• 대한바이러스학회지
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• 제27권2호
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• pp.115-128
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• 1997

Three cDNA fragments located within NS5 region of HCV were synthesized by RT using viral RNA extracted from blood sample of Korean patient as a template. The cDNAs were amplified by PCR, cloned into the T-vector, and the nucleotide sequences were determined. Comparative analysis of the nucleotide and amino acid sequence of NS5 cDNAs showed that it is closely related with HCV type 1b. The cloned NS5 cDNA showed 91-94% homology at the nucleotide sequence level and 96-98% homology at the amino acid sequence level with several strains of the HCV type 1b. The NS5 cDNAs were subcloned into E. coli expression vectors to construct pRSETA5-1, pTHAN5-1, pRSETC5-2, pRSETBB1, pRESTCB1 and pRSETB-H3. Expression of the NS5 proteins was achieved by inducing the promoter with isopropyl-thio-${\beta}$-D-galactoside (IPTG) and confirmed by SDS-polyacrylamide gel electrophoresis. The NS5 proteins were immunoreactive against sera from Korean hepatitis C patients in Western blot analysis. Among the recombinant NS5 proteins, pRSETAS-1 plasmid derived protein, coded from aa2022 to aa2521 of HCV polyprotein, showed the strongest immunoreactivity against sera from Korean hepatitis C patients in immunoblot analysis. These results suggest that NS5 proteins would be useful as an antigen for detection of antibody against HCV in the blood samples.

• Journal of Microbiology and Biotechnology
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• 제10권6호
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• pp.823-828
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• 2000

The complete nucleotide sequence of hepatitis B virus DNA isolated from Korean patient serum was determined and characterized, and its phylogenetic relation was then investigated. The viral genome was 3,215 base pairs long and included four well known open reading frames (i.e. surface antigens, core antigens, X protein and DNA polymerase). The sequence of the surface antigen showed that the HBV genome under investigation, designated HBV 315, was characteristic of subtype adr. A phylogenetic analysis using the total genome sequence revealed that HBV315 was grouped into genomic group C together with isolates from Japan, China, Thailand, Polynesia, and New Caledonia. The mean percent similarity between HBV315 and other HBV isolates in genomic group C was 97.25%, and that with other genomic groups ranged from 86.16% to 91.25%. The predicted amino acid sequences of HBV315 were compared with two closely related subtype adr isolates, M38636 and D12980. The results showed that the X gene product was identical in the three strains, while there were significant amino acid sequence differences between HBV315 and M38636 in the Pre-S1 and Pre-S2 regions.

• 대한바이러스학회지
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• 제26권1호
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• pp.31-45
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• 1996

C형 간염바이러스 (HCV)는 각 개체간에 뉴클레오티드 서열상의 다양성을 나타내고, 이러한 유전적 다양성이 임상병리적 증상과 밀접한 연관이 있을 것으로 고려되어 왔다. 본 연구에서는 HCV E1과 NS5B 부위의 염기서열 분석을 통해 한국의 C형 간염바이러스의 분포와 다양성에 관해 분석하고, 발생계통도를 그려 HCV간의 진화적 거리를 확인하였다. 염기서열분석은 서울대학교 병원과 충남대학교 병원으로부터 얻은 56개의 HCV-양성 혈청을 대상으로 RT-PCR과 PCR 과정을 통해 얻은 유전자 산물을 클로닝하여 수행되었다. 56개의 혈청중 53개의 샘플에서 HCV RNA가 검출되었다. 이들 53개 샘플에 대한 분석 곁과, 유전형 1a, 1b, 2a, 2b, 7a가 각각 5.7, 45.3, 45.3, 1.9, 1.9%로 분포하고 있고, 1b형과 2a형이 한국에서의 주요한 HCV 유전형으로 밝혀졌다. 본 연구는 염기서열 분석을 통해 한국에서 1b형과 마찬가지로 2a형도 높은 빈도로 분포하고 있고, 비록 분포 빈도는 낮지만 1a 형과 7a 형도 존재하고 있음을 밝힌 최초의 보고이다.

• Asian Pacific Journal of Cancer Prevention
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• 제15권22호
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• pp.9635-9641
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• 2014

