• BMB Reports
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• v.35 no.5
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• pp.524-531
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• 2002

Thrombotic thrombocytopenic purpura (TTP) is characterized by widespread platelet thrombi in arterioles and capillaries. Unusually large or multimeric von Willebrand factor, as well as one or ore platelet-agglutinating factors, have been implicated in the pathogenesis of TTP. But, the actual mechanisms of platelet agglutination have not been satisfactorily explained. Recent studies suggested the 37-kDa platelet-agglutinating protein (PAP) p37 to be partially responsible for the formation of platelet thrombi in patients with TTP. We studied mobility in SDS-PAGE, the sequence of N-terminal amino acid residues, DNA and antigenic characteristics of PAP p37, which might be related to the pathogenesis of TTP. PAP p37 was purified from the plasma of a 31-year-old male Korean patient with acute TTP. The findings are as follows: (1) We compared PAP p37 with thrombin through the use of SDS-PAGE, either with or without $\beta$-mercaptoethanol. PAP p37 did not appear to be cleaved between the A- and B-chains of prethrombin 2. However, thrombin did cleave between those of prethrombin 2, but linked with disulfide bridge. (2) N-terminal 21 amino acid sequence of PAP p37 was T-F-G-S-G-E-A-D-X-G-L-R-P-L-F-E-K-K-S-L-E. It appeared to be identical to that of 285-305 amino acid residues of human prothrombin (prethrombin 2). (3) No prothrombin gene DNA mutation was revealed. (4). The antigenicity of PAP p37 was similar to thrombin, which was a result of the competitive binding against the anti-thrombin antibody. With these results, we conclude that PAP p37 has similar characteristics to prethrombin2.

• Asian Pacific Journal of Cancer Prevention
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• v.17 no.2
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• pp.575-579
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• 2016

The aim of this research is to determine the characteristics of endothelial factors and the hemostatic system conditions with malignant neoplasms patients, which were exposed to ionizing radiation. Total number of examined people is 223, 153 of them are patients with gastrointestinal tract oncopathology. The article presents the results of the analysis of numerical indicators of endothelial condition and platelet hemostasis of patients, who had cancer. They lived in the regions of Kazakhstan, subjected to contamination of radionuclides as a result of nuclear weapons tests, which took place in this region from 1949 to 1989. These results then were compared to cancer patients and healthy individuals with no radiation risk. The study revealed the presence of higher levels of endothelial dysfunction and following trigger of the hemostatic system in patients with malignant tumors of the gastrointestinal tract. In particular, the defined high degree of endothelial dysfunction include endothelium-dependent vasodilation, content desquamated endothelial cells in peripheral blood and von Willebrand factor. These indicators have a clear correlation with the degree of disorder of studied parameters of the hemostasis, which can cause the development of thrombotic complications.

• The Korean Journal of Physiology and Pharmacology
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• v.25 no.1
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• pp.27-38
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• 2021

Excessive salt intake induces hypertension, but several gamma-aminobutyric acid (GABA) supplements have been shown to reduce blood pressure. GABA-salt, a fermented salt by L. brevis BJ20 containing GABA was prepared through the post-fermentation with refined salt and the fermented GABA extract. We evaluated the effect of GABA-salt on hypertension in a high salt, high cholesterol diet induced mouse model. We analyzed type 1 macrophage (M1) polarization, the expression of M1 related cytokines, GABA receptor expression, endothelial cell (EC) dysfunction, vascular smooth muscle cell (VSMC) proliferation, and medial thicknesses in mice model. GABA-salt attenuated diet-induced blood pressure increases, M1 polarization, and TNF-α and inducible nitric oxide synthase (NOS) levels in mouse aortas, and in salt treated macrophages in vitro. Furthermore, GABA-salt induced higher GABAB receptor and endothelial NOS (eNOS) and eNOS phosphorylation levels than those observed in salt treated ECs. In addition, GABA-salt attenuated EC dysfunction by decreasing the levels of adhesion molecules (E-selectin, Intercellular Adhesion Molecule-1 [ICAM-1], vascular cell adhesion molecule-1 [VCAM-1]) and of von Willebrand Factor and reduced EC death. GABA-salt also reduced diet-induced reductions in the levels of eNOS, phosphorylated eNOS, VSMC proliferation and medial thickening in mouse aortic tissues, and attenuated Endothelin-1 levels in salt treated VSMCs. In summary, GABA-salt reduced high salt, high cholesterol diet induced hypertension in our mouse model by reducing M1 polarization, EC dysfunction, and VSMC proliferation.

