No abstract available.
Antiviral Agents/therapeutic use ; Hepacivirus/drug effects/physiology ; Hepatitis C/*drug therapy ; Humans

OBJECTIVETo study the inhibition effect of HCV NS5A on p53 protein transactivity and its possible mechanism.

METHODSLuciferase reporter gene system was used for the study of p53 transactivity on p21 promoter and electrophorectic mobility-shift assay (EMSA) was applied to observe whether HCV NS5A could suppress the binding ability of p53 protein to its specific DNA sequence.

RESULTSEndogenous p53 protein could stimulate p21 promoter activity, and the relative luciferase activity increased significantly (3.49 x 10(5) vs 0.60 x 10(5), t = 5.92, P<0.01). Exogenous p53 protein also up-regulated p21 promoter driving luciferase expression, comparing to the control group (0.47 x 10(5)), the relative luciferase activity increased (5.63 x 10(5)) obviously (t = 10.12, P<0.01). HCV NS5A protein inhibited both endogenous and exogenous p53 transactivity on p21 promoter in a dose-dependent manner (F > or = 20.71, P<0.01). In the experiment of EMSA, p53 could bind to its specific DNA sequence, but when co-transfected with HCV NS5A expressing vector, the p53 binding affinity to its DNA decreased.

CONCLUSIONHCV NS5A can inhibit p53 protein transactivity on p21 promoter through its inhibiting of p53 binding ability to the specific DNA sequence.

Hepacivirus ; genetics ; Humans ; Promoter Regions, Genetic ; Transcriptional Activation ; drug effects ; Tumor Suppressor Protein p53 ; drug effects ; genetics ; metabolism ; physiology ; Viral Core Proteins ; genetics ; Viral Nonstructural Proteins ; genetics ; pharmacology

OBJECTIVETo investigate the influence of miR-122 on IFN-α treatment for HCV infection.

METHODSHuh7.5.1 cells infected with HCV were treated with miR-122 mimics (20 nmol/L, 100 nmol/L, 400 nmol/L) and/or IFN-α (1000 IU/ml). The relative expression of HCV RNA was detected by real-time polymerase chain reaction (PCR). Huh7.5.1 cells were treated with different amounts of HCV (107 copies, 106 copies and 105 copies) and/or IFN-α (1000 IU/ml).

RESULTSIFN-α suppressed the replication of HCV in a time-dependent manner, resulting in a ≊ 83% reduction of HCV at 48 h. MiR-122 mimics facilitated replication of HCV RNA in a dose-dependent manner (P<0.05). The antiviral effect of IFN-α was inverted to levels of miR-122 mimics (20 nmol/L, 100 nmol/L, 400 nmol/L), (73.3% ± 3.5% compared with 84% ± 4.5%, P>0.05; 64.67% ± 5.5% compared with 84% ± 4.5%, P>0.05; 56.33% ± 5.1% compared with 84% ± 4.5%, P<0.05). The antiviral effect of IFN-α was inverted to HCV load (105 copies group compared with 107 copies group, P<0.05).

CONCLUSIONMiR-122 facilitates replication of HCV RNA in the cell culture system; and the expression of miR-122 may partly counteract the anti-HCV effect of IFN-α.

Antiviral Agents ; pharmacology ; Cell Line, Tumor ; Hepacivirus ; drug effects ; genetics ; physiology ; Humans ; Interferon-alpha ; pharmacology ; MicroRNAs ; genetics ; RNA, Viral ; genetics ; Transfection ; Virus Replication ; drug effects ; genetics

OBJECTIVETo investigate the inhibitive effects of small interfering RNA (siRNA) on hepatitis C virus (HCV) replication in cells infected by HCV in vitro.

