• Childhood Kidney Diseases
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• v.26 no.2
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• pp.101-106
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• 2022

Nephrotic syndrome (NS) is a hypercoagulable state in which children are at risk of venous thromboembolism. A higher risk has been reported in children with steroid-resistant NS than in those with steroid-sensitive NS. The mortality rate of cerebral venous sinus thrombosis (CVST) is approximately 10% and generally results from cerebral herniation in the acute phase and an underlying disorder in the chronic phase. Our patient initially manifested as a child with massive proteinuria and generalized edema. He was treated with albumin replacement and diuretics, angiotensin-converting enzyme inhibitor, and deflazacort. Non-contrast computed tomography showed areas of hyperattenuation in the superior sagittal sinus when he complained of severe headache and vomiting. Subsequent magnetic resonance imaging revealed empty delta signs in the superior sagittal, lateral transverse, and sigmoid sinuses, suggesting acute CVST. Immediate anticoagulation therapy was started with unfractionated heparin, antithrombin III replacement, and continuous antiproteinuric treatment. The current report describes a life-threatening CVST in a child with steroid-dependent NS, initially diagnosed by contrast non-enhanced computed tomography and subsequently confirmed by contrast-enhanced magnetic resonance imaging, followed by magnetic resonance venography for recanalization, addressing successful treatment.

• Journal of Microbiology and Biotechnology
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• v.18 no.3
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• pp.503-511
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• 2008

An edible marine red alga, Grateloupia filicina, collected from Jeju Island of Korea was hydrolyzed by cheap food-grade carbohydrases (Viscozyme, Celuclast, AMC, Termamyl, and Ultraflo) to investigate its anticoagulant activity. Among the tested enzymatic extracts of G. filicina, a Termamyl extract showed the highest anticoagulant activity. Anion-exchange chromatography on DEAE-cellulose and gel-permeation chromatography on Sepharose-4B were used to purify the active polysaccharide from the crude polysaccharide fraction of G. filicina. The purified sulfated polysaccharide (0.42 sulfate/total sugar) showed ${\sim}1,357kDa$ molecular mass and was comprised mainly of galactose(98%) and 1-2% of glucose. The sample showed potential anticoagulant activity on activated partial thromboplastin time (APTT) thrombin time (TT) assays. The purified G. filicina anticoagulant (GFA) inhibited the coagulation factor X (92%), factor II (82%), and factor VII (68%) of the coagulation cascade, and the molecular interaction (protein-polysaccharide) was highly enhanced in the presence of ATIII (antithrombin III). The dissociation constant of polysaccharide towards serine proteins decreased in the order of FXa (58.9 nM) >FIIa (74.6 nM) >FVII (109.3 nM). The low/less cytotoxicity of the polysaccharide benefits its use in the pharmaceutical industry; however, further studies that would help us to elucidate the mechanism of its activity are needed.

• The Journal of Internal Korean Medicine
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• v.17 no.1
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• pp.34-50
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• 1996

The present experiment was desinged to investigate the effects of Tongdosan water extracts on the Cardiovascular System in the Experimental Animals. Thus, the changes of blood pressure and heart rate were measured after oral admini- stration. Measurment of Mortality rate was observed for measuring the effect of Tongdosan water extract. Tongdosan water extract against pulmonary thrombo- embolism induced by collagen the mixture(0.1ml/10g, 2mg/kg B.W) plus serotonin(5mg/kg B.W) in mouse. The effect of Tongdosan water extract was examined by observing the change of collagen-induced platelet aggregation, coagulation activity, ex vivo and in vitro fibrinolytic activity of euglobulin fraction in rats. The results were summarized as followings. 1. Tongdosan dropped the blood pressure in spontaneous hypertensive rat. 2. The drug increased the auricular blood flow in rabbit. 3. The drug relaxed the artery contraction by pretreated norepinephrine in rat. 4. The drug inhibited the death rate of mouse which was led to thromboembo- lism by serotonin and collagen. 5. The drug inhibited the platelet aggregation in rat. 6. The drug prolonged the prothrombin time and activated partial thromboplastin time on the test of plasma coagulation factor activity in rat, but was not valuable. 7. The drug reduced the fibrinogen lyses time of rat ex vivo assay and lyses area was increased. 8. Tongdosan reduced fibrinogen lyses time of rat in vitro assay. According to the above mentioned results, Tongdosan increased the blood flow and dropped the blood pressure by the dilation of blood vessel. And the drug presented the antithrombin acivity, inhibited the platelet aggregation.