Background: This retrospective study was aimed to investigate the efficacy of prophylactic agents in hepatocellular carcinoma (HCC) patients receiving TACE and compare the difference between lamivudine and entecavir. Materials and Methods: A consecutive series of 203 HBV-related HCC patients receiving TACE were analyzed including 91 patients given prophylactic agents. Virologic events, defined as an increase in serum HBV DNA level to more than 1 log10 IU/ml higher than the nadir level, hepatitis flares due to HBV reactivation and progression free survival (PFS) were the main endpoints. Results: Some 48 (69.6%) reached virologic response. Prophylaxis significantly reduced virologic events (8.8% vs 58.0%, p=0.000) and hepatitis flares (1.1% vs 13.4%, p=0.001). Patients presenting undetectable HBV DNA levels displayed a significantly improved PFS as compared to those who never achieved undetectable HBV DNA. Prophylaxis and e-antigen positivity were the only significant variables associated with virologic events. In addition, prophylaxis was the only independent protective factor for hepatitis flares. Liver cirrhosis, more cycles of TACE, HBV DNA negativity, a lower Cancer of the Liver Italian Program score, non-metastasis and no hepatitis flares were protective factors for PFS. Prophylactic lamivudine demonstrated similar efficacy as entecavir. Conclusions: Prophylactic agents are efficacious for prevention of HBV reactivation in HCC patients receiving TACE. Achievement of undetectable HBV DNA levels displayed a significant capability in improving PFS. Moreover, persistent tumor residual lesions, positive HBV DNA and hepatitis B flares might be causes of tumor progression in these patients.

• 대한수의학회지
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• 제39권4호
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• pp.825-831
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• 1999

This study was performed to determine the etiological agents in concurrent disorders in gastrointestinal and respiratory tract. Most of dogs had clinical signs including nasal and ocular discharge, coughing, vomiting, and diarrhea. Of the 22 dogs, seropositive rates of each virus were 54.5% (12/22) against canine distemper virus, 90.9% (20/22) against canine adenovirus 1, 36.4% (8/22) against canine adenovirus 2, 18.2% (4/22) against canine parvovirus, 81.8% (18/22) against canine hepatitis virus and 59.1% (13/22) against canine coronavirus. Canine distemper virus and canine parvovirus infection were 54.6% (12/22) in histopathological examination. In addition, mixed infections of canine distemper virus and adenovirus 2 were 9.1% (2/22). While simple infection of canine adenovirus 2 were 9.1% (2/22). E coli and Staphylococcus spp were isolated in facts as a rate of 72.7% (16/22) and 40.9% (9/22), respectively. Conclusionally, it is also estimated that environmental stress might be one of the causative factors.

• Parasites, Hosts and Diseases
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• 제56권1호
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• pp.61-70
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• 2018

We developed a Rapid Diagnostic Test (RDT) kit for detecting IgG/IgM antibodies against Zika virus (ZIKV) using monoclonal antibodies to the envelope (E) and non-structural protein 1 (NS1) of ZIKV. These proteins were produced using baculovirus expression vector with Sf9 cells. Monoclonal antibodies J2G7 to NS1 and J5E1 to E protein were selected and conjugated with colloidal gold to produce the Zika IgG/IgM RDT kit (Zika RDT). Comparisons with ELISA, plaque reduction neutralization test (PRNT), and PCR were done to investigate the analytical sensitivity of Zika RDT, which resulted in 100% identical results. Sensitivity and specificity of Zika RDT in a field test was determined using positive and negative samples from Brazil and Korea. The diagnostic accuracy of Zika RDT was fairly high; sensitivity and specificity for IgG was 99.0 and 99.3%, respectively, while for IgM it was 96.7 and 98.7%, respectively. Cross reaction with dengue virus was evaluated using anti-Dengue Mixed Titer Performance Panel (PVD201), in which the Zika RDT showed cross-reactions with DENV in 16.7% and 5.6% in IgG and IgM, respectively. Cross reactions were not observed with West Nile, yellow fever, and hepatitis C virus infected sera. Zika RDT kit is very simple to use, rapid to assay, and very sensitive, and highly specific. Therefore, it would serve as a choice of method for point-of-care diagnosis and large scale surveys of ZIKV infection under clinical or field conditions worldwide in endemic areas.

• Genomics & Informatics
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• 제7권4호
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• pp.187-194
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• 2009

Cytochrome P450 2E1 (CYP2E1) is a member of the cytochrome P450 superfamily, and it is a key enzyme responsible for the metabolic activation of many smallmolecular-weight compounds such as alcohol, which is classified as a human carcinogen. In this study, we identified 19 single nucleotide polymorphisms (SNPs) in CYP2E1 in Korean population. In these SNPs, we examined possible genetic association of CYP2E1 polymorphisms with HBV clearance and the risk of hepatocellular carcinoma (HCC). Five common polymorphic sites were selected, CYP2E1 polymorphisms at rs381-3867, rs3813870, rs2070673, rs2515641 and rs2480257, considering their allele frequencies, haplotype-tagging status and LDs for genotyping in larger-scale subjects (n=1,092). Statistical analysis demonstrated that CYP2E1 polymorphisms and haplotypes show no significant association with HBV clearance, HCC occurrence and onset age of HCC (p>0.05). Previous studies, however, have shown contradictory findings on associations of CYP2E1 polymorphisms with CYP2E1 activities and HCC risk. Comparing the contrasting results of previous researches suggest that CYP2E1 polymorphism is associated with CYP2E1 activity induced by ethanol, but is not directly associated with HCC risk. CYP2E1 variation/haploype information identified in this study will provide valuable information for future studies on CYP2E1.