METHODSThe HCV RNA transcripts prepared by pFL-JC1 were transfected into Huh-7.5.1 cells. Na ve Huh-7.5.1 cells were incubated with the supernatants of transfected cells and the expression of HCV core protein in infected cells was detected by indirect immunofluorescence. The infected cells were transfected with 4, 40 and 200 nmol/L of NS5B siRNA for 24 h, 48 h and 72 h, respectively. The normal Huh-7.5.1 cells were transfected with 4, 40 and 200 nmol/L of NS5B siRNA. Group of blank, lipofectamine 2000, unrelated siRNA and IFNα-2b (1000 IU/ml) served as controls. The HCV RNA and PKR mRNA levels were examined by quantitative RT-PCR.

RESULTSThe HCV core protein in HCV infected cells was detected. Compared with control groups, the HCV RNA levels in infected cells significantly decreased when transfected with 40 and 200 nmol/L of siRNA for 24 h; 4, 40 and 200 nmol/L of siRNA for 48 h and 72 h (P<0.05). The HCV RNA levels in infected cells treated with IFNα-2b (1000 IU/ml) for 24 h, 48 h and 72 h were significantly lower than those in control groups (P<0.05 or P<0.01). The PKR mRNA levels in Huh-7.5.1 cells transfected with siRNA of three concentrations did not have significant difference, as compared with control groups (P>0.05).

CONCLUSIONsiRNA against HCV NS5B region can effectively inhibit HCV replication in HCV infected cells, but can not activate the dsRNA-dependent protein kinase (PKR).

Cell Line, Tumor ; Hepacivirus ; drug effects ; genetics ; physiology ; Humans ; RNA, Small Interfering ; pharmacology ; Transfection ; Viral Nonstructural Proteins ; genetics ; Virus Replication ; drug effects

Hepatitis c virus (HCV) infection has become one of the global public health problem,while there is no vaccine to prevent HCV infection, the so-called "cocktail" therapy that use a combination of drugs targeting multiple steps in the HCV infection cycle could achieve better curative effect. the process of HCV entering into host cell is the important step of drug intervention, in which HCV envelope protein El and E2, Host cell factors including Heparan sulfate(HS), CD81, scavenger receptor class B type I (SR-BI), Occludin (OCLD), Claudin (CLDN), low densitity lipoprotein receptor (LDLR), dendritic cell-specific ICAM-3-grabbing nonintegrin (DC-SIGN), Liver/lymph node specific ICAM-3-grabbing integrin(L-SIGN), trans- ferrin receptor 1 (TfR1) and so on play a important role. The virus and the host factors can be used as targets of hcv entry inhibitors many studies have shown that as novel and promising compounds, HCV entry inhibitors combinating with other drugs can be more effective in the treatment of HCV, this paper have re- viewed targets and inhibitors of HCV enterring into host cell since 1990s.
Animals ; Antiviral Agents ; pharmacology ; Hepacivirus ; drug effects ; physiology ; Hepatitis C ; genetics ; metabolism ; virology ; Humans ; Receptors, Virus ; genetics ; metabolism ; Viral Envelope Proteins ; genetics ; metabolism ; Virus Internalization ; drug effects

OBJECTIVEHomeopathic nosodes have seldom been scientifically validated for their anticancer effects. This study was conducted to examine if a recently developed hepatitis C nosode has demonstrable anticancer potential in cancer cells in vitro.

METHODSAnticancer effects of Hepatitis C 30C (Hep C 30), if any, were initially tested on three cancer cell lines, HepG2 (liver cancer), MCF-7 (breast cancer) and A549 (lung cancer) and one normal liver cell line WRL-68 cells and subsequently a more thorough study using further scientific protocols was undertaken on HepG2 cells (against WRL-68 cells as the normal control) as HepG2 cells showed better anticancer response than the other two. Three doses, one at 50% lethal dose (LD50) and the other two below LD50, were used on HepG2 cells subsequently. Protocols like apoptosis induction and its possible signaling mechanism were deployed using immunoblots of relevant signal proteins and confocal microscopy, with particular reference to telomerase and topoisomerase II (Top II) activities, two strong cancer biomarkers for their direct relationship with divisional activities of cells and DNAs.