• Toxicological Research
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• v.17
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• pp.97-104
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• 2001

Three types of proteases (MEF-1, MEF-2 and MEF-3) were purified from the egg cases of Ten-odera sinensis using ammonium sulfate fractionation, gel filtration on Bio-Gel P-60 and affinity chromatography on DEAE Affi-Gel blue gel. The proteases were assessed homogeneous by SDS-polyacrylamide gel electrophoresis and have molecular weight of 31,500, 32,900 and 35,600 Da, respectively. The N-terminal regions of the primary structure were compared and they were found to be different each other. MEFs readily digested the $A\alpha$ - and B$\beta$-chains of fibrinogen and more slowly the ${\gamma}$-chain. The action of the enzymes resulted in extensive hydrolysis of fibrinogen and fibrin, releasing a variety of fibrinopeptides. MEF-1 was inactivated by Cu$^{2+}$ and Zn$^{2+}$ and inhibited by PMSF and chymostatin. MEF-2 was inhibited by PMSF, TLCK. soybean trypsin inhibitor. MEF-3 was only inhibited by PMSF and chymostatin. Antiplasmin was not sensitive to MEF-1 but antithrombin III inhibited the enzymatic activity qf MEF-1. MEF-2 specifically bound to anti plasmin Among the chromogenic protease substrates, the most sensitive one to the hydrolysis of MEFs was benzoyl-Phe-Val-Arg-p-nitroanilide with maximal activity at pH 7.0 and 3$0^{\circ}C$. MEF-1 preferentially cleaved the oxidized B-chain of insulin between Leu15 and Tyr16. In contrast, MEF-2 specifically cleaved the peptide bond between Arg23 and Gly24. D-dimer concentrations increased on incubation of cross-linked fibrin with MEF-1, indicating the enzyme has a strong fibrinolytic activity.ity.

• Proceedings of the Korean Society for Applied Microbiology Conference
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• 2000.04a
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• pp.178-181
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• 2000

In order to develop the novel gene expression system, we introduced new control elements which could influence the foreign gene expression in animal cells. When the foreign genes are introduced into the genome of higher eukaryotic cells, the expressions from these integrated genes are often low and can vary greatly depending on the positions of the integration sites due to the complex nature of the chromatin structures (1). First we screened the various DNA sequence elements which can function as an insulator of gene expression from these position effects and can cooperate with the SV40 enhancer/promoter. Among the several DNA elements from the various sources, we identified the particular DNA element which confers the increased frequency of the positive colonies, assayed by the reporter gene from stable selections indicating significantly reduced position effects. This element also showed the several fold-increased expression level as well as the copy-number dependent expression with host cell specificity. Second we modified the transcription termination element where we introduced the specific terminator in combination with SV40 polyA signal. This modified terminator showed the increased efficiency and the level of the gene expression. By combining these two elements, we made the animal cell expression system and tested successfully for the recombinant protein productions of TGF ${\beta}$-soluble receptor, Antithrombin III, and single chain Pro-Urokinase. [Supported by grants from MOCIE]

• Biotechnology and Bioprocess Engineering:BBE
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• v.5 no.4
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• pp.234-241
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• 2000

