Portable vibration-assisted filtration device for on-site isolation of blood cells or pathogenic bacteria from whole human blood

Kim, Yong Tae,Park, Kyun Joo,Kim, Seyl,Kim, Soon Ae,Lee, Seok Jae,Kim, Do Hyun,Lee, Tae Jae,Lee, Kyoung G. Elsevier 2018 Talanta Vol.179 No.-

<P><B>Abstract</B></P> <P>Isolation of specific cells from whole blood is important to monitor disease prognosis and diagnosis. In this study, a vibration-assisted filtration (VF) device has been developed for isolation and recovery of specific cells such as leukocytes and pathogenic bacteria from human whole blood. The VF device is composed of three layers which was fabricated using injection molding with cyclic olefin copolymer (COC) pellets consisting of: a top layer with coin-type vibration motor (Ф = 10mm), a middle plate with a 1μm or 3μm-pore filter membrane to separate of <I>Staphylococcus aureus</I> (<I>S. aureus</I>) cells or leukocytes (i.e. white blood cells) respectively, and a bottom chamber with conical-shaped microstructure. One milliliter of human whole blood was injected into a sample loading chamber using a 3μm-pore filter equipped in the VF device and the coin-type vibration motor applied external vibration force by generating a rotational fluid which enhances the filtration velocity due to the prevention of the cell clogging on the filter membrane. The effluent blood such as erythrocytes, platelet, and plasma was collected at the bottom chamber while the leukocytes were sieved by the filter membrane. The vibration-assisted leukocyte separation was able to finish within 200s while leukocyte separation took 1200s without vibration. Moreover, we successfully separated <I>S. aureus</I> from human whole blood using a 1μm-pore filter equipped VF device and it was further confirmed by genetic analysis. The proposed VF device provides an advanced cell separation platform in terms of simplicity, fast separation, and portability in the fields of point-of-care diagnostics.</P> <P><B>Highlights</B></P> <P> <UL> <LI> A vibration-assisted filtering device (VF) was developed for isolation of specific cells. </LI> <LI> 6-fold faster separation of leukocytes has been accomplished. </LI> <LI> Pathogenic bacteria can be successfully recovered from human blood. </LI> <LI> The VF device provides a cell isolation platform for on-site disease diagnostics. </LI> </UL> </P> <P><B>Graphical abstract</B></P> <P>[DISPLAY OMISSION]</P>

랫드 유선암에 대한 면역치료 방법 개발

정자영,이국경,염영나,이종근,김현배,김세일,강호일,이영순,김옥희 식품의약품안전청 1997 식품의약품안전청 연보 Vol.1 No.-

This study was enamined on the effect of adoptive immunotherapy in T,12-dimethylbena la]aathracene(B?BA) -in13uce6 rat mammary carcinogenesis. Flfty- sik-day-old female Sprague-Dawley 40 rats were divided into four groups. fLs a positive controtp animals in Group I were intubated with DMBA. Smg/100g body weight and single dose, at onset of experimeat. Group ll was treatedadoptive iinrnunotherapy with polyinosinic- polycytidylic acid(Poly I:C) and Group 111 was treated withInterleultin(IL-2). Group nr is negative control, therefore no treated group. All animals were sacrificedat 16 weeks after DMBA in.tubation. Mammary gland wet weight, dry fat free tissue weight, incidence oftumor, and number of lobules, alveolar buds, terminal end buds, ftnd terminal ducts were examined,Histopathogenesis of DMBA-iudvced rat mammary tumors were performed by whole mount method androutine histologic technique. As a result, the induced mammary tumors of Group I was 60%. The effectof adoptive irnrnunotherapy in DMBA-induced rat mammary carcin☞genesis of Group rl and ITI were 33% and 0%, respectively. Histopathological types of induced-mammary tumors were adeaorna,adenocarciaoma and carcinosarcoma. Morphological changes of the mammary gland after treated withDMBA was analysed by whole mount and histopathological methul- The number of the terminal endbuds and_terminal ducts are significantly higer in DMBA treated Groups(p<0.01) than non-treatedGroup ini fbservation of wllole mount method. Tn microscopy observation, lobules containing alveolarbuds and hyperplastic alueolar nodules were significantly higher in DMBA treated Groups(p<0.01)than aon- treated Group. Especially, hyperplastic alveolar nodules is very sensitive parameter to assessthe mamraary carcinogenesis and the effect of adoptive immunotherapy.