RESULTSHep C 30 induced apoptosis, caused distorted cell morphology typical of apoptotic cells, increased reactive oxygen species generation and produced increased DNA nicks. Further it enhanced pro-apototic signal proteins like Bax, cytochrome c and inhibited anti-apoptotic signal proteins, Bcl-2, cytochrome c and caspase-3, changed mitochondrial membrane potential and caused externalization of phosphatidylserine. The drug also decreased expression of two cancer biomarkers, Top II and telomerase, consistent with its anticancer effect.

CONCLUSIONHep C 30 has demonstrable anticancer effects against liver cancer cells in vitro.

Antineoplastic Agents ; pharmacology ; Apoptosis ; drug effects ; Cell Survival ; drug effects ; DNA Topoisomerases, Type II ; metabolism ; Hep G2 Cells ; Hepacivirus ; Humans ; Liver Neoplasms ; drug therapy ; enzymology ; pathology ; Materia Medica ; Mitochondria ; drug effects ; physiology ; Telomerase ; metabolism

Adult ; Aged ; Alcohol Drinking ; adverse effects ; Alcoholism ; complications ; Hepacivirus ; physiology ; Hepatitis C, Chronic ; pathology ; virology ; Humans ; Interferon-alpha ; adverse effects ; therapeutic use ; Liver ; immunology ; pathology ; Liver Cirrhosis ; etiology ; pathology ; Male ; Middle Aged ; RNA, Viral ; biosynthesis ; Virus Replication ; drug effects

BACKGROUND/AIMS: Enhanced replication of hepatitis C virus (HCV) is well described in the setting of moderate to severe immunosuppression. The aims of this retrospective study were to determine the incidence of enhanced HCV replication in hepatocellular carcinoma (HCC) patients undergoing transarterial chemolipiodolization (TACL) and to identify the factors associated with enhanced replication of HCV. The clinical pattern of enhanced HCV replication was compared with hepatitis B virus (HBV) reactivation during TACL. METHODS: This study enrolled 49 anti-HCV-seropositive patients who were diagnosed with HCC between January 2005 and December 2010 and who underwent TACL using epirubicin and/or cisplatin with consecutive HCV RNA copies checked. For comparison, 46 hepatitis B surface antigen1-positive patients with HCC who were treated with TACL were also enrolled. The frequency, associated factors, and clinical outcomes of enhanced HCV replication were analyzed and compared with those of HBV reactivation during TACL. RESULTS: Enhanced replication of HCV occurred in 13 (26.5%) of the 49 anti-HCV-seropositive patients during TACL. Of these 13 patients, 4 developed hepatitis, but none of the subjects developed decompensation due to the hepatitis. No significant clinical factors for enhanced HCV replication during TACL were found. Compared with HBV reactivation, the frequency of hepatitis attributed to enhanced HCV replication was significantly lower than that for HBV reactivation (8.2% vs. 23.9%, P=0.036). CONCLUSIONS: TACL can enhance HCV replication; however, the likelihood of hepatitis and decompensation stemming from enhanced HCV replication was lower than that for HBV reactivation in patients undergoing TACL.
Adult ; Aged ; Antineoplastic Agents/*administration & dosage/adverse effects/pharmacology ; Carcinoma, Hepatocellular/complications/*therapy ; Chemoembolization, Therapeutic/*adverse effects ; Drug Therapy, Combination ; Female ; Hepacivirus/drug effects/*physiology ; Hepatitis B/complications/epidemiology/virology ; Hepatitis B Surface Antigens/blood ; Hepatitis B virus/drug effects/*physiology ; Hepatitis C/complications/epidemiology/virology ; Humans ; Liver Neoplasms/complications/*therapy ; Male ; Middle Aged ; RNA, Viral/analysis ; Retrospective Studies ; Virus Activation ; *Virus Replication

