Basic, Research : Production of Hepaptitis E Virus-like Particle (HEVLP) in Human Hepatoma Cell Line using Baculovirus-based Expression System

( Eun Byul Lee ),( Jung Eun Choi ),( Jung Hee Kim ),( Wonhee Hur ),( Tian Zhu Li ),( Sung Woo Kim ),( Sung Woo Hong ),( Young Ki Lee ),( Sung Min Kim ),( Joon Ho Lee ),( Seung Kew Yoon ) 대한간학회 2013 춘·추계 학술대회 (KASL) Vol.2013 No.1

Background: Hepatitis E virus (HEV) is a hepatotropic virus and has shown the ability of self-assembly through its N-terminally truncated ORF2 (Nt-ORF2). Recently, many researchers have attempted to use the virus like particles (VLPs) as a gene delivery vehicle. In this study, we established the liver specific delivery system using HEV-LP produced in Huh7 cells by recombinant baculovirus expression system. Methods: To produce HEV-LP in mammalian cells, the recombinant baculovirus containing the CMV promoter derived Nt-ORF2 gene was generated in the insect cells, Sf21. Subsequently, the baculovirus was used to infect into the human hepatoma cell line, Huh7 cells. The expression of Nt-ORF2 in Huh7 cells was confirmed by western blot analysis. In addition, the formation of HEV-LP particle was observed by electron microscopy. Next, to determine the liver-specific entrance of HEV-LP, FITC-conjugated HEV-LP infected into the cancer cell lines derived from various organs. After infection, the fluorescence existed in infected cells was detected by confocal microscopy. Results: Nt-ORF2 was highly expressed in Huh7 cells infected with recombinant baculoviruses. Nt-ORF2 expressed in the cells showed the ability of self-assembly and release from the cells. The HEV-LPs produced from Huh7 cells was observed 25 nm in diameter. Furthermore, HEV-LP could enter into the hepatoma cell lines within 4 hr post-infection. Conclusions: These results suggest that HEV-LP was efficiently produced and infected in hepatoma cells and established HEV-LP system could be used as the valuable liver specific transporter to deliver the therapeutic agents.

The Relationship between Smartphone Addiction and Quality of Sleep in Nursing Students

Jung-yeon Lee,Han-byul Yoo,Ye-eun Lee,Soo-hyun Lee,Jeong-eun Lee,Hyun-jung Lee,Su-jeong Lee 한국간호과학회 2019 한국간호과학회 학술대회 Vol.2019 No.10

MicroRNA-99a Attenuates HCV Replication through the Downregulation of Subtilisin Kexin Isozyme-1 (SKI-1) / Site-1 Protease (S1P)

( Eun Byul Lee ),( Seung Kew Yoon ),( Jung-hee Kim ),( Wonhee Hur ),( Sung Min Kim ),( Joon Ho Lee ) 대한간학회 2016 춘·추계 학술대회 (KASL) Vol.2016 No.1

Aims: MicroRNAs modulate various biological processes through dysfunction of target genes. Accumulating evidence indicates that a number of miRNAs can regulate or be regulated by hepatitis C virus (HCV) infection. Recently, it has been reported that expression level of miR-99a was inversely correlated with sustained virological response in patients with chronic hepatitis C. However, the exact role of miR-99a and its target in the pathogenesis of HCV infection remains unexplored. Here, we investigated the restrictive effects of miR-99a on the replication of HCV in relation to lipid metabolism and identified subtilisin kexin isozyme-1/site-1 protease (SKI1/S1P) as a novel target of miR-99a. Methods: The levels of miR-99a were evaluated in the serum of patients with chronic HCV infection, Huh7 cells infected with HCVcc and HCV full-genomic replicon (FGR) cells by miRNA quantitative real- time PCR (qRT-PCR). In addition, the effect of miR-99a-5p on HCV replication was analyzed by measuring the levels of HCV RNA after treatment with miR-99a-5p mimics in Huh7 cells infected with HCVcc and FGR cells. The levels of miR-99a target genes involved in lipid metabolism were also assessed by western blot analysis and qRT-PCR. The change of lipid accumulation by miR-99a-5p over-expression was quantitatively measured by Nile Red staining. Results: The expression level of miR-99a-5p was significantly down-regulated in sera from patients chronically infected with HCV compared to healthy subjects. Moreover, the expression level of miR-99a was decreased in both of FGR cells and Huh7 cells infected with HCVcc. Notably, treatment with miR-99a-5p mimics significantly down-regulated HCV replication. In this study, we identified SKI1/S1P as a novel target of miR-99a in relation to HCV replication. SKI1/S1P expression was significantly suppressed by treatment with miR-99a-5p mimics in HCV replicating cells. Regarding the role of SKI1/S1P in lipogenesis, the forced expression of miR-99a-5p attenuated the increase in the amount of intracellular lipid droplets after oleic acid treatment. Conclusions: Our data provide new mechanistic insights of role of miR-99a as an anti-viral host factor on HCV replication by suppression of lipid accumulation through down-regulated of SKI1/S1P. This research was supported by Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Science, ICT & Future Planning (2015R1C1A1A020 37212)