B16 흑색종 세포에서 분리 동정된 암전이 관련 유전자의 기능에 관한 연구

염영나,김세일,황명실,이국경,주종호,조현영,강호일,김옥희 식품의약품안전청 2000 식품의약품안전청 연보 Vol.4 No.-

암전이로 많은 암환자가 사망한다. 암전이는 신생혈관형성, 침윤, 세포부착 등의 세부과정이 단계져이고 복합적으로 진행괵는페, 이러한 과정을 이해하가 위할 동물 모델 시스템으로 816 흑색종을 이용한 in uf'vo metastasis assay가 사용되고 있다. 전이능력 차이가 있는 Bl6 흑색종을 DDRT-PC료으로 nIRNA 발현차이를 』:이는 VASP와 Iysosom리 sialidase를 확인한 바 있어, mRNA 발현 차이가 실지로 단백질 수준에서 변화가 있는지 또한 VASP 과발찬이 전이활성에 어떠한 영향을 보이는지 조사하였다. 전이능의 차이를 보.이는 Bl6 흑색종 세포(Bl6-Fl, Bl6-FIO, Bl6-BL6, Bl6-B16fung)에서 VASP과 Iysosomal sialidase의 단백질 수준에서의 발현정도를 VASP의 경우 westerri blotting으로 fysosomal sialidaset· 활성측정 방법으로 분석한 결과에서 큰 차이를 보이지 않았고 전이능력과 삶관 관계를 보이지 않았다. Bl6-Fl세포와 MCF7에 tFASP 유전자를 transfection시켜 침윤 정도를 조사한 결과 VfISP의 과발현으로 matrigel과 type R「 collagen에 대한 adhesion예는 차이를 없었고, 침윤 활성의 감소 효과를 보여 줌에 따라 VASP 단백질이 세포 이동과 침운에 과여하고 있음을 입즛하였다. Tumor metastasis is the leading cause of death in cancer patients. The process ef cancer metastasis consists of linked sequential steps, incEuding invasion, detachment,intravasation, circulation, adhesion, extravasation, and growth in distant organs. In viuoexperimental metastasis assay using Bl6 murine melanoma is generally used to investfgatemetastasis process. In the previous study of the DDRT-PCR of Bl6 metastatic varients havingthe same genetic backgrourid, we suggested that the VASP and Iysosomal sialidase , haddifferential expression in the mRNA level. In this study, we compared the protein expressionlevel of VASP using theTmetilod of western blotting and Iysosomal sialidase activity in the Bl6metastatic varients which arrf Bl6-Fl, Bl6-fIO, Bl6-B36, and Bl6-316/tung. The VASP andIysosomal silaidase have no efference in the protein expression levels of the Bl6 metas·taticvarients and have no correlation with metastatic potential of the Bl6 metastatic varients. Andthe VASP-transfected Bl6-F.1 cell have less Invassive activity than control cell and have nodiffernce in the adhesion to Matrigel and type R# collagen. This result suggests that VASPoverexpression can regulate ffle cell motility and invasion.

Quantification and visualization of metastatic lung tumors in mice

Lee Ha Neul,Kim Seyl,Park Sooah,Jung Woonggyu,Kang Jin Seok 한국독성학회 2022 Toxicological Research Vol.38 No.4

