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Generation and Characterization of a Neutralizing Human Monoclonal Antibody to Hepatitis B Virus PreS1 from a Phage-Displayed Human Synthetic Fab Library

  • Jo, Gyunghee;Jeong, Mun Sik;Wi, Jimin;Kim, Doo Hyun;Kim, Sangkyu;Kim, Dain;Yoon, Jun-Yeol;Chae, Heesu;Kim, Kyun-Hwan;Hong, Hyo Jeong
    • Journal of Microbiology and Biotechnology
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    • v.28 no.8
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    • pp.1376-1383
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    • 2018
  • The hepatitis B virus (HBV) envelope contains small (S), middle (M), and large (L) proteins. PreS1 of the L protein contains a receptor-binding motif crucial for HBV infection. This motif is highly conserved among 10 HBV genotypes (A-J), making it a potential target for the prevention of HBV infection. In this study, we successfully generated a neutralizing human monoclonal antibody (mAb), 1A8 (IgG1), that recognizes the receptor-binding motif of preS1 using a phage-displayed human synthetic Fab library. Analysis of the antigen-binding activity of 1A8 for different genotypes indicated that it can specifically bind to the preS1 of major HBV genotypes (A-D). Based on Bio-Layer interferometry, the affinity ($K_D$) of 1A8 for the preS1 of genotype C was 3.55 nM. 1A8 immunoprecipitated the hepatitis B virions of genotypes C and D. In an in vitro neutralization assay using HepG2 cells overexpressing the cellular receptor sodium taurocholate cotransporting polypeptide, 1A8 effectively neutralized HBV infection with genotype D. Taken together, the results suggest that 1A8 may neutralize the four HBV genotypes. Considering that genotypes A-D are most prevalent, 1A8 may be a neutralizing human mAb with promising potential in the prevention and treatment of HBV infection.

Molecular Cloning and Expression in Escherichia coli of a Rabbit Globin Gene (유전공학적 방법에 의한 토끼 글로빈 유전자의 재조합과 대장균에서의 발현)

  • Jang, Sung-Key;Park, Hyune-Mo
    • The Korean Journal of Zoology
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    • v.27 no.2
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    • pp.103-116
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    • 1984
  • The structural gene of rabbit hemoglobin was cloned into Pst I site of pBR322 in E. coli. The complementary DNA (cDNA) was synthesized from rabbit globin mRNA with avian myeloblastosis viral reverse transcriptase, and then RNA was destroyed at pH 11. The double stranded cDNA was synthesized with both Klenow fragment of E. coli DNA polymerase I and reverse transcriptase and then the hairpin loop was opened with Sl nuclease. Double stranded cDNA was subsequently tailed with dCTP and annealed to dGMP-tailed vector DNA. After transformation and initial screening of appropriate clones by plasmid size, the cloned colonies were identified by in situ colony hybridization using by plasmid size, the cloned colonies were identified by in situ colony hybridization using $[^32P]$-labeled cDNA probes and characterized the inserts with restriction endonucleases. The expression of cloned globin gene was investigated by standard radioimmunoassay using rat anti-rabbit Hb serum as primary antibody and goat antirat IgG serum as secondary antibody. The result suggested that the chimeric proteins (the part of $\\beta$-lactamase from the vector pBR322 and globin from rabbit) were supposedly produced in E. coli and the product had the antigenic determinant of rabbit hemoglobin.

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Efficient Plant Regeneration from Alfalfa Callus by Osmotic Stress Treatment (알팔파 캘러스로부터 삼투압 스트레스 처리에 의한 효율적인 식물체 재분화)

  • Kim, J.S.;Lee, D.G.;Lee, S.H.;Woo, H.S.;Lee, B.H.
    • Journal of Animal Science and Technology
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    • v.46 no.5
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    • pp.879-886
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    • 2004
  • Effects of culture mediwn supplements and osmotic stress treatment on embryogenic callus induction and somatic embryogenesis were investigated in order to optimize tissue culture conditions of alfalfa(Medicago sativa L.). SH mediwn containing 5mgIL 2,4-D and 0.2mgIL kinetin was optimal for embryogenic callus induction from cotyledon tissue of alfalfa. Somatic embryos were formed when the embryogenic callus was cultured on SH mediwn supplemented with ImgIL 2,4-D and 2mgIL BA. Supplementation of 5mM L-proline and IgIL casein hydrolysate into the regeneration mediwn further increased plant regeneration frequency. Osmotic stress treatment of callus appeared to improve the frequency of somatic embryo formation, but the frequency of somatic embryo formation differed by the osmotic stress treatment using different osmotic stressors. The highest plant regeneration frequency of 30.7% was observed when embryogenic callus was treated with 0.7M sucrose for 18h. Efficient regeneration system established in this study will be useful for molecular breeding of alfalfa through genetic transformation.

