• BMB Reports
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• v.31 no.3
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• pp.281-286
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• 1998

Hepatitis C virus (HCV) core proteins from two different isolates (HCV-1 and HCV-RH) were expressed in Spotioptera Jrugiperda (Sf9) insect cells. The RH core consisted of two major species of proteins (21 kDa and 19 kDa). On the other hand, the HCV-1 core was approximately 16 kDa in a SDS-PAGE gel. Both core proteins were phosphorylated in vivo on serine residues. Furthermore, the RH core but not HCV-1 core formed dimers, indicating that the protein conformation of the core in these two isolates is dfferent from one another. Immunofluorescence studies showed that the RH core was present in the cytoplasm, whereas the HCV-1 core was localized predominantly to the nucleus in recombinant baculovirus-infected insect cells. Since the major difference between the two isolates is the codon 9 of the core protein, a single amino acid substitution appears to play a major role in the protein conformation and these properties may reflect the different biological functions of core proteins in HCV-infected cells.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.20
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• pp.8837-8842
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• 2014

Background: Hepatitis C virus (HCV) infection is an important cause of liver cancer in Thailand. The highest prevalence of anti-HCV positive among Thai blood donors is found in the northeastern region. The present analysis of the genotype distribution among anti-HCV positive northeastern-Thai blood donors was conducted to provide a base for the epidemiological pattern of HCV infection in this region. Materials and Methods: A total of 112 HCV seropositive healthy blood donors were randomly selected and tested for the presence of HCV-RNA by RT-PCR. HCV-RNA positive samples were genotyped by direct sequencing at core region genomes and confirmed by phylogenetic analysis. Results: HCV viremia was found in 94.6% (106/112) of HCV seropositive blood donors. There were 3 major genotypes distributed among this population. HCV genotype 3a was the most prevalent (71.7%) followed by genotypes 1a (7.5%), 1b (7.5%), 6i (3.8%), 6f (2.8%) and 6n (1.9%). Conclusions: HCV genotype 3a in asymptomatic infections in northeastern Thailand is significantly higher than other previous reports. Subgenotype 6 prevalence is less than in neighboring countries and distribution patterns differ. The findings are relevant as predictors for using interferon therapy in this population.

• BMB Reports
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• v.30 no.3
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• pp.234-236
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• 1997

Chronic hepatitis C virus (HCV) infection is associated with the rapid development of cirrhosis and hepatocellular carcinoma. It has been reported that the amount of HCV RNA may be correlated with the progression of hepatitis and may be a prognostic marker for treatment of HCV patients. The direct detection of HCV RNA by reverse transcription-polymerase chain reaction (RT-PCR) is widely used to determine the presence of circulating virions. The most relevant limit of this approach is the lack of quantitative information about the viral titer. In the present study, we developed the method for HCV quantitation using competitive reverse transcription (CRT)-PCR using the deleted HCV standard. The serially diluted standard was added in titrated amounts to the target HCV RNA. The mixture was then reverse transcribed and amplified in the same reaction tube. The methods were evaluated using over 110 HCV-PCR positive samples in Koreans. About 59% of the samples were judged to contain $10^{5}-10^{6}$ copies of HCV RNA in 1 ml of serum.

• Microbiology and Biotechnology Letters
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• v.50 no.3
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• pp.422-429
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• 2022

The hepatitis C virus (HCV) genome contains a positive-sense single-stranded RNA molecule, and it is classified into 8 genotypes and 87 subtypes. Globally, over 350,000 people die from liver cirrhosis and hepatocellular carcinoma caused by HCV each year. Here, the genotype distribution of HCV was estimated in the population in Cheonan, Korea using Sanger sequencing. In addition, the correlation between HCV RNA level and genotype was assessed using real-time polymerase chain reaction (PCR); similarly, the correlation of HCV RNA level with isolation year (2007-2016) was determined using 463 consecutive serum samples obtained from patients at Dankook University Hospital, Cheonan, Korea. In 2007, genotype 1b (54.2%) was predominant, followed by genotypes 2a (41.7%), 1a (2.1%) and 3a (2.1%); whereas in 2016, the predominant genotype was 2a (49.0%), followed by genotypes 1b (46.9%), 3b (2%), and 4a (2%). Neither age nor sex was correlated with HCV genotype. Furthermore, the mean HCV RNA level decreased significantly from 2012 to 2016 (p < 0.05). However, no significant correlations between genotype and HCV RNA level were found. Overall, the findings revealed that genotypes 2a and 1b were the most common in Cheonan, and the prevalence of HCV genotype 1b tended to decrease over the past decade.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.18
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• pp.7725-7730
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• 2014