• Journal of Chest Surgery
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• v.40 no.3 s.272
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• pp.180-192
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• 2007

Background: As determined from the recent investigations of discordant cardiac xenotransplantation, hyperacute rejection occurs mainly at the endothelial cells in donor microvascular systems, but this does not occur at cardiac valve leaflets or at medium-to-large caliber vessels. On the basis of this background, this study was performed to look into the biocompatibility for transplantation of a middle or large diameter xenogenic blood vessel by conducting xenogenic arterial transplantation with the carotid artery in a pig-to-goat model. Material and Method: The experimental group was composed of 10 pairs of pig-to-goat combinations. They were divided into each period of 1 week, and 1, 3, 6 and 12 months. Four carotid artery grafts obtained through collection of the bilateral carotid arteries from two pigs were preserved at $-70^{\circ}C$ without other treatment, and then they were transplanted into the bilateral carotid arteries of two goats. Doppler ultrasonography was done on a periodic basis after transplantation to evaluate the patency of the grafted blood vessel. At the ends of a predetermined period, the grafts were explanted from the goats and they underwent gross examination. Hematoxylin-eosin and Masson's trichrome staining were conducted. In addition, in order to examine the immunological rejection of the grafted xenogenic blood vessel, immunohistochemical staining was conducted with T-lymphocyte indicator and von Willebrand factor. Result: Two goats at the each one-week period and the one-year period died during the experimental period because of a reason unrelated to the experimental procedure, and the remaining 8 goats survived until the end of each experiment period. On Doppler ultrasonography, unilateral carotid artery occlusion was found in a goat, whose period was specified as 3 months, among the 8 survived goats. However, the vascular patency was maintained well and there was no graft that formed aneurysms in the other goats. On gross examination, the region of vascular anastomosis was preserved well, and calcification of the grafted blood vessel was not shown. Histologically, the endothelial cells of the graft disappeared one week after transplantation, and then there was progressive spread of the recipients' endothelial cells from the anastomotic site. The reendothelialization occurred over the whole graft at one month after transplantation. The neointimal thickening and adventitial inflammation became severe by 3 months after transplantation, but this lessened at 6 months and 12 months, respectively. The rate of CD3 positive cells was very low among the infiltrated inflammatory cells. Conclusion: The fresh-frozen xenogenic artery kept its patency without being greatly influenced by xenogenic immune reaction.

• Journal of Oral Medicine and Pain
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• v.24 no.3
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• pp.335-345
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• 1999

