• 한국어병학회지
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• 제13권1호
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• pp.7-13
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• 2000

1993년부터 한국 서해안의 새우 양식장에서는 양식새우의 대량폐사가 일어났다. 외부증상은 두흉갑과 표피에 흰 반점이 나타났고 어류주화세포에는 배양되지 않았다. 열($50^{\circ}C$) 및 강산(pH 3)에는 쉽게 실활되었으나 강알카리(pH 11)에는 내성이 강했다. 바이러스의 입자 형태는 Rod Shaped한 형태를 보였다. 바이러스 단백질의 분석결과는 Hypodermal Hematopoietic Necrosis Baculovirus(HHNBV)와 유사했고 바이러스 핵산분석 결과는 약 114kb로 Penaeid Acute Viremia(PAV)와 유사했다.

• Molecules and Cells
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• 제38권2호
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• pp.122-129
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• 2015

DBM-2198, a six-membered azasugar nucleotide (6-AZN)-containing phosphorothioate (P = S) oligonucleotide (AZPSON), was described in our previous publication [Lee et al. (2005)] with regard to its antiviral activity against a broad spectrum of HIV-1 variants. This report describes the mechanisms underlying the anti-HIV-1 properties of DBM-2198. The LTR-mediated reporter assay indicated that the anti-HIV-1 activity of DBM-2198 is attributed to an extracellular mode of action rather than intracellular sequence-specific antisense activity. Nevertheless, the antiviral properties of DBM-2198 and other AZPSONs were highly restricted to HIV-1. Unlike other P = S oligonucleotides, DBM-2198 caused no host cell activation upon administration to cultures. HIV-1 that was pre-incubated with DBM-2198 did not show any infectivity towards host cells whereas host cells pre-incubated with DBM-2198 remained susceptible to HIV-1 infection, suggesting that DBM-2198 acts on the virus particle rather than cell surface molecules in the inhibition of HIV-1 infection. Competition assays for binding to HIV-1 envelope protein with anti-gp120 and anti-V3 antibodies revealed that DBM-2198 acts on the viral attachment site of HIV-1 gp120, but not on the V3 region. This report provides a better understanding of the antiviral mechanism of DBM-2198 and may contribute to the development of a potential therapeutic drug against a broad spectrum of HIV-1 variants.

• 대한화장품학회지
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• 제47권2호
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• pp.171-178
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• 2021

본 연구에서는 프로바이오틱스 유래 엑소좀 유사 나노베지클을 분리하고, 피부에 대한 여러 가지 생리활성을 평가했다. 프로바이오틱스의 한 종인 Lactococcus lactis subsp. lactis (LL)를 배양하고 고압균질기와 한외여과를 통해 70 ~ 200 nm 크기를 갖는 LL 유래 엑소좀 유사 나노베지클(LVs)을 분리했다. 나노입자추적분석 결과 1.81 × 10¹¹ particles/mL로 나타났다. LVs를 섬유아세포와 피부각질세포에 처리하여 피부 주름과 장벽 개선과 관련된 효능을 확인했다. 우선 섬유아세포에서 fibrillin (FBN1) 유전자 발현량이 23%, 피부각질세포에서 fibronectin (FN1)과 filaggrin (FGN) 유전자 발현량이 각각 65%, 400% 증가했다. 그리고 각질형성능은 대조군 대비 30% 증가함을 확인할 수 있었다. 또한, UV 조사한 피부각질세포에 LVs를 처리했을 때 collagen type I alpha 1 (COL1A1)이 대조군 대비 약 83% 증가하는 결과를 보여주었다. 이로써 프로바이오틱스 유래 엑소좀 유사 나노베지클은 장벽 개선과 관련하여 화장품 및 의약품 소재로 이용할 수 있음을 확인했다.

