• Korean Journal of Veterinary Research
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• v.25 no.1
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• pp.7-17
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• 1985

Many investigators have been pursuing various attempts so far to produce hepatitis B surface antigen(HBsAg) vaccines using the techniques such as isolation from plasma of chronic HBsAg carrier, recombinant DNA technique or preparation of synthetic peptides specific for immunogenic determinants. Hepatitis B virus can not grow on any cell lines by the tissue culture technique at the present time. The plasma of chronic HBsAg carrier is expensive and its source is limited. The HBsAg from the recombinant DNA technique gave still very low yield. Another approach, therefore, has been initiated to develop a synthetic hepatitis B virus vaccine. The possible use of several distinct synthetic vaccines in prophylaxis can be facilitated by availability of full synthetic immunogens. Peptides synthesized for potential application as antiviral vaccines have been mostly tested in the form of conjugates with carrier proteins, although the free synthetic peptide can be immunogenic. To understand basic knowledges on the antigenicity and immunogenicity of a synthetic peptide specific for major immunogenic determinant of HBsAg, a nonapeptide, $H_2N^{139}Cys-Thr-Lys-Pro-Thr-Asp-Gly-^{146}Asn-Aba$ COOH, which corresponds to HBsAg amino acid residues 139 to 147, was synthesized by the Merrifield's solid-phase method with a slight modification. The antigenicity and immunogenicity of this specific synthetic peptide were examined comparing with purified plasma-derived natural HBsAg. The results obtained are as follows; 1. The peptide synthesized showed the identical amino acid composition to the theoretical value. The degree of purification and molecular weight were acertained by methods of high performance liquid chromatography and mass spectrometry. 2. Using m-maleimidobenzoyl-N-hydroxysuccinimide ester as a conjugating agent, the synthetic peptide was conjugated to rabbit albumin and ${\gamma}$-globulin, tetanus and diphtheria toxoids, and keyhole limpet hemocyanin. Their conjugation yields were 8.3, 9.5, 15.8, 13.5, and 11.2%, respectively. 3. The natural HBsAg was purified from plasma of chronic HBsAg carrier. By the electron microscopic observation of the purified natural HBsAg preparation, no Dane particles were observed and the preparation showed negative DNA polymerase activity. 4. Antigenicity of the synthetic peptide and the plasma-derived natural HBsAg was determined by competition radioimmunoassay using $^{125}I$-natural HBsAg. Their 50% inhibitions appeared as $90{\mu}g/ml$ and $0.12{\mu}g/ml$ for the synthetic peptide and the natural HBsAg, respectively. This indicates that the former was about 750-fold less antigenic than the latter. 5. Immunogenicity of the synthetic peptide was determined by administering the peptide-carrier conjugates into rabbits with and without Freund's complete adjuvant. Regardless the carrier proteins and adjuvant, positive immune responses to the synthetic peptide were observed. The higher antibody titers, however, were shown in the groups administered with Freund's complete adjuvant. 6. Immunizing dose 50% in mice of the various peptide-carrier conjugates was 5.47, 6.00, 65.16, 31.25 and $13.03{\mu}g/dose$ for rabbit albumin and ${\gamma}$-globulin, tetanus and diphtheria toxoids, and keyhole limpet hemocyanin, respectively, while the natural HBsAg showed $0.65{\mu}g/dose$. 7. It was postulated that homologous proteins prefer to heterologous ones as the carriers.

• Journal of Life Science
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• v.22 no.2
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• pp.259-265
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• 2012

A bacterial strain, CK214, exhibiting high motility on an LB agar (1.5%, w/v) surface was isolated from the environment. The formation of unusual agar shrinking around colonies on agar plates was observed. The strain grew on minimal media containing pure agar as a sole carbon source. The cell-free culture supernatant of CK214 generated a reduced form of sugar in the in vitro reaction with the use of pure agar as a substrate, suggesting the secretion of an agar-degrading enzyme. The CK214 strain showed swarming motility on the solid media containing a wide range of concentrations of agar (0.5, 1.0, 1.5, 2.0% w/v). Various tests, including Gram staining, API analysis, and phylogenetic analysis based on 16S rDNA sequences identified that the CK214 strain was a G(+) rod-shaped bacterium grouped in genus Paenibacillus. Electron microscopic analysis demonstrated that the P. CK214 strain is peritrichously flagellated. Through transposon random mutagenesis, several agar-degrading activity defective mutants (ADMs) were generated. These mutants will be used in the future experimentation for the study of the correlation between agar-degrading activity and motility.

