection with hepatitis C virus (HCV) has become the fastest-rising cause of cancer-related deaths. Genetic variations may play an important role in the development of HCC in HCV patients. Ghrelin exerts anti-inflammatory, antifibrotic and hepatoprotective effects on chronically injured hepatic tissues. Ghrelin gene shows several single nucleotide polymorphisms (SNPs) including − 604G/A, Arg51Gln, and Leu72Met. Hemochromatosis gene (HFE) mutations namely C282Y and H63D may cause hepatic iron overload, thus increasing the risk of HCC in HCV patients.

Aim

To investigate the association of progression of HCC with ghrelin and HFE gene polymorphisms in HCV Egyptian patients.

Methods

Seventy-nine chronic HCV patients (thirty-nine developed HCC and forty did not), and forty healthy control subjects were included in the study. The polymorphisms were evaluated by PCR/RFLP analysis, and related protein levels were measured by either ELISA or colorimetric assays.

Results

The three tested SNPs on ghrelin gene were detected in the studied groups, only one SNP (Arg51Gln) showed significantly higher GA, AA genotypes and A allele frequencies in hepatitis C patients who developed HCC than in hepatitis C patients without HCC and controls. Of the two mutations studied on HFE gene only H63D heterozygous allele was detected, and its frequency did not statistically differ among studied groups.

Conclusion

Our results suggest that A allele at position 346 of the ghrelin gene is associated with susceptibility to HCC in hepatitis C patients.     

Liver protein profiling in chronic hepatitis C: identification of potential predictive markers for interferon therapy outcome

The current anti-hepatitis C virus (HCV) therapy, based on pegylated-interferon alpha and ribavirin, has limited success rate and is accompanied by several side effects. The aim of this study was to identify protein profiles in pretreatment liver biopsies of HCV patients correlating with the outcome of antiviral therapy. Cytosolic or membrane/organelle-enriched protein extracts from liver biopsies of eight HCV patients were analyzed by two-dimensional fluorescence difference gel electrophoresis and mass spectrometry. Overall, this analysis identified 21 proteins whose expression levels correlate with therapy response. These factors are involved in interferon-mediated antiviral activity, stress response, and energy metabolism. Moreover, we found that post-translational modifications of dihydroxyacetone kinase were also associated with therapy outcome. Differential expression of the five best performing markers (STAT1, Mx1, DD4, DAK, and PD-ECGF) was confirmed by immunoblotting assays in an independent group of HCV patients. Finally, we showed that a prediction model based on the expression levels of these markers classifies responder and nonresponder patients with an accuracy of 85.7%. These results provide evidence that the analysis of pretreatment liver protein profiles is valuable for discriminating between responder and nonresponder HCV patients, and may contribute to reduce the number of nonresponder patients exposed to therapy-associated risks.     

Serum levels of preS antigen (HBpreSAg) in chronic hepatitis B virus infected patients

Background

Viruses that are incorporating host cell proteins might trigger autoimmune diseases. It is therefore of interest to identify possible host proteins associated with viruses, especially for enveloped viruses that have been suggested to play a role in autoimmune diseases, like human herpesvirus 6A (HHV-6A) in multiple sclerosis (MS).

Results

We have established a method for rapid and morphology preserving purification of HHV-6A virions, which in combination with parallel analyses with background control material released from mock-infected cells facilitates qualitative and quantitative investigations of the protein content of HHV-6A virions. In our iodixanol gradient purified preparation, we detected high levels of viral DNA by real-time PCR and viral proteins by metabolic labelling, silver staining and western blots. In contrast, the background level of cellular contamination was low in the purified samples as demonstrated by the silver staining and metabolic labelling analyses. Western blot analyses showed that the cellular complement protein CD46, the receptor for HHV-6A, is associated with the purified and infectious virions. Also, the cellular proteins clathrin, ezrin and Tsg101 are associated with intact HHV-6A virions.

Conclusion

Cellular proteins are associated with HHV-6A virions. The relevance of the association in disease and especially in autoimmunity will be further investigated.