법의학적 개인식별 및 친생자 감정시 여러 개의 single tandem repeats(STR) 유전좌위 검색이 필요하다. 그 이유는 STR 유전좌위는 대립유전자 수가 적고 이형접합도가 낮아 서로 다른 개체간에도 동일한 유전좌위를 가질 확률이 높기 때문에 개인식별에 대한 기여도가 떨어지게 된다. 따라서 여러 개의 다양한 STR 유전좌위들을 동시에 분석함으로써 우연적으로 개체간에 유전자형이 일치할 가능성을 낮추어야 감정의 신뢰성을 높일 수 있으며 이에는 각 STR 유전좌위에 대한 유전좌위의 분포가 인종별, 지역별로 달라 이에 대한 유전자분포를 구하는 것이 선행조건이다. 이에 본 연구에서는 법의학적 개인식별 및 친자감정시 기초자료로 활용하기 위하여 서로 혈연관계가 없는 201명의 한국인 혈액에서 DNA를 추출하여 STR 유전좌위증 human coagulation factor XIII A subunit gene(F13A0l Locus), human c-fes/fps proto-oncogene(FESFPS Locus), human von Willebrand factor gene (vWA Locus)등 FFv Triplex 유전자를 중합효소반응에 의하여 동시에 증폭하고, 폴리아크릴아마이드겔을 이용한 전기영동 및 질산은 염색을 시행한 후 FFv Triplex유전자의 유전자형 및 대립유전자 빈도 등을 분석하여 다음과 같은 결과를 얻었다. (1) F13A01유전자는 5개의 대립유전자, 12개의 유전자형을 검출하였으며, 이형접합도는 60.7%로 나타났고 대립유전자 및 유전자빈도는 3.2, 4, 5, 6, 16 대립유전자에서 각각 0.34 3, 0.114, 0.062, 0.475, 0.005로 나타났으며, 대립유전자 7, 8, 9, 10, 11, 12, 13, 14, 15는 검출되지 않았다. (2) F13A01 대립유전자다양성 (allelic diversity value)은 0.641, 개인식별력(PD)은 0.814를 보였으며 대립유전자다양성 및 이형접합도가 다른 민족과 비교할 때 다소 낮았다. (3) FESFPS유전자는 8개 대립유전자 모두 나타났으며, 15개의 유전자형을 검출하였으며, 이형접합도는 66.7%로 나타났고 대립유전자 및 유전자빈도는 7, 8, 9, 10, 11, 11, 12, 13, 14 대립유전자에서 각각 0.002, 0.002, 0.005, 0.032, 0.507, 0.264, 0.197, 0.007로 나나났다. (4) FESFPS 대립유전자다양성(allelic diversity value)은 0.641, 개인식별력(PD)은 0.804를 보였다. (5) vWA유전자는 9개의 대립유전자, 23개의 유전자형을 검출하였으며, 이형접합도는 80.1%로 나타났고 대립유전자 및 유전자 빈도는 11, 12, 14, 15, 16, 17, 18, 19, 20 대립유전자 에서 각각 0.002, 0.002, 0.219, 0.032, 0.187, 0.279, 0.189, 0.072, 0.017로 나타났으며, 대립 유전자 13, 21는 검출되지 않았다. (6) vWA 대립유전자다양성(allelic diversity value)은 0.799, 개인식별력(PD)은 0.924로 매우 높게 나타냈다.

• International Journal of Oral Biology
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• v.32 no.1
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• pp.13-22
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• 2007

The stem cell research is emerging as a cutting edge topic for a new treatment for many chronic diseases. Recently, dental stem cell would be possible for regeneration of tooth itself as well as periodontal tissue. However, the study of the cell characterization is scarce. Therefore, we performed the genetic profiling and the characterization of mouse fetus/neonate derived dental tissue and cell to find the identification during dental development. We separated dental arch from mandibles of 14.5 d fetal mice and neonate 0 d under the stereoscope, and isolated dental cells primarily from the tissues. Then, we examined morphology and the gene expression profiles of the primary cells and dental tissues from fetus/neonate and adult with RT-PCR. Primary dental cells showed heterogeneous but the majority was shown as fibroblast-like morphology. The change of population doubling time levels (PDLs) showed that the primary dental cells have growth potential and could be expanded under our culture conditions without reduction of growth rate. Immunocytochemical and flow cytometric analyses were performed to characterize the primary dental cell populations from both of fetus (E14.5) and neonate. Alpha smooth muscle actin (${\alpha}-SMA$), vimentin, and von Willebrand factor showed strong expression, but desmin positive cells were not detected in the primary dental cells. Most of the markers were not uniformly expressed, but found in subsets of cells, indicating that the primary dental cell population is heterogeneous, and characteristics of the populations were changed during culture period. And mesenchymal stem cell markers were highly expressed. Gene expression profile showed Wnt family and its related signaling molecules, growth factors, transcription factors and tooth specific molecules were expressed both fetal and neonatal tissue. The tooth specific genes (enamelin, amelogenin, and DSPP) only expressed in neonate and adult stage. These expression patterns appeared same as primary fetal and neonatal cells. In this study we isolated primary cells from whole mandible of fetal and neonatal mice. And we investigated the characteristics of the primary cells and the profile of gene expressions, which are involved in epithelial-mesenchymal interactions during tooth development. Taken together, the primary dental cells in early passages or fetal and neonatal mandibles could be useful stem cell resources.