Hepatocellular carcinoma (HCC) is the third most common cause for cancer death in the world, now being especially linked to chronic hepatitis C virus (HCV) infection. This case-control study consisting of 65 HCC patients and 82 patients with other malignant tumours as controls was conducted to determine the association of HCV markers with HCC. Serum of each participant was obtained for detection of HCV Ab and RNA by DNA enzyme immunoassay (DEIA). Twenty six per cent (26.0%) of HCC patients had positive anti-HCV which was significantly greater than the control group (p=0.001). HCC patients significantly have a risk of exposure to HCV infection almost 3 times than the control group (OR=2.87, 95% C.I=1.1-7). Anti-HCV seropositive rate was significantly (p=0.03) higher among old age HCC patients and increases with age. Males with HCC significantly showed to have more than 9 times risk of exposure to HCV infection (OR=9.375, 95 % CI=1.299-67.647) than females. HCV-RNA seropositive rate was (70.8%) significantly higher among HCC patients compared to (22.2%) the control group (p=0.019). The most prevalent genotype (as a single or mixed pattern of infection) was HCV-1b. This study detected a significantly higher HCV seropositive rate of antibodies and RNA in HCC patients.

• IMMUNE NETWORK
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• v.6 no.1
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• pp.20-26
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• 2006

Background: Rheumatoid arthritis (RA) is a chronic and systemic inflammatory disease that is characterized by invasive synovial hyperplasia, leading to progressive joint destruction. Recent studies have described that RA is caused by virus, bacteria or outside material. Approximately 2 to 20% of RA cases arc reported to be associated with infected hepatitis C virus (HCV). However, the mechanisms underlying virus-induced RA are still unknown. Moreover, few molecular studies have addressed the inflammatory aspects of HCV-associated autoimmune RA. In this study, we aimed to determine whe ther or not another HCV core protein transactivates the IL-8 gene expression, prototypic chemokine, in synovial cell. Methods: To establish the HCV core expressing stable synovial cell line, pCI-neo-core, a plasmid encoding HCV core protein, were transfected to HIG-82 cell line that is an established cell line from rabbit periaricular soft tissue. We examined the morphological changes and cell cycle distribution of HIG-82 cells with expression of HCV core protein by inverted microscopy and flow cytometry analysis, respectively. Also, we determined the mRNA levels of Interleukin (IL)-6 and IL-8 related to the inflammation by RT-PCR and then analyzed regulation of IL-8 expression by the NF-${\kappa}B$ pathway. Results: Our study showed no significant differences in morphology and cell cycle between HIG-82 control cell line and HIG-82 expressing HCV core protein. However, expression of HCV core protein induces the IL-8 mRNA expression in HIG-82 core cells via activated NF-${\kappa}B$ pathway. Conclusion: These results suggest that HCV core protein can lead to enhanced IL-8 expression. Such a proinflammatory role may contribute to the etiologic pathogenesis in RA patients with HCV infection.