• Proceedings of the Korean Society of Plant Pathology Conference
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• 1994.06a
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• pp.11-26
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• 1994

Crown gall of stonefruit and nut trees is one of the very few plant diseases subject to efficient biological control. The disease is caused by the soil-inhabiting bacteria Agrobacterium tumefaciens and Agrobacterium rhizogenes and the original control organism was a non-pathogenic isolate of A. rhizogenes strain K84. Control is achieved by dipping planting material in a cell suspension of strain K84 which specifically inhibits pathogenic strains containing a nopaline Ti plasmid. Because the agrocin 84-encoding plasmid (pAgK84) is conjugative, it can be transmitted from the control strain to pathogenic strains which, as a result, become immune to agrocin 84 and cannot be controlled. To prevent this happening, the transfer genes on pAgK84 were located and then largely eliminated by recombinant DNA technology. The resulting construct, strain K1026, is transfer deficient but controls crown gall just as effectively as does strain K84. Field data from Spain confirm that pAgK84 can transfer to pathogenic recipients from strain K84 but not from strain K1026. The latter has been registered in Australia as a pesticide and is the first genetically engineered organism in the world to be released fro commercial use. It is recommended as a replacement for strain K84 to prevent a breakdown in the effectiveness of biological control of crown gall. Several reports indicate that both strains K84 and K1026 sometimes control crown gall pathogens that are resistant to agrocin 84. A possible reason for this is that both strains produce a second antibiotic called 434 which inhibits growth of nearly all isolates of A. rhizogenes, both pathogens and non-pathogens. Crown gall of grapevine is caused by another species, Agrobacterium vitis. It is resistant to agrocin 84 and cannot be controlled by strains K84 or K1026. It is different from other crown gall pathogens in several characteristics, including the fact that, although a rhizosphere coloniser, its also lives systemically in the vascular tissue of grapevine. Pathogen free propagating material can be obtained from tissue culture or, less surely, by heat therapy of dormant cuttings. A number of laboratories are searching for a biocontrol strain that will prevent, or at least delay, reinfection. A non-pathogenic A. vitis strain F/25 from South Africa looks very promising in this regard.

• Journal of Society of Preventive Korean Medicine
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• v.28 no.2
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• pp.113-130
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• 2024

Objectives : This study was conducted to evaluate the cytotoxic protective effects of Jinsimyusaeng-hwan (Modified Ojayeonjong-hwan) against oxidative stress. Methods : 1,1-diphenyl-2-picrylhydrazyl (DPPH), 2,2'-azinobis-(3-ethyl-benzothiazoline-6-sulfonic acid)(ABTS) radical scavenging activity and ferric reducing ability of plasma (FRAP) method were used to estimate the antioxidant activity of Jinsimyusaeng-hwan. C2C12 myoblasts were used to reevaluate the antioxidant effects. And apoptosis analysis, mitochondrial membrane potential analysis, measurement of intracellular reactive oxygen species levels were conducted to investigate antioxidant activity of Jinsimyusaeng-hwan. Results : In comparison of DPPH free radical and ABTS cationic radical scavenging activity, it increased as the concentration of water extracts of Jinsimyusaeng-hwan(WEJ) and 70% ethanol extracts of Jinsimyusaeng-hwan (EEJ) increased. In the results of comparing the total phenol content and reducing power using the FRAP method, extracts with high total phenol content also showed high reducing power. In comparison of the protective effect against H₂O₂-induced oxidative stress in C2C12 myoblasts, WEJ had no significant effect, but the EEJ inhibited H₂O₂-mediated cytotoxicity in a concentration-dependent manner. The cytotoxic protective effect of EEJ against oxidative stress in C2C12 myoblasts was correlated with their inhibitory effects on H₂O₂-induced apoptosis and cell cycle arrest. In H₂O₂-treated C2C12 myoblasts, the apoptosis inhibitory effects of EEJ were associated with suppression of mitochondrial dysfunction and DNA damage. The protective effect of EEJ against H₂O₂-induced oxidative stress in C2C12 myoblasts were directly related to the inhibition of ROS generation. Conclusion : Jinsimyusaeng-hwan extracts have cytotoxic protective effects against oxidative stress, and it was better in 70% ethanol extract than in water extract.