• Journal of Embryo Transfer
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• v.33 no.3
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• pp.129-137
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• 2018

Acute vascular rejection has been known as a main barrier occurring in a xenograted tissue of alpha 1,3-galactosyltransferase knock-out (GalT KO) pig into a non-human primate (NHP). Adenosine which is a final metabolite following sequential hydrolysis of nucleotide by ecto-nucleotidases such as CD39 and CD73, act as a regulator of coagulation, and inflammation. Thus xenotransplantation of CD39 and CD73 expressing pig under the GalT KO background could lead to enhanced survival of recipient NHP. We constructed a human CD39 and CD73 expression cassette designed for endothelial cell-specific expression using porcine Icam2 promoter (pIcam2-hCD39/hCD73). We performed isolation of endothelial cells (pAEC) from aorta of 4 week-old GalT KO and membrane cofactor protein expressing pig ($GalT^{-MCP/-MCP}$). We were able to verify that isolated cells were endothelial-like cells using immunofluorescence staining analysis with von Willebrand factor antibody, which is well known as an endothelial maker, and tubal formation assay. To find optimal condition for efficient transfection into pAEC, we performed transfection with GFP expression vector using four programs of nucleofection, M-003, U-023, W-023 and Y-022. We were able find that the program W-023 was optimal for pAEC with regard to viability and transfection efficiency by flow cytometry and fluorescent microscopy analyses. Finally, we were able to obtain $GalT^{-MCP/-MCP}/CD39/CD73$ pAEC expressing CD39 and CD73 at levels of 33.3% and 26.8%, respectively. We suggested that pACE isolated from $GalT^{-MCP/-MCP}$ pig might be provided as a basic resource to understand biochemical and molecular mechanisms of the rejections and as an alternative donor cells to generate $GalT^{-MCP/-MCP}/CD39/CD73$ pig expressing CD39 and CD73 at endothelial cells.

• Journal of Nutrition and Health
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• v.35 no.1
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• pp.5-14
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• 2002

Alteration in the syntesis or enhanced inactivation of nitric oxide(NO) can induce impairment of endothelial cell function. Insulin dependent diabetes mellitus(IDDM) is characterized by impaired endothelial function and vascular disease. NO is produced through L-arginine pathway To elucidate the hypothesis that the decreased production on NO in IDDM reflects vascular damage and the NO production can be manipulated by either dietary fat(7% of kg diet) or the oral supplementation with L-arginine(2g/kg bw), plasma markers for vascular endothelial damage and plasma lipid profiles were measured in streptozotocin(STZ)-induced diabetic rats. Diabetic or normal Sprague-Dawley rats were fed 6 different experimental diets for 4 weeks(SO : soybean oil, SOA: soybean oil + L-arginine supplementation, BT : beef tallow, BTA_ beef tallow + L-arginine supplementation, OV olive oil, OVA : olive oil + L-arginine supplementation). Plasma glucose, total cholesterel, HDL-cholesterol, LDL-cholesterol and triglyceride were measured. Endothelial markers, plasma von Willebrand factor(vWf), thromboxane B$_2$, and 6-keto PGF1$\alpha$ of aorta were measured by ELISA. Plasma NO production was evaluated through the measurement of nitrite by EIA. Feeding saturated fatty acid(SFA, BT) increased relative liver size(RLS) in diabetic rats compared to either polyunsatunted fatty acid(PUFA, SO) or monounsaturated fatty acid(MUFA, OV) The supplementation of L-arginine inhibited the liver and kidney enlargement in olive oil find diabetic rats. Plasma glucose was lower in diabetic animal find the olive oil compared to fed beef tallow and the supplementation L-arginine decreased it in diabetic rats find beef tallow significantly(p < 0.05). Plasma TXB$_2$ levels were increased due to diabetes and the value of beef tallow group showed highest value. Plasma vWf concentration of beef tallow group was higher value in normal rats and was elevated more in diabetes. In diabetic groups, the vWf concentration of olive oil group was lower than beef tallow or soybean oil group. The supplementation of L-arginine in diabetic rats decreased plasma TXB$_2$ and vWf levels significantly(p < 0.05). NO production was higher in normal olive oil fed rats and was tend to be decreased in diabetic rats and the supplementation of L-arginine recovered to normal value(p < 0.05), Olive oil supplemented with L-arginine tended to lower plasma total cholesterol and LDL-cholesterol after 4 week treatment. These results suggest that generalized vascular endothelial changes based on plasma TXB$_2$and vWf occurs in diabetic rats. and olive oil with L-arginine supplementation contributes to a better control of the hyperglycemia, endothelial changes and hypercholesterolemia accompanying diabetes as compared with beef tallow or soy bean oil in this rat model.