• 대한수의학회지
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• 제37권4호
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• pp.883-897
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• 1997

Present study was undertaken in order to find out the most significant laboratory test for the early diagnosis of ethylene glycol poisoning in dogs. Seven dogs weighing from 5.0kg to 19.3kg were administered orally 6ml or l0ml of ethylene glycol per kilogram of body weight. In addition to clinical observations, blood samples were taken from the jugular vein on 0, 1, 3, 6, 12, 24 and 48 hours or until death after the adminisration and examined for the hematological, blood chemical, venous blood gas and electrolyte changes. Urine samples were simultaneously taken and examined for pH, protein, glucose and calcium oxalate crystals in the sediments. The results were as follows : 1. All dogs showed marked hypothermia in addition to the characteristic signs of ethylene glycol poisoning until death. 2. No significant hematological changes were observable after the administration of ethylene glycol except the secondary dehydration and stress leukogram. 3. Chemical values related to the liver function(serum glutamic pyruvic transaminase, total serum protein, serum albumin, serum total bilirubin) showed no significant changes during the experimental period in all dogs. 4. Chemical values related to the renal function(BUN and creatinine) showed significant(p<0.0l, p<0.001) increase from 24 hours to death in all dogs. 5. All dogs showed significant(p<0.01, p<0.001) decrease in blood pH from 3 hours to death after the ethylene glycol administration. 6. All dogs showed significant(p<0.05, p<0.01, p<0.001) increaes in anion gap from 1 hour to death after the ethylene glycol administration. 7. All dogs showed significant(p<0.05, p<0.01) decrease in urine pH from 1 hour to 6 hours after the administration of ethylene glycol. 8. Characteristic envelope form of calcium oxalate crystals were found in the urinary sediment from 6 hours after the administration of ethylene glycol in all dogs. 9. Histologically calcium oxalate crystals were found in the renal rubles in all dogs. 10. From these results, it may be concluded that the examinations of urinary sediments for the calcium oxalate crystals, blood or urinary pH and blood anion gap are most reliable tests for the early diagnosis of ethylene glycol poisoning in dogs.

• Journal of Microbiology and Biotechnology
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• 제31권3호
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• pp.358-367
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• 2021

The World Health Organization (WHO) has declared the coronavirus disease 2019 (COVID-19) as an international health emergency. Current diagnostic tests are based on the reverse transcription-quantitative polymerase chain reaction (RT-qPCR) method, which is the gold standard test that involves the amplification of viral RNA. However, the RT-qPCR assay has limitations in terms of sensitivity and quantification. In this study, we tested both qPCR and droplet digital PCR (ddPCR) to detect low amounts of viral RNA. The cycle threshold (CT) of the viral RNA by RT-PCR significantly varied according to the sequences of the primer and probe sets with in vitro transcript (IVT) RNA or viral RNA as templates, whereas the copy number of the viral RNA by ddPCR was effectively quantified with IVT RNA, cultured viral RNA, and RNA from clinical samples. Furthermore, the clinical samples were assayed via both methods, and the sensitivity of the ddPCR was determined to be equal to or more than that of the RT-qPCR. However, the ddPCR assay is more suitable for determining the copy number of reference materials. These findings suggest that the qPCR assay with the ddPCR defined reference materials could be used as a highly sensitive and compatible diagnostic method for viral RNA detection.

• 한국미생물·생명공학회지
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• 제37권4호
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• pp.377-382
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• 2009