• BMB Reports
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• v.37 no.2
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• pp.192-198
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• 2004

The hepatitis C virus (HCV) core protein is believed to be one of viral proteins that are capable of preventing virus-infected cell death upon various stimuli. But, the effect of the HCV core protein on apoptosis that is induced by various stimuli is contradictory. We examined the possibility that the HCV core protein affects the ceramide-induced cell death in cells expressing the HCV core protein through the sphingomyelin pathway. Cell death that is induced by $C^2$-ceramide and bacterial sphingomyelinase was analyzed in 293 cells that constitutively expressed the HCV core protein and compared with 293 cells that were stably transfected only with the expression vector. The HCV core protein inhibited the cell death that was induced by these reagents. The protective effects of the HCV core protein on ceramide-induced cell death were reflected by the reduced expression of $p21^{WAF1/Cip1/Sid1}$ and the sustained expression of the Bcl-2 protein in the HCV core-expressing cells with respect to the vector-transfected cells. These results suggest that the HCV core protein in 293 cells plays a role in the modulation of the apoptotic response that is induced by ceramide. Also, the ability of the HCV core protein to suppress apoptosis might have important implications in understanding the pathogenesis of the HCV infection.

• Journal of Microbiology
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• v.42 no.4
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• pp.361-364
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• 2004

Internal ribosome entry site (IRES) of the hepatitis C virus (HCV) is known to be essential for HCV replication and most conserved among HCV variants. Hence, IRES RNA is a good therapeutic target for RNA-based inhibitors, such as ribozymes. We previously proposed a new anti-HCV modulation strategy based on trans-splicing ribozymes, which can selectively replace HCV transcripts with a new RNA that exerts anti-HCV activity. To explore this procedure, sites which are accessible to ribozymes in HCV IRES were previously determined by employing an RNA mapping method in vitro. In this study, we evaluate the intracellular accessibility of the ribozymes by comparing the trans-splicing activ­ities in cells of several ribozymes targeting different sites of the HCV IRES RNA. We assessed the intra­cellular activities of the ribozymes by monitoring their target-specific induction degree of both reporter gene activity and cytotoxin expression. The ribozyme capable of targeting the most accessible site iden­tified by the mapping studies then harbored the most active trans-splicing activity in cells. These results suggest that the target sites predicted to be accessible are truly the most accessible in the cells, and thus, could be applied to the development of various RNA-based anti-HCV therapies.

• Journal of Microbiology and Biotechnology
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• v.16 no.10
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• pp.1634-1639
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• 2006

Hepatitis C virus (HCV)-encoded nonstructural protein 5B (NS5B) possesses RNA-dependent RNA polymerase activity, which is considered essential for viral proliferation. Thus, HCV NS5B is a good therapeutic target protein for the development of anti-HCV agents. In this study, we isolated two different kinds of nuclease-resistant RNA aptamers with 2'-fluoro pyrimidines against the HCV NS5B from a combinatorial RNA library with 40 nucleotide random sequences, using SELEX technology. The isolated RNA aptamers were observed to specifically and avidly bind the HCV NS5B with an apparent $K_d$ of 5 nM and 18 nM, respectively, in contrast with the original RNA library that hardly bound the target protein. Moreover, these aptamers could partially inhibit RNA synthesis of the HCV subgenomic replicon when transfected into Huh-7 hepatoma cell lines. These results suggest that the RNA aptamers selected in vitro could be useful not only as therapeutic agents of HCV infection but also as a powerful tool for the study of the HCV RNA-dependent RNA polymerase mechanism.

• BMB Reports
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• v.37 no.6
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• pp.735-740
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• 2004

Hepatitis C virus (HCV) is a causal agent of the chronic liver infection. To understand HCV morphogenesis, we studied the assembly of HCV structural proteins in insect cells. We constructed recombinant baculovirus expression vectors consisting of either HCV core alone, core-E1, or core-E1-E2. These structural proteins were expressed in insect cells and were examined to assemble into particles. Neither core-E1 nor core-E1-E2 was capable of assembling into virus-like particles (VLPs). It was surprising that the core protein alone was assembled into core-like particles. These particles were released into the culture medium as early as 2 days after infection. In our system, HCV structural proteins including envelope proteins did not assemble into VLPs. Instead, the core protein itself has the intrinsic capacity to assemble into amorphous core-like particles. Furthermore, released core particles were associated with HCV RNA, indicating that core proteins were assembled into nucleocapsids. These results suggest that HCV may utilize a unique core release mechanism to evade the hosts defense mechanism, thus contributing to the persistence of HCV infection.