AN IMMUNOHISTOCHEMICAL STUDY ON THE DISTRIBUTION OF CGRP CONTAINING NERVE FIBERS AFTER PULP EXPOSURE IN RAT MOLAR (흰쥐대구치 치수노출후 치수조직내 CGRP함유 신경섬유의 분포에 관한 면역조직화학적 연구)

  • Kim, Eun-Soung;Park, Il-Yoon;Moon, Joo-Hoon
    • Restorative Dentistry and Endodontics
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    • v.24 no.2
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    • pp.372-380
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    • 1999
  • The purpose of this study was to investigate the distribution of calcitonin gene-related peptide(CGRP) containing nerve fivers after pulp exposure in rats. The Spague-Dawley rats weighing about 250 - 300g were used. The animals were devided into normal control group and experimental groups. Experimental animals were sacrified on 2, 4, 7, 10 days after pulp exposure. The maxillary teeth and alveolar bone were removed and immersed in the 4% paraformaldehyde plus 0.1M phosphate buffer (pH 7.4). Serial frozen $50{\mu}m$ thick sections were cut with a cryostat. In the immunohistochemical staining procedure, the rabbit CGRP antibody was used as a primary antibody. The sections were incubated for 48 hours at $4^{\circ}C$, and placed into biotinylated anti-rabbit IgG as a secondary antibody and incubated in ABC (avidin-biotin complex), The sections were visualized by 0.05% 3.3 diaminobenzidine tetrahydrochloride. The results of this study were as follows: 1. In control group, CGRP containing nerve fibers ran parallel to the long axis of root and reached the coronal pulp. They were distributed on Raschkow plexus under the odontoblastic layer. 2. In 2 day group after pulp exposure, tissue necrosis and acute inflammation occurred and CGRP containing nerve fibers increased. In 4 day group, the necrotic tissue extended to the pulp and CGRP containing nerve fibers were distributed around the inflammation zone. 3. In 7 day group after pulp exposure, pulp necrosis occurred, and in 10 day group, the abscess under the necrotic pulp extended to the root apex area and CGRP containing nerve fibers were not observed in root canals. 4.The sprouting of CGRP nerve fibers was most remarkable at the pulp chamber under injury in 4 day group, and it was found at inflammation zone under the necrotic tissue in 7 day group and the remaining root pulp tissue in 10 day group. As mentioned above, CGRP nerve fibers had a tendency to increase around the inflammatory zone, especially around the acute inflammation tissue, when compared with control group. It is suggested that CGRP nerve fibers maybe related to the control of inflammatory response of pulp tissue.

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The Relationship between Helicobacter pylori Infection and Iron-Deficiency: Seroprevalence Study in 937 Pubescent Children (Helicobacter pylori 감염과 철 결핍의 관계: 937명의 사춘기 환아의 혈청 유병률 연구)