Histopathological examination is important for the diagnosis of various diseases. Conventional histopathology provides a two-dimensional view of the tissues, and requires the tissue to be extracted, fixed, and processed using histotechnology techniques. However, there is an increasing need for three-dimensional (3D) images of structures in biomedical research. The objective of this study was to develop reliable, objective tools for visualizing and quantifying metastatic tumors in mouse lung using micro-computed tomography (micro-CT), optical coherence tomography (OCT), and field emission-scanning electron microscopy (FE-SEM). Melanoma cells were intravenously injected into the tail vein of 8-week-old C57BL/6 mice. The mice were euthanized at 2 or 4 weeks after injection. Lungs were fixed and examined by micro-CT, OCT, FE-SEM, and histopathological observation. Micro-CT clearly distinguished between tumor and normal cells in surface and deep lesions, thereby allowing 3D quantification of the tumor volume. OCT showed a clear difference between the tumor and surrounding normal tissues. FE-SEM clearly showed round tumor cells, mainly located in the alveolar wall and growing inside the alveoli. Therefore, whole-tumor 3D imaging successfully visualized the metastatic tumor and quantified its volume. This promising approach will allow for fast and label-free 3D phenotyping of diverse tissue structures.

항암제가 apoptosis에 미치는 영향

염영나,김세일,이국경,정자영,강호일,김옥희,이경태,길광섭 식품의약품안전청 1997 식품의약품안전청 연보 Vol.1 No.-

녹차 등에 의한 Peroxisome Proliferator-activated Receptor Transactivation의 변화 연구

이국경,조현영,주종호,김세일,황명실,염영나,강호일,김옥희 식품의약품안전청 2000 식품의약품안전청 연보 Vol.4 No.-

녹차는 동서양에서 오래 전부터 즐겄 마션오던 차이다. 암 예방효과가 있다고 여겨진 녹차가 랫드에서 과산화소체의 수와 관련효소인 paluitoyl CoA oxydase(PCoA)의 활성을 증가시킨다고 최근에 보고죄었다. 본 연군에서는 녹차에 의한 과산화소체의 증가간 peroxisome pro넓craters-activated receptor a (PPARO)의 활성을 통하여 일어나는 가를 알아보기 위하여 transient transfection assay법을 이용하였다. Cos-1 세포에 PP·ARu를 transfection 시킨 후 녹차, 홍차, 우롱차의 추출액과 녹차의 주성분인 epigallocatechin gallate (EGCG), epigallate catechin (EGC), epicatechin gallate (ECG),epicatechin (EC), gallic acid를 투여하였다. 또한 강력한 과산화소체 증가물질인 Wy-14,643과의 상호작용을 관찰하기 위하여 Wy-씨,643에 차 추출액 또는 성분들을 병용투여하였다. 녹차와 홍차 추출액과 EGCG(lOoM), EGC는 대조판에 비하여 PPARa의 활성을 1.5-2배 정도 증가시켰으며 차의 종류별로는 륵차>흥차>우롱차』둥글레차의 순으로 PPARC를 활성시켰다. 병용투여의 경우, 녹차는 Wy-14,643에 의한 PPARa의 활썽을 4-6배정도까지 강력하게 증가시켰다. 따라서 녹차는 PPARa에 직접적으로 작용하여 과산화소첸의 수를 증가시키는 것 이외에, 알려지지 않은 기전을 통하여 PPARa 의 활성조절에 관여하는 것으로 여겨진다. 또한 PPAaa의 찰성이 중간발효산물인 우롱차의 경우에는 낮은 반면 완전발효산물인 흥차에서 다시 높아지는 결과는 EGCG 이외에도 PPARa의 활성에 영향을 미치는 또 다른 물질이 존재함을 쩨시하고 있다. Tea is a number of peroxisomesnITnlber of oeroxisomespopular beverage. Recently it was reported that green tea increases the in rats. In this stfdy, to know that the rrlechanism to increase theanri activities of neroxilomal pnfvmpl il pxprrrf ihrourh activation of peroxisome proliferator-activated receptor u(PPARa), Cell trallsient transfection studies were carried out to investigate the interactions of tea extracts and major tea components with PPARa, cloned frorrl moust. Green tea and black tea extract, and epigallocatechlh gallate (ECCG), amajor component of fresh green tea leaves, slightly increased 1.5-2 times of the activatioR ofPPARC compared wiff DMEitf alone treatment. Especially the effective doses of tea extractsand EGCG were lower than doses necessary to EGCG-induced cell death and inhibition ofmetastasis by inhibiting urokinase activity In ether reports. Also at these concentrations, gieentea and bfack tea extracts enhanced the activation of PPARa induced by Wy-14,643(20 r M) ina dose dependent Tnanner. But even though oolong tea is semi-fermented during themanufacturing process, oolong tea extract showed less active than black tea_ These resvltsdemonshate that green tea possesses an unknown potent regulatory role in activation of PPARO byperoxisoine protiferators and some chemicals in addtion to EGCG may have role in thisregulation and exist iii the futl-fermented balck tea.