Evaluation of Circulating MicroRNA Biomarkers in the Acute Pancreatic Injury Dog Model

Lee, Han-Byul,Park, Hyun-Kyu,Choi, Hyun-Ji,Lee, Sora,Lee, Sang-Joon,Lee, Ji-Young,Cho, Eun-Ho,Han, Hyo-Jeong,Seok, Ju-Hyung,Son, Woo-Chan MDPI 2018 INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES Vol.19 No.10

<P>This study aimed to evaluate the usefulness of four microRNAs (miRNAs) in an acute pancreatic injury dog model. Acute pancreatitis was induced by infusion of cerulein for 2 h (7.5 μg/kg/h). The levels of well-known miRNAs, microRNA-216a (miR-216a) and microRNA-375 (miR-375), and new candidates microRNA-551b (miR-551b), and microRNA-7 (miR-7), were measured at 0, 0.5, 1, 2, 6, 12, and 24 h with serum amylase and lipase, and histopathological examination was performed. Among the four miRNAs, miR-216a and miR-375, and serum enzymes were significantly increased by cerulein treatment. The expression levels of miRNAs and serum enzymes peaked at 2–6 h with a similar pattern; however, the overall increases in miR-216a and miR-375 levels were much higher than those of the serum enzyme biomarkers. Increased levels of miR-216a and miR-375 were most highly correlated to the degree of individual histopathological injuries of the pancreas, and showed much greater dynamic response than serum enzyme biomarkers. Twenty-four-hour time-course analysis in this study revealed time-dependent changes of miRNA expression levels, from initial increase to decrease by predose level in acute pancreatitis. Our findings demonstrate that, in dogs, miR-216a and miR-375 have the potential to sensitively detect pancreatitis and reflect well the degree of pancreatic injury, whereas miR-551b and miR-7 do not.</P>

Clutter Suppression Method of Iron Tunnel Using Cepstral Analysis for Automotive Radars

LEE, Han-Byul,LEE, Jae-Eun,LIM, Hae-Seung,JEONG, Seong-Hee,KIM, Seong-Cheol 'Institute of Electronics, Information and Communi 2017 IEICE TRANSACTIONS ON COMMUNICATIONS - Vol.100 No.2

3,4,5-Tricaffeoylquinic Acid Inhibits the Lipopolysaccharide-Stimulated Production of Inflammatory Mediators in Keratinocytes

Lee, Seon Ae,Jung, Eun Byul,Lee, Seon Hwa,Kim, Yun Jeong,Bang, Hyoweon,Seo, Seong Jun,Choi, Young Wook,Kim, Manh Heun,Lee, Min Won,Lee, Chung Soo S. Karger AG 2012 Pharmacology Vol. No.

<P>Abstract</P><P><B><I>Background and Purpose:</I></B> Microbial product lipopolysaccharide (LPS) has been shown to be involved in the pathogenesis of inflammatory skin diseases. Caffeoyl derivatives have demonstrated anti-inflammatory and antioxidant effects. However, the effect of 3,4,5-tricaffeoylquinic acid (3,4,5-triCQA) on the production of microbial product-induced inflammatory mediators in keratinocytes has not yet been studied. <B><I>Experimental Approach:</I></B> Using human keratinocytes, we investigated the effect of 3,4,5-triCQA on the LPS-stimulated production of inflammatory mediators in relation to the nuclear factor (NF)-κB, Akt and ERK pathways. <B><I>Results:</I></B> 3,4,5-triCQA inhibited the LPS-induced expression of Toll-like receptor 4, and the production of cytokines and chemokines in keratinocytes. 3,4,5-triCQA, Bay 11-7085, Aκt inhibitor and ERK inhibitor each attenuated the LPS-induced production of inflammatory mediators by inhibiting the NF-κB, Akt and ERK pathways.<B><I> Conclusions and Implications:</I></B> 3,4,5-triCQA may attenuate the LPS-stimulated production of inflammatory mediators in keratinocytes by suppressing the Toll-like receptor 4 expression-mediated activation of the Akt, ERK and NF-κB pathways. 3,4,5-triCQA may exert a preventive effect against microbial product-induced inflammatory skin diseases.</P><P>Copyright © 2012 S. Karger AG, Basel</P>

NDRG2-mediated Modulation of SOCS3 and STAT3 Activity Inhibits IL-10 Production

Lee, Eun-Byul,Kim, Ae-Yung,Kang, Kyeong-Ah,Kim, Hye-Ree,Lim, Jong-Seok The Korean Association of Immunobiologists 2010 Immune Network Vol.10 No.6