  • Kim, Sang-Jong;Kim, Bong-Lim;Kim, Soon-Ki;Choe, Yon-Ho
    • Pediatric Gastroenterology, Hepatology & Nutrition
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    • v.5 no.2
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    • pp.129-135
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    • 2002
  • Purpose: The purpose of this study was to investigate the relationship between Helicobacter pylori (H. pylori) infection and iron-deficiency anemia in pubescent children, susceptible to iron deficiency due to the high iron requirements for growth. Methods: Hemoglobin, serum iron, total iron-binding capacity, serum ferritin, and serum IgG antibodies to H. pylori were measured in 937 children (475 boys and 462 girls). Their ages ranged from 10 to 18 years. The prevalences of H. pylori infection were compared between groups, based on the presence or absence of anemia, hypoferritinemia, iron deficiency, and iron-deficiency anemia. The levels of hemoglobin, serum iron, total iron-binding capacity, transferrin saturation, and serum ferritin were obtained according to the presence or absence of H. pylori infection. Results: The prevalences of anemia, iron deficiency, iron-deficiency anemia, and H. pylori infection were 8.1%, 9.1%, 3.1%, and 20.8%, respectively. The H. pylori-positive rates in anemia, hypoferritinemia, and iron-deficiency group were 34.2%, 29.5%, and 35.3%, respectively, compared to 19.6% in the non-anemia group, 19.2% in the non-hypoferritinemia group, and 19.4% in the non-iron deficiency group. The H. pylori-positive rate in the iron-deficiency anemia group was 44.8% in comparison with 20.0% in the non-iron-deficiency anemia group. Hemoglobin and iron levels did not show any significant differences between the H. pylori-positive and -negative groups, whereas the serum ferritin level decreased significantly in the H. pylori-infected group. Conclusion: H. pylori infection is thought to be associated with iron deficiency in pubescent children.

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A Health Risk Assessment of Tributyltin Compounds in Fishes and Shellfishes in Korea. (국내 유통중인 어패류 섭취에 따른 유기주석화합물의 인체 위해성 평가)

  • Choi, Shi-Nai;Choi, Hye-Kyung;Song, Hoon;Oh, Chang-Hwan;Park, Jong-Sei
    • Journal of Food Hygiene and Safety
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    • v.17 no.3
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    • pp.137-145
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    • 2002
  • Tributyltin compounds have been increasingly used in the form of plastic stabilizers, catalytic agents, industrial agricultural biocides, antifouling paint, and pesticides. Among these organotin compounds, large amounts of tributyltin(TBT) and triphenyltin(TPT) have been used as antifouling agents because they have a superior ability to prevent marine organism from being encrusted on ship bottoms and in culturing nets. Environmental pollution by these organotin compounds in the aquatic environment were undertaken. The international maritime Organization's established a provisional tolerable daily intake(TDI) of 1.6[micro]g TBTO/kg/ B.W. The Food and Agiculture Organization (of the United Nations)/world Health Organization's (FAO/WHO) proposed a TDI of 0.5ug TPT/kg BW/d. This study is conducted monitoring of TBT on seafoods in Korea and risk assessment for exposure on TBT in seafoods. Total hazard index(using Reference Dose : 0.3 ug TBTO/kg B.W/day) of intake exposure on seafoods is 0.04 as the 50th percentile, 0.08 as the 95th percentile. This value is estimated by Monte-Carlo simulation using Crystal Ball(Decisioneering Co., 2001).

A Clinical and Pathological Analysis of Children with Membranoproliferative Glomerulonephritis According to the Clinical Manifestations at Presentation (발견 양상에 따른 소아 막증식성 사구체신염의 임상적 및 병리조직학적분석)

  • Jeon Chang-Ho;Kang Mi-Seon;Chung Woo-Yeong
    • Childhood Kidney Diseases
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    • v.8 no.2
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    • pp.186-194
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    • 2004
  • Purpose: Membranoproliferative glomeulonephritis(MPGN) has been diagnosed in an increasing number of asymptomatic cases. These cases have been detected by school urinary screening test even though the total cases of MPGN show a decreasing trend. We have analyzed the clinical and pathological characteristics of children with MPGN according to the clinical manifestations at the time of disease presentation. Methods: A total of 18 patients who had been diagnosed with idiopathic MPGN by percutaneous renal biopsy from January 1990 to February 2004 were involved in our study. The patients were divided into 2 groups as the school urinary screening(A) group and the symptomatic(S) group according to the clinical manifestations at the time of disease presentation. Results: Out of the total 18 patients, 8(44.4%) were in the S group and 10(55.6%) were in the A group. The mean serum total protein, albumin and $C_3$ levels in the S group were significantly lower than those levels of the A group, respectively($4.9{\pm}1.2\;g/dL,\;vs\;7.0{\pm}0.5\;g/dL\;P=0.002,\;2.8{\pm}0.9\;g/dL\;vs.\;4.1{\pm}0.3\;g/dL\;P=0.002,\;63.9{\pm}36.4\;mg/dL\;vs.\;100.8{\pm}39.5\;g/dL\;P=0.041$). The mean total protein amount of 24 hour collected urine in the S group were significantly higher than that of the A group($3684.0{\pm}2601.3\;mg/m^2\;vs.\;559.4{\pm}4.6.9\;mg/m^2$, respectively, P=0.001). Hypocomplementemia was observed in 11(61.1%) out of 18 patients at the time of disease onset, 7(87.5%) in the S group and 4(40%) in the A group. However the hypocomplementemia was decreased in 6(33.3%) out of 18 patients at the time of final follow-up, 3(37.5%) in the S group and 3(30%) in the A group. According to the pathologic type, hypocomplementemia was observed 8 patients(61.5%) with type I disease, 1 patients (100%) with type II disease, 2 patients(50%) in type III disease at the disease onset, but 4 patients(30.8%) in type I disease, 1 patient(100%) in type II disease, 1 patient(33.3%) with type III disease at the time of last follow-up. The incidence of cellular crescent formation and tubular atropy. as observed on light microscopy, were higher in the S group compared to the A group. Mean grade of capillary wall thickening and, mesangial proliferation were significantly higher in the S group. Conclusion: MPGN, as diagnosed in patients with only asymptomatic urinary abnormalities, has been increasing, it is more frequent in asymptomatic patients than in patients with presenting symptoms. Our result suggests that MPGN should be considered in the renal biopsy diagnosis regardless of serum $C_3$ level when urinary abnormalities are found by school urinary screening test.