Metabolomics represents an emerging and powerful discipline that provides an accurate and dynamic picture of the phenotype of bio-systems through the study of potential metabolites that could be used as therapeutic targets and for the discovery of new drugs. Hepatitis C virus (HCV) is a leading cause of liver disease worldwide, and is a major burden on public health. It is hypothesized that an animal model of HCV infection would produce unique patterns of endogenous metabolites. Herein, a method for the construction of efficient networks is presented with regard to the proteins of bear bile powder (PBBP) that protect against HCV as a case study. Ultra-performance liquid chromatography, coupled with electrospray ionization/quadrupole-time-of-flight high definition mass spectrometry (UPLC-HDMS), coupled with pattern recognition methods and computational systems analysis were integrated to obtain comprehensive metabolomic profiling and pathways of the large biological data sets. Among the regulated pathways, 38 biomarkers were identified and two unique metabolic pathways were indicated to be differentially affected in HCV animals. The results provided a systematic view of the development and progression of HCV, and also could be used to analyze the therapeutic effects of PBBP, a widely used anti-HCV medicine. The results also showed that PBBP could provide satisfactory effects on HCV infection through partially regulating the perturbed pathway. The most promising use in the near future would be to clarify the pathways for the drugs and obtain biomarkers for these pathways to help guide testable predictions, provide insights into drug action mechanisms, and enable an increase in research productivity toward metabolomic drug discovery.
Animals ; Antiviral Agents ; chemistry ; metabolism ; pharmacology ; Bile ; chemistry ; metabolism ; Hepacivirus ; drug effects ; physiology ; Hepatitis C ; drug therapy ; virology ; Humans ; Male ; Metabolomics ; Proteins ; chemistry ; metabolism ; pharmacology ; Proteomics ; Spectrometry, Mass, Electrospray Ionization ; Tupaiidae ; Ursidae

Hepatitis C Virus (HCV) is associated with a severe liver disease and increased frequency in the development of hepatocellular carcinoma. Overexpression of HCV core protein is known to transform fibroblast cells. Phospholipase D (PLD) activity is commonly elevated in response to mitogenic signals, and has also been overexpressed and hyperactivated in some human cancer cells. The aim of this study was to understand how PLD was regulated in the HCV core protein-transformed NIH3T3 mouse fibroblast cells. We observed that PLD activity was elevated in the NIH3T3 cells overexpressing HCV core protein over the vector alone-transfected control cells, however, expression levels of PLD protein and protein kinase C (PKC) in the HCV core protein-transformed cells was similar to the control cells. Phorbol 12-myristate 13-acetate (PMA), which is known to activate PKC, stimulated PLD activity significantly more in the core protein-transformed cells, in comparison with that of the control cells. PLD activity assay using PKC isozyme-specific inhibitor and PKC translocation experiment showed that PKC-delta was mainly involved in the PMA- induced PLD activation in the core-transformed cells. Moreover, in cells overexpressing HCV core protein, PMA also stimulated p38 kinase more potently than that of the control cells, and an inhibitor of p38 kinase abolished PMA-induced PLD activation in cells overexpressing HCV core protein. Taken together, these results suggest that PLD might be implicated in core protein-induced transformation.
Animals ; Cell Line, Transformed ; *Cell Transformation, Viral ; Fibroblasts/enzymology/virology ; Hepacivirus/genetics/*physiology ; Mice ; NIH 3T3 Cells ; Phospholipase D/*metabolism ; Protein Kinase C/antagonists & inhibitors/physiology ; Protein Transport/drug effects ; Research Support, Non-U.S. Gov't ; Tetradecanoylphorbol Acetate/*analogs & derivatives/pharmacology ; Transfection ; Up-Regulation ; Viral Core Proteins/genetics/*metabolism ; p38 Mitogen-Activated Protein Kinases/physiology