Poster Session 1 : Analytical biochemistry Biochemical pharmacology and natural product chemistry Biotechnology Carbohydrate chemistry and metabolism ; New aminopeptidase N inhibitors, betulinic acid and indomethacin, induce apoptosis of B16 melanoma

( Young Na Yum ),( Hyun Young Cho ),( Seyl Kim ),( Myung Sil Hwang ),( Eun Kung Yoon ),( Hyo Min Lee ),( Ki Hwa Yang ),( Seung Hee Kim ) 한국생화학분자생물학회 (구 한국생화학회) 2001 생화학분자생물학회 춘계학술발표논문집 Vol.2001 No.-

Peroxisome proliferator에 의해 발현 억제되는 유전자의 검색

이국경,강호일,김세일,염영나,정자영,황명실,김옥희 식품의약품안전청 1998 식품의약품안전청 연보 Vol.2 No.-

설치류예서 peroxisome pl·oliferator(PPs)의 장기투여는 간세포의 증식을 일으키며 이에 따른 간장 무게의 증가를 유발한다. 이후 간장 무게의 증가된 상태가 계속 유지되다가 수개월 후에 간암을 발생한다. 본 실험은 PPs에 의하여 간장 무게가 증가된 간세포의 상태를 유전자 발현순준에서 이해하기 위하여 dfferential display-reverse tran scrlption-polymerase chairl reaction(DD-HT-PCR) 기법을 이용하먼 발현이 감소된 유전자를 검색하였다. 이에 82개의 발현 감소가 예측되는 CBNA 절편을 분리하였고 reverse northerB blot으로 9종윽 유전자가 발현 감소됨을 확인할 수 있었다. 9종의 선별된 CDNA 절편의 염기서열을 확인한 곁과 2종은 transferrin과 of-inhibifur 111에 대해 각각 100%읜 동질성을 갖는 CDNA 절편이었으며, 또 다른 2종의 CDNA 절펀은 long Inter spersed repetitive nuclear element(LINE)와 98%의 동질성을 보였단. 나떠지 5종윽 CDNA 절편은 매우 낮은 농질성을 보이거나 꿀색이 되지 않았다. Transferrin의 감소는 peroxisome pro?iferator actiuated receptor(PPAR)가 transferrin의 transcription factor인 HNF-4(hepatocyte nuclear factor-4)와 경쟁적인 작용함에 기인한다.본 실험에서 PPs에 의해 발현 감소되는 것으로 처음 밝혀진 α1-inhibitor III는 종양세포에서 발현이 감소하는 유전자로 알려져 잇으며 또한 역시 PPs에 의해 발현감소되는 retrotransposon인 LINE과 아직 동정되지 않은 5종의 유전자들의 발현 감소는 PPs의 작용범위가 알려진 범위보다 넓을수 있음을 제시하고 있다. 따라서 PPs는 현재까진 알려진 transcription factor외에도 다른 transcription factor와 경쟁적으로 aL울괴거나 또는 다른 경로를 통하여 단백질의 발현에 관여함으로써 간세포의 증식 또는 대사에 영향을 끼칠 수 있을 것으로 여겨진다. Peroxisome proliferators (PPs) increase the liver weight. maintain the increased weight of liver. and eventuallf influce liver tumor in rodents. To understand the hyferplastic status of liver in-duced by long-terrE treatuent of PPs, the techaique ef differential display-reverse transcription-poly-morass chain reaction (D3)-RT-PCR ) was rerformed to detect the transcriptionally down-regulatedgenes that would be relateil te the homeostasis or altered function of rat liver treated with PPs. To iso-late HNA from rat liver maintaining increased weight, the rats were given 0.5% clofibrate-containingdiet for 3 weeks. Using DD~RT-PCR teehnique,82 suspected CDNA fragnlents shewing transcriptionallydown-regulated patterns were isolated. Only 9 CDNA fragments aiuoBg these fragraents were decreasedtranscriftionally by 4 other PPs (Wy-14,643, di(2-ethylhexyl) phthalate, di(2-ethylhexyl) adipate, andperchloroethylene). Two CHNA fragments have 100% similarity wi·th transferrin and α1-inhibitor lIt,respectively. Two other CBNA fragmeBts have 98% sirailarity with long interspersed repetitive nuclearelements (LINE). The expression of transferrin was already knowB to be regulated by HNF-4(hepatecyte nuclear factor-4) and decreased by competitive action of peroxisome proliferator-activated.scepter :fPAR) . But the t.anscriptionally down-.egulatioa of of-inhibitor lIT by PPs was firstlyshown in-tliis study. The transcriptional reductions of a nlunber of LINE interspersed within raammali-an genome and the unidentified genes imply that the scope of genomic targets by PPs may be more enor-mous than what we have krㄴown. Therefore these results svggest that PPs may have an influence on cetlcycle or homeostasis througt competitive action against transcription factors or through other pathway.