사람과 동물 유래의 혈장, 세포, 조직 등을 이용하여 생물의약품을 생산하기 위해서는 바이러스 안전성 확보가 필수적이다. 바이러스 안전성 보증을 위해 생물의약품 제조공정은 바이러스 불활화/제거 단계를 포함하여야 한다. 짧은 파장자외선(UVC) 조사는 바이러스 불활화 효과가 매우 높은 것으로 알려졌지만, UVC 조사로 인한 단백질의 변성과 대상 물질에 동일하게 조사를 할 수 있는 기계적 장치 개발의 어려움으로 인해 UVC 조사는 생물의약품 제조 공정에 사용되지 못했다. 최근에 이러한 결점을 해결한 연속 유동 UVC 반응기(UVivatec)가 개발되었다. UVivatec의 바이러스 불활화 효과 및 단백질 회수율을 검증하기 위해 단백질 의약품을 대상으로 적용가능성을 조사하였다. 최적화된 $3,000\;J/m^2$ 조사 공정에서 단백질의 회수율은 98%이상이었다. UVC 조사에 의한 human immunodeficiency virus(HIV), hepatitis A virus(HAV), bovine herpes virus(BHV), bovine viral diarrhea virus(BVDV), porcine parvovirus(PPV), bovine parvovirus(BPV), minute virus of mice(MVM), reovirus type 3(REO), bovine parainfluenza virus type 3(BPIV) 불활화 효과를 평가하였다. HAV, PPV, BPV, MVM, REO와 같은 비외피(nonenvelope) 바이러스는 $3,000\;J/m^2$ 조사량에 의해 검출한계 이하로 완벽하게 불활화되었다. HIV, BVDV, BPIV 같은 외피(envelope) 바이러스도 $3,000\;J/m^2$ 조사량에 의해 검출한계 이하로 완벽하게 불활화되었다. 또한 BHV도 매우 민감하게 불활화되었다. UVC 조사에 의한 각 바이러스들의 로그 감소율은 HIV는 ${\geq}3.89$, HAV는 ${\geq}5.27$, BHV는 5.29, BVDV는 ${\geq}5.96$, PPV는 ${\geq}4.37$, BPV는 ${\geq}3.55$, MVM은 ${\geq}3.51$, REO는 ${\geq}4.20$, BPIV는 ${\geq}4.15$이었다. 이와 같은 결과에서 UVivatec을 이용한 UVC 조사는 바이러스 불활화에 매우 효과적인 방법임을 확인하였다.

• Journal of Microbiology
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• 제42권1호
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• pp.25-31
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• 2004

Sf9 cells have obvious advantages for the conventional production technology of vaccine. They are useful tools for high concentration and large-scale cultures. Sf9 cells were grown to maximal concentration, 8${\times}$l0$\^$6/ cells/$m\ell$ in a 500$m\ell$ spinner flask, with a doubling time at the exponentially growing phase of 24.5 hours, using serum-free media. To explore the ability of Sf9 cells to be infected by the Japanese encephalitis (JE) virus Beijing-l strain, Sf9 cells were infected with the virus. By 4-5 days post-infection, 10-15 % of the Sf9 cells showed cytopathic effect (CPE), from granularity to the formation of syncytia and multinucleated giant cells continuously observed over a period of 35 days. Positive fluorescent reactions were detected in 30-40% of cells infected with the JE virus Beijing-l strain, and the uninfected Sf9 cells were completely negative. Virus particles, propagated in Sf9 and Vero cells, were concentrated by sedimentation on 40% trehalose cushions by ultracentrifugation, and showed identical patterns of viral morphogenesis. Complete virus particles, 40 to 50 nm in diameter, were observed, and JE virus envelope (E) proteins, at 53 kDa, were found in the western blot analysis to the anti-JE virus E protein monoclonal antibody and reacted as a magenta band in the same position to the glycoprotein staining. To evaluate whether the infectious virus was produced in Sf9 cells inoculated with the JE virus Beijing-l stain, Sf9 cells were inoculated with the virus, and sample harvested every 5 days. The titers of the JE virus Beijing-l strain rose from 1.0${\times}$l0$\^$5/ to 1.5${\times}$l0$\^$6/ pfu/$m\ell$. The infected Sf9 cells could be subcultured in serum-free medium, with no change in the plaque sizes formed by the JE virus Beijing-l strain in the plaque assay. It is suggested that the ability of the JE virus Beijing-l strain to infect Sf9 cells in serum-free media will provide a useful insect cell system, where the JE virus replication, cytopathogenicity and vaccine immunogen can be studied.

• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 1996년도 춘계학술대회
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• pp.19-29
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• 1996

The neurological manifestations of AIDS include dementia, encountered even in the absence of opportunistic superinfection or malignancy. The AIDS Dementia Complex appears to be associated with several neuropathological abnormalities, including astrogliosis and neuronal injury or loss. How can HIV-1 result in neuronal damage if neurons themselves are only rarely, if ever, infected by the vitus\ulcorner In vitro experiments from several different laboratiories have lent support to the existence of HIV- and immune-related toxins. In one recently defined pathway to neuronal injury, HIV-infected macrophages/microglia as well as macrophages activated by HIV-1 envelope protein gp120 appear to secrete excitants/neurotoxins. These substances may include arachidonic acid, platelet-activating factor, free radicals (NO - and O$_2$), glutamate, quinolinate, cysteine, cytokines (TNF-${\alpha}$, IL1-B, IL-6), and as yet unidentified factors emanating from stimulated macrophages and possibly reactive astrocytes. A final common pathway for newonal suscepubility appears to be operative, similar to that observed in stroke, trauma, epilepsy, and several neurodegenerative diseases, including Huntington's disease, Parkinson's disease, and amyotrophic lateral sclerosis. This mechanism involves excessive activation of N-methyl-D-aspartate (NMDA) receptor-operated channels, with resultant excessive influx of Ca$\^$2+/ leading to neuronal damage, and thus offers hope for future pharmacological intervention. This chapter reviews two clinically-tolerated NMDA antagonists, memantine and nitroglycerin; (ⅰ) Memantine is an open-channel blocker of the NMDA-associated ion channel and a close congener of the anti-viral and anti-parkinsonian drug amantadine. Memantine blocks the effects of escalating levels of excitotoxins to a greater degree than lower (piysiological) levels of these excitatory amino acids, thus sparing to some extent normal neuronal function. (ⅱ) Niuoglycerin acts at a redox modulatory site of the NMDA receptor/complex to downregulate its activity. The neuroprotective action of nitroglycerin at this site is mediated by n chemical species related to nitric oxide, but in a higher oxidation state, resulting in transfer of an NO group to a critical cysteine on the NMDA receptor. Because of the clinical safety of these drugs, they have the potential for trials in humans. As the structural basis for redox modulation is further elucidated, it may become possible to design even better redox reactive reagents of chinical value. To this end, redox modulatory sites of NMDA receptors have begun to be characterized at a molecular level using site-directed mutagenesis of recombinant subunits (NMDAR1, NMDAR2A-D). Two types of redox modulation can be distinguished. The first type gives rise to a persistent change in the functional activity of the receptor, and we have identified two cysteine residues on the NMDARI subunit (#744 and #798) that are responsible for this action. A second site, presumably also a cysteine(s) because <1 mM N-ethylmaleimide can block its effect in native neurons, underlies the other, more transient redox action. It appears to be at this, as yet unidentified, site on the NMDA receptor that the NO group acts, at least in recombinant receptors.

• Journal of Microbiology and Biotechnology
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• 제25권4호
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• pp.537-546
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• 2015

Classical swine fever (CSF) is a highly contagious disease of pigs caused by CSF virus (CSFV). E2 is the major viral envelope protein of immune dominance that induces neutralizing antibodies and confers protection against CSFV infection. The B/C domains of E2 are variable among CSFV isolates, which could affect immunogenicity and binding to antibodies. We attempted to characterize the epitope recognized by a monoclonal antibody 2B6 (mAb-2B6) raised against the E2 B/C domains of the vaccine C-strain and to examine if mutations in the epitope region would affect antibody binding and viral neutralization. The epitope specific for mAb-2B6 recognition is linear, spanning five residues ⁷⁷⁴DGXNP⁷⁷⁸ in the B/C domains. The residue N777 is indispensable for the specificity. The epitope exists only in group 1 strains, but not in those of group 2. The recombinant viruses containing individual mutations on the epitope region lost the reactivity to mAb-2B6. The mutant virus RecC-N777S had low replication potential, about 10-fold decrease in the yield of progeny virus particles, whereas the mutant virus RecC-P778A reverted to proline upon continuous passaging. The mutations on the mAb-2B6 epitope region did not affect neutralization by anti-C-strain polyclonal sera from pigs. Deletion from aa774 covering the mAb-2B6 epitope, but not that from aa781, also affected binding with the polyclonal antibodies from vaccinated pigs, although the major binding region for the vaccinated antibodies is aa690-773.

• Journal of Microbiology and Biotechnology
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• 제28권8호
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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.