Background: N-myc downstream regulated gene 2 (NDRG2) is a member of the NDRG gene family. Our previous report indicated a possible role for NDRG2 in regulating the cytokine, interleukin-10 (IL-10), which is an important immunosuppressive cytokine. Several pathways, including p38-MAPK, NF-${\kappa}B$, and JAK/STAT, are used for IL-10 production, and the JAK/STAT pathway can be inhibited in a negative feedback loop by the inducible protein, SOCS3. In the present study, we investigated the effect of NDRG2 gene expression on IL-10 signaling pathway that is modulated via SOCS3 and STAT3. Methods: We generated NDRG2-overexpressing U937 cell line (U937-NDRG2) and treated the cells with PMA to investigate the role of NDRG2 in IL-10 production. U937 cells were also transfected with SOCS3- or NDRG2-specific siRNAs to examine whether the knockdown of SOCS3 or NDRG2 influenced IL-10 expression. Lastly, STAT3 and SOCS3 induction was measured to identify the signaling pathway that was associated with IL-10 production. Results: RT-PCR and ELISA assays showed that IL-10 was increased in U937-mock cells upon stimulation with PMA, but IL-10 was inhibited by overexpression NDRG2. After PMA treatment, STAT3 phosphorylation was decreased in a time-dependent manner in U937-mock cells, whereas it was maintained in U937-NDRG2 cells. SOCS3 was markedly reduced in U937-NDRG2 cells compared with U937-mock cells. IL-10 production after PMA stimulation was reduced in U937 cells when SOCS3 was inhibited, but this effect was less severe when NDRG2 was inhibited. Conclusion: NDRG2 expression modulates SOCS3 and STAT3 activity, eventually leading to the inhibition of IL-10 production.

Establishment of Delivery System Targeting Liver Using Hepatitis E Virus-Like Particles

Eun Byul Lee,Jung-Hee Kim,Jung Eun Choi,Seung Kew Yoon 한국간담췌외과학회 2014 한국간담췌외과학회 학술대회지 Vol.2014 No.4

Continuous Transforming Growth Factor β₁Secretion by Cell-Mediated Gene Therapy Maintains Chondrocyte Redifferentiation

Lee, Dug Keun,Choi, Kyoung Baek,Oh, In Suk,Song, Sun U.,Hwang, Sally,Lim, Chae-Lyul,Hyun, Jong-Pil,Lee, Hyeon-Youl,Chi, Guang Fan,Yi, Youngsuk,Yip, Vivian,Kim, Jeannie,Lee, Eun Byul,Noh, Moon Jong,Lee Mary Ann Liebert 2005 Tissue engineering Vol.11 No.1

<P>One of the most important factors in the production of cartilage is transforming growth factor beta1 (TGF-beta1). To obtain sustained release of TGF-beta1, a cell-mediated gene therapy technique was introduced. We infected chondrocytes with a retroviral vector carrying the TGF-beta1 gene. The single clone derivative showed sustained TGF-beta1 secretion. It also showed constitutive type II collagen expression. Whereas the TGF-beta1 protein itself is unable to induce formation of cartilage in vivo, human chondrocytes engineered to express a retroviral vector encoding TGF-beta1 showed cartilage formation in vivo when cells were injected into nude mice intradermally. These data suggest that cell-mediated gene therapy using TGF-beta1 as a transgene would be a promising treatment for osteoarthritis.</P>

A Study on the Rationalization of Criteria for Facilities Handling Toxic Chemicals in Consideration of the Threshold Quantity

Eun Byul Lee(이은별),Byung Tae Yoo(유병태) 위기관리 이론과 실천 2018 Crisisonomy Vol.14 No.9

2015년 화학물질관리법이 전면 개정됨에 따라 화학사고의 예방 및 대응을 위한 다양한 제도가 신설 되었고, 특히 유해화학물질 취급시설의 설치 및 관리기준에 상당히 강화되었다. 이를 계기로 사업장의 안전관리가 강화되어 사고예방 효과도 있었지만, 일부 기준들은 취급량 및 입지조건 등을 고려하지 않고 일괄적으로 적용되고 있어 산업계에서는 어려움을 호소하고 있는 실정이다. 본 연구에서는 이러한 기준에 대해 보다 현실적이면서 합리적인 방안을 마련하고자 하였다. 이를 위하여 국내·외 유사 화학물질 안전관리제도를 비교⋅분석하였으며 화학물질 취급시설 이해관계자 100명을 대상으로 설문조사를 실시하였다. 설문 결과를 바탕으로 화학물질 취급시설 설치 및 관리 기준을 산업단지 입주여부와 장외영향평가 소량취급 여부를 구분하여 차등적용 및 관리 할 수 있는 합리적인 방안을 제시하였다. The Korean government revised the Chemical Control Act in 2015 to promote chemical safety, particularly enhancing the standards for the installation and management of chemical-handling facilities. Although it pro-vided a significant accident prevention effect due to the stronger workplace safety management, industry has been complaining that some standards are applied universally without considering specific chemical usages and locations. In this study, we attempted to provide a more rational approach to these standards by comparing and analyzing the similar safety management systems around the world. We also conducted a survey of 100 stakeholders in chemical-handling facilities. Based on the findings, we proposed a reasonable method to apply installation and management standards of chemical handling facilities, depending on whether or not they are located within an industrial complex and they handle only a small quantity of ORA.