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Prospective Study of the Immunologic Factors Affecting the Prognosis of Severe Community-Acquired Pneumonia (중증 지역사회획득 폐렴환자의 예후에 영향을 미치는 면역지표에 대한 연구)

  • Hwang, Jae-Kyung;Lee, Ho-Moeng;Song, Kwang-Sik;Park, Gye-Young;Park, Jeong-Woong;Park, Jae-Kyung;Jeong, Seong-Hwan;Ahn, Jeong-Yeal;Seo, Yiel-Hea;Nam, Gui-Hyun
    • Tuberculosis and Respiratory Diseases
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    • v.50 no.4
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    • pp.437-449
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    • 2001
  • Background : In the severe community-acquired pneumonia, it has been known that the immune status is occasionally suppressed. This study was performed to identify the immunologic markers related with the prognostic factors in severe community-acquired pneumonia. Methods : 23 patients with severe community-acquired pneumonia were involved in this study, and divided into survivor (16) and nonsurvivor (7) groups. In this study, the medical history, laboratory tests(complete blood counts, routine chemistry profile, immunoglobulins, complements, lymphocyte subsets, cytokines, sputum and blood culture, urine analysis), and chest radiographs were scrutinized. Results : 1) Both groups had lymphopenia(total lymphocyte count $995.6{\pm}505.7/mm^3$ in the survivor and $624.0{\pm}287.6/mm^3$ in the nonsurvivor group). 2) The T-lymphocyte count of the nonsurvivor group($295.9{\pm}203.0/mm^3$) was lower than the survivor group($723.6{\pm}406.5/mm^3$) (p<0.05). 3) The total serum protein(albumin) was $6.0{\pm}1.0(2.7{\pm}0.7)\;g/d{\ell}$ in the survivor and $5.2{\pm}1.5(2.3{\pm}0.8)g/d{\ell}$ in the nonsurvivor group. The BUN of the nonsurvivor group($41.7{\pm}30.0mg/d{\ell}$) was higher than that of the survivor group($18.9{\pm}9.8mg/d{\ell}$)(p<0.05). The creatinine concentration was higher in the nonsurvivor group($1.8{\pm}1.0mg/d{\ell}$) than that in the survivor group($1.0{\pm}0.3mg/d{\ell}$)(p<0.05). 4) The immunoglobulin G level was higher in the survivor group($1433.0{\pm}729.5mg/d{\ell}$) than in the nonsurvivor group($849.1{\pm}373.1mg/d{\ell}$) (p<0.05). 5) The complement $C_3$ level was $108.0{\pm}37.9mg/d{\ell}$ in the survivor group and $88.0{\pm}32.1mg/d{\ell}$ in the nonsurvivor group. 6) A cytokine study showed an insignificant difference in both groups. 7) Chronic liver disease, DM, and COPD were major underlying diseases in both groups. Conclusion : These results suggest that decreased a T-lymphocyte count and immunoglobulin G level, and an increased BUN and creatinine level may be associated with the poor prognosis of severe community-acquired pneumonia.