• Pediatric Infection and Vaccine
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• v.14 no.2
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• pp.171-178
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• 2007

Purpose : We investigated clinical characteristics of real-time PCR proven EBV viremia patients who were not serologically diagnosed but clinically suspected, and compared it to serologically proven EBV infected patients. Methods : The study population consisted of 45 patients, who were suspected acute EBV infection at Wonju Christian Hospital Department of Pediatrics, Yonsei University Wonju College of Medicine from Jan. 2004 to Dec. 2006. real-time PCR of cell free serum was performed to prove EBV viremia. Then we chose $102.5copies/{\mu}g$ DNA as a suitable cutoff level for EBV associated diseases. Results : There are 4 patients diagnosed as EBV infection by serologically and 15 patients diagnosed as EBV viremia by real-time PCR quantitative measurement. The most common presenting symptoms and signs of EBV viremia was fever in 11 cases (73%). Atypical lymphocytosis was found in 4 cases (27%). Increased AST, ALT levels were observed in 13 cases (87%), 12 cases (80%), respectively. We could diagnose 5 cases of EBV viremia younger than one year of age. They revealed clinical symptoms which could be found in EBV infection. The serologically diagnosed patients had hepatomegaly and splenomegaly in 3 cases (75%). All serologically confirmed patients have leukocytosis above $20,000/mm^3$, among them 2 cases (50%) had higher percentage (>15%) of atypical lymphocytes. The AST/ALT level above 50 IU/L were demonstrated in all cases. Conclusion : Serologically unproven real-time PCR EBV viremia patients revealed similar clinical findings with that of serologically proven EBV infected patients. So, it is meaningful to perform EBV real-time PCR for the diagnosis of EBV infection especially for the cases younger than 1 year of age.

• Applied Microscopy
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• v.32 no.2
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• pp.91-105
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• 2002

Urea transport in the kidney is mediated by a family of transporter proteins that includes renal urea transporters (UT-A) and erythrocyte urea transporters (UT-B). The cDNA of five isoforms of rat UT-A, UTA1, UT-A2, UT-A3, UT-A4, and UT-A5 have been cloned. The purpose of this study was to examine the expression of UT-A (L194), which marked UT-A1, UT-A2 and UT-A4. Male Sprague-Dawley rats, weighing approximately 200 g, were divided into three group: control rats had free access to water, dehydrated rats were deprived of water for 3 d, and water loaded rats had free access to 3% sucrose water for 3 d before being killed. The kidneys were preserved by in vivo perfusion through the abdominal aorta with the 2% paraformaldehyde-lysine- periodate (PLP) or 8% paraformaldehyde solution for 10 min. The sections were processed for immunohistochemical studies using pre-embedding immunoperoxidase method and immunogold method. In the normal rat kidney, UT-A1 was expressed intensely in the cytoplasm of the inner medullary collecting duct (IMCD) cell and UT-A2 was expressed on the plasma membrane of the terminal portion of the shortloop descending thin limb (DTL) cells (type I epithelium) and of the long-loop DTL cells (type II epithelium) in the initial part of the inner medulla. Immunoreactivity for UT-A1 in the IMCD cells, was decreased in dehydrated animals whereas strongly increased in water loaded animals compared with control animals. In the short-loop DTL, immunoreactivity for UT-A2 was increased in intensity in both dehydrated and water loaded groups. However, in the long-loop DTL of the outer part of the inner medulla, immunoreactivity for UT-A2 was markedly increase in intensity in dehydrated group, but not in water loaded group. In conclusion, in the rat kidney, UT-A1 is located in the cytoplasm of IMCD cells, whereas UT-A2 is located in the plasma membrane of both the short-and long-loop DTL cells. Immunohistochemistry studies revealed that UT-A1 and UT-A2 may have a different role in urea transport and are regulated by different mechanisms.