• 대한간학회지
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• 제24권4호
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• pp.384-391
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• 2018

Backgrounds/Aims: The objective of our study was to determine the epidemiological, laboratory, and serological characteristics of patients with chronic hepatitis B virus (HBV) infection and normal transaminases. The study also aimed to evaluate liver damage by measuring the liver fibrosis (LF) grade and to identify possible factors associated with the presence of fibrosis. Methods: A retrospective observational study was conducted in patients with chronic HBV infection and classified as inactive carriers or immune-tolerant. Epidemiological variables of age, sex, immigrant, alcohol consumption, and body mass index (BMI), as well as virological variables (HBV DNA) and transaminase level were collected throughout the follow-up. The LF grade was evaluated by transient elastography. The cutoff value for significant fibrosis (SF) was liver stiffness ${\geq}7.9kPa$. Results: A total of 214 patients were included in the analysis, and 62% of them had a BMI ${\geq}25kg/m^2$. During follow-up, 4% of patients showed transaminase elevation (<1.5 times normal). Most patients had a viral DNA level <2,000 IU/mL (83%). Data on LF were available in 160 patients; of these, 14% had SF, 9% F3, and 6% F4. The variables associated with the presence of SF were transaminase alteration during follow-up, as 23% of patients with SF had elevated transaminases versus 3% of patients without SF (P<0.005), and BMI, as the vast majority of patients with SF (88%) had a BMI ${\geq}25kg/m^2$ versus 56% of patients without SF (P<0.05). Conclusions: In patients with chronic HBV infection and normal transaminases, liver damage does not seem to be related to DNA levels, alcohol consumption, or immigrant status. SF seems to be associated with transaminase alteration during follow-up and elevated BMI. It is therefore recommended to measure LF grade with validated non-invasive methods in such patients.

• 대한수의학회지
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• 제44권4호
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• pp.655-662
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• 2004

The aim of this work was the validation of a rapid real-time PCR assay based on TaqMan technology for the unequivocal identification of infectious canine hepatitis (ICH) virus, to be used directly on DNA purified from blood specimens. A real-time PCR system targeting at the E3 ORFA gene sequence of canine adenovirus type 1 was optimized and validated through comparative analysis of samples using conventional PCR system. The real-time PCR assay based on TaqMan technology could disclose 23 (37.7%) out of 61 samples as PCR positive. In contrast, 18 (29.5%) samples were found PCR positive when conventional PCR was applied on these samples. The use of the ABI Prism 7700 sequence detection system allowed the efficient determination of the amplified product accumulation through a fluorogenic probe. The entire real-time TaqMan PCR assay, including DNA extraction, amplification, and detection could be completed within 3 hours. The detection method of real-time TaqMan PCR assay was 1,000 times more sensitive than conventional PCR. Real-time TaqMan probe and primer set developed and optimized in this study is a sensitive, rapid and accurate method for detection of ICH virus and can be effective screening tool for the detection of ICH in a diagnostic laboratory routines.

• 생명과학회지
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• 제28권9호
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• pp.1007-1015
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• 2018

E6AP (E6-associated protein)는 C형 간염바이러스(hepatitis C virus, HCV)의 코어 단백질 유비퀴틴화와 프로테오좀 분해를 유도하여 캡시드 조립을 저해함으로써 HCV 복제를 억제하는 것으로 알려져 있다. 반면에 HCV 코어 단백질은 숙주의 항바이러스 방어계에 대항하고 자신의 유비퀴틴-의존적 프로테아좀 분해를 막기 위하여 DNA 메틸화를 통하여 E6AP 발현을 저해하는 전략을 진화과정에서 획득하였다. 본 연구에서는 HCV 코어 단백질이 E6AP 발현을 저해하는 기전을 밝혀내고자 하였다. HCV 코어 단백질은 HepG2 세포에서 DNA 메틸화 효소들인 DNMT1, 3a 및 3b의 단백질 수준과 효소 활성을 증가시켜 프로모터 과메틸화를 통하여 E6AP 발현을 저해하였지만 p53를 발현하지 않는 Hep3B 세포에서는 이러한 효과들이 관찰되지 않았다. 흥미롭게도 Hep3B 세포에 p53만 과발현시키면 HCV 코어 단백질이 없더라도 DNMT가 활성화되고 프로모터 과메틸화를 통하여 E6AP 발현이 저해되었다. 또한 p53 녹다운 및 과발현 실험을 통하여 p53 활성화가 HCV 코어 단백질의 효과에 필수적임을 알 수 있었다. 이로 인하여 Hep3B 보다 HepG2 세포에서 낮은 수준의 유비퀴틴화된 HCV 코어 단백질이 검출되었다. 따라서 HCV 코어 단백질은 p53-의존적으로 자신의 유비퀴틴-매개성 프로테아좀 분해를 저해한다.