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Effects of Supplementing Herbs on Growth Performances, Blood Composition and Diarrhea in Hanwoo Calves (한방제재 첨가급여가 한우송아지의 성장, 혈액성상 및 설사에 미치는 영향)

  • Kim, Byung-Ki;Choi, Chang-Bon;Lee, Sang-Oug;Baek, Kyung-Hoon;Jung, Dae-Jin;Hwang, Eun-Gyeong
    • Journal of Animal Science and Technology
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    • v.53 no.5
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    • pp.451-459
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    • 2011
  • The current study was conducted to determine the effects of herbal supplement on growth performances and diarrhea in Hanwoo calves. Total 24 Hanwoo calves (3 treatments: 8 calves per treatment) were randomly assigned to either Control (no treatment), Treatment 1 (0.3% herb supplement), or Treatment 2 (0.5% herb supplement) diets for 150 days. Total body weight gain and total feed intake per head in Treatment 1 was 142.8 kg and 545.9 kg, respectively, and it was higher (p<0.05) compared to other groups. The blood total cholesterol range was 86.43~97.00 mg/dl, triglycerides 13.26~13.86 mg/dl, GOT 76.97~79.60 mg/dl, GPT 19.54~20.97 mg/dl, WBC 8.75~9.95 k/${\mu}{\ell}$, RBC 10.14~11.91 M/${\mu}{\ell}$, and hemoglobin 10.74~11.20 g/$d{\ell}$, respectively, with no significant (p>0.05) differences among treatments. The blood immunoglobulin G levels were 5.74~6.05 mg/ml which tended to decrease as experimental period extended. Total number of pathogens in feces showed peaks at 1~2 months after the initiation of experiment, and tended to decrease thereafter. Total number of Eimeria spp., E. coli and BVDV in feces showed no significant differences but control group showed higher counts than both treatment groups. During overall period, the incidence of pathogenic diarrhea in calves of 2 treatment groups was much lower than control group calves (C: 24, vs T1: 9, T2: 13 heads), however, it was not significant (p>0.05). In conclusion, supplementation of herbs in Hanwoo calf diets might be beneficial to improve growth performances and prevent diarrhea.

Analysis of the Change in Seropositive Rate of the Epstein-Barr Virus in Koreans: A Single-Center Study (한국인의 Epstein-Barr Virus 항체 양성률 변화양상에 대한 분석: 단일기관 연구)

  • Kim, Su Kyung;Choi, Joon-Sik;Kim, Dongsub;Kang, Cheol-In;Chung, Doo Ryeon;Peck, Kyong Ran;Kang, Eun-Suk;Kim, Yae-Jean
    • Pediatric Infection and Vaccine
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    • v.27 no.2
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    • pp.117-126
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    • 2020
  • Purpose: Epstein-Barr virus (EBV) infection is related to infectious mononucleosis or nasopharyngeal cancer, and its epidemiology may change according to the socioeconomic development of communities. This study aimed to evaluate the recent epidemiology of EBV seropositive rate in Korea. Methods: We retrospectively reviewed EBV serology test results obtained from a part of clinical care at Samsung Medical Center, Seoul, South Korea, from January 2000 to December 2017. Results: The EBV seropositive rate in 26,527 subjects during the study period was 81.0% (21,485/26,527): 44.4% (2,716/6,122) in subjects aged 0-9 years, 75.8% (2,077/2,739) in those aged 10-19 years, and 94.5% (16,692/17,666) in those aged ≥20 years. The EBV seropositive rate decreased from 89.4% (8,592/9,616) in 2000-2008 to 76.2% (12,893/16,911) in 2009-2017 (P<0.001). Especially, the EBV seropositive rate in subjects aged 0-19 years significantly decreased from 2000-2008 to 2009-2017 (0-9 years, 62.8% [1,172/1,866] in 2000-2008 and 36.3% [1,544/4,256] in 2009-2017; 10-19 years, 83.8% [745/858] in 2000-2008 and 70.8% (1,332/1,881) in 2009-2017) (P<0.001). Conclusions: The EBV seropositive rate in children has decreased in the last 20 years. As the age of patients with primary EBV infection increased, there is a need for interest in clinical manifestation, such as infectious mononucleosis, in adolescents and young adults.