Suppression of chemically-induced liver tumors by castration or estradiol-3-benzoate treatment in F344 rats.

Kang, Jin Seok,Ahn, Byeongwoo,Kim, Chuel Kyu,Han, Beom Seok,Che, Jeong-Hwan,Kim, Seyl,Jang, Dong Deuk,Yang, Ki-Hwa National Hellenic Research Foundation 2005 Oncology reports Vol.14 No.2

<P>Epidemiological data reveal that the incidence of liver cancer is markedly higher in men than women. To clarify the mechanism responsible for the induction of higher incidence of liver tumors in male animals, we investigated the modifying effect of sex hormones in diethylnitrosamine (DEN)-induced rat hepatocarcinogenesis. F344 male rats (n=120) were divided into two experiments, experiment I (Exp I) and experiment II (Exp II). In each experiment, 60 rats were randomly allocated into four groups. The mini-osmotic pumps containing doses of 47.5 mg (Exp I) or 23.75 mg (Exp II) of DEN were inserted into the abdominal cavity of each animal to initiate liver carcinogenesis. Animals in group 2 were castrated one week prior to DEN treatment, and animals in groups 3 and 4 were treated with 1 or 10 microg of estradiol-3-benzoate (EB), respectively, one week prior to DEN treatment. Animals in group 1 were treated with DEN alone and sham-operated at the same time. All animals were sacrificed 26 weeks after DEN treatment. In Exp I, liver tumor incidence of group 3 decreased significantly compared with that of group 1 (p<0.05), and tumor multiplicities of groups 2, 3 and 4 were decreased significantly compared to that of group 1 (p<0.01). In Exp II, tumor incidence of group 3 was significantly different (p<0.05) when compared to that of group 1. Immunohistochemical expression of ERalpha was shown in normal appearing cells, but not in tumor cells. Western blot analysis confirmed that ERalpha expression was higher in normal liver tissue compared to tumor tissues. Taken together, we conclude that castration or EB treatment has an inhibitory effect in DEN-induced hepatocarcinogenesis in F344 rats. The reason for ERalpha loss in tumor cells should be further elucidated.</P>

Quantification and visualization of lung metastatic tumor in mice

Ha Neul Lee,Se Ryeong Jeong,Nahyeon Gu,Kanghee Ryu,Seyl Kim,Kibeom Park,Sooah Park,Woonggyu Jung,Jin Seok Kang 한국실험동물학회 2020 한국실험동물학회 학술발표대회 논문집 Vol.2020 No.7