• Journal of Yeungnam Medical Science
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• v.15 no.2
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• pp.286-296
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• 1998

Different techniques for culturing respiratory epithelial cells have been developed to overcome the limitations of studies on in vivo and on bioptic material. Traditionally, culture systems are divided into organ cultures, explant cultures and dissociated cell cultures. The first two contain both epithelial and non-epithelial cells. However, in monolayer cultures of dissociated cells only epithelial cells are present, the effects observed are caused by a pure epithelial responses. The purpose of this study is to establish primary culture method of human nasal epithelium (HNEC) by monolayer culture of dissociated cells to evaluate the role of the epithelial cells in the allergic and non-allergic nasal inflammatory reactions. HNEC was prepared by primary culture method of monolayer culture of dissociated cells from human inferior nasal turbinate mucosa of septal deviation patients. Primary cultured cells were characterized by indirect immunofluorescence assay and transmission electron microscopy. The immunoreactivities of cytokeratin-pan and cytokeratin No. 8 were observed in cultured HNEC. However, the immnoreactivities of vimentin and von Willebrand factor were not observed in cultured HNEC. The tonofilaments and desmosome were observed in cultured HNEC. The cultured epithelial cells were identified to be pure nasal epithelial cells. The monolayer culture of dissociated cells could successfully be employed for further study to investigate the role of the epithelial cells in allergic or non-allergic nasal inflammatory diseases.

• IMMUNE NETWORK
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• v.1 no.1
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• pp.87-94
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• 2001

Background: Cytokine-mediated ex vivo expansion has been proposed as a means of increasing the number of cord blood (CB) hematopoietic stem cells for transplantation. As well as stem cell number, stromal cells are necessary for functional maturation of hematopoiesis. The purpose of this study was to analyze the development of stromal cells during ex vivo expansion of CB $CD34^+$ cells. Methods : $CD34^+$ cells were purified from CB by magnetic bead selection. The levels of of interleukin-3, interleukin-$1{\beta}$, interleukin-6, granulocyte macrophagecolony stimulating factor and tumor necrosis factor-${\alpha}$ were measured in culture supernatants on 0, 1, 2, and 3 weeks, using ELISA techniques. CB $CD34^+$ cells were expanded in Iscoves modified Dulbeccos medium in the presence of several cytokines. The expression of E-selectin, vascular cell adhesion molecule-1, intercellular adhesion molecule-1, platelet/endothelial cell adhesion molecule-1, von Willebrand factor, vimentin, and CD14 in newly developed stromal cells was examined by immunocytochemical method. Relevant extracellular matrix (ECM) proteins and proper cytokines were also assayed for the most suitable condition for expansion of stromal cells. Results: Several cytokines were found to have been produced by CB $CD34^+$ cells as well as bone marrow-derived $CD34^+$ cells. During ex vivo expansion of CB $CD34^+$ cells, stromal cells appeared in the culture by day 4 and expanded over the following 7-10 days before being confluent by day 2 1. These cells expressed surface markers characteristic of cells of endothelial lineage. Furthermore, these stroaml cells also expanded effectively when treated with thrombopoietin+flt-3 ligand+stem cell factor+leukemia inhibitory factor or 0.1% poly-L-lysine-coated wells. Conclusion: Stromal cells were developed during ex vivo expansion of CB $CD34^+$ cells and that this development could be enhanced further by treating the stromal cells with cytokines or ECM.