The Saccharomyces cerevisiae PMR1 gene encodes a Ca2+-ATPase localized in the Golgi. We have investigated the effects of PMR1 disruption in S. cerevisiae on the glycosylation and secretion of three heterologous glycoproteins, human ${\alpha}$1-antitrypsin (${\alpha}$1-AT), human antithrombin III (ATHIII), and Aspergillus niger glucose oxidase (GOD). The pmr1 null mutant strain secreted larger amounts of ATHIII and GOD proteins per a unit cell mass than the wild type strain. Despite a lower growth rate of the pmr1 mutant, two-fold higher level of human ATHIII was detected in the culture supernatant from the pmr1 mutant compared to that of the wild-type strain. The pmr1 mutant strain secreted ${\alpha}$1-AT and the GOD proteins mostly as core-glycosylated forms, in contrast to the hyperglycosylated proteins secreted in the wild-type strain. Furthermore, the core-glycosylated forms secreted in the pmr1 mutant migrated slightly faster on SDS-PAGE than those secreted in the mnn9 deletion mutant and the wild type strains. Analysis of the recombinant GOD with anti-${\alpha}$1,3-mannose antibody revealed that GOD secreted in the pmr1 mutant did not have terminal ${\alpha}$1,3-linked mannose unlike those secreted in the mnn9 mutant and the wild type strains. The present results indicate that the pmr1 mutant, with the super-secretion phenotype, is useful as a host system to produce recombinant glycoproteins lacking high-mannose outer chains.

• The Journal of Internal Korean Medicine
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• v.20 no.1
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• pp.57-72
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• 1999

This study was designed to elucidate the anti-inflammatory, cardiovascular, anti-thrombotic, and analgesic effect of Whalrakdan. The anti-inflammatory effects was measured by the method of carrageenin induced edema, protein leakage test using CMC-pouch, and the effect of Whalrakdan on the cardiovascular system was observed by the change of flow rate of Ringer solution in the vascular system in the ear of rabbit. and the contraction and dilatation of rat tail artery. Death rate, platelet aggregation, plasma coagulation activity, antithrombin activity was observed for the measurement of the anti-thrombotic effect of Whalrakdan, and the analgesic effect was measured by the acetic acid method and hot plate method. The result was as follows: 1. After 2 or 3hour of Whalrakdan administration, carrageenin induced edema and CMC-pouch protein leakage was significantly decreased. 2. The slight anagesic effect of Whalrakdan extract was confirmed by the observation of writhing syndrome, paw licking time, and escape time. 3. The droplet of Ringer solution increased according to the increase of concentration of Whalrakdan extract, and the vasoconstriction decreased dependantly to the concentration of Whalrakdan extract. 4. The anti-thrombotic effect of Whalrakdan was observed by the decrease of death rate, the inhibition of platelet aggregation, and the increase of anti-thrombin activity.

• Archives of Pharmacal Research
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• v.29 no.5
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• pp.418-423
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• 2006

Tabanus anticoagulant protein (TAP) was isolated from the whole body of the tabanus, Tabanus bivittatus, using three purification steps (ammonium sulfate fractionation, gel filtration on Bio-Gel P-60, and ion exchange chromatography on DEAE Sephadex gel). The purified TAP, with a molecular weight of 65 kDa, was assessed to be homogeneous by SDS-polyacrylamide gel electrophoresis, and an isoelectric point of 7.9 was determined by isoelectric focusing. The internal amino acid sequence of the purified protein was composed of Ser-Leu-Asn-Asn-Gln-Phe-Ala-Ser-Phe-lle-Asp-Lys-Val-Arg. The protein was activated by $Cu^{2+}\;and\;Zn^{2+}$, and the optimal conditions were found to be at pH $3\sim6\;and\;40\sim70^{\circ}C$. Standard coagulation screen assays were used to determine thrombin time and activated partial thromboplastin time. Chromogenic substrate assays were performed for thrombin and factor Xa activity. TAP considerably prolonged human plasma clotting time, especially activated partial thromboplastin time in a dose-dependent manner; it showed potent and specific antithrombin activity in the chromogenic substrate assay. Specific anti-factor Xa activity in TAP was not detected. Overall, this result suggested that TAP has significant anticoagulant activity on blood coagulation system.

• Journal of Chest Surgery
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• v.35 no.3
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• pp.239-243
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• 2002

In this case, a 39 year-old man was admitted with Budd-Chiari syndrome associated with complete superior vena cava(SVC) obstruction causing general edema and hepatic failure. Conservative medical therapy was failed. And after the radiologist failed to invasive procedure of balloon dilatation, we attempted the inferior vena cava to right atrium bypass graft. Operation was done through median sternotomy and extended vertical oblique abdominal incision. A 24 mm Dacron tube was placed from the inferior vena cava just below the left renal vein to the right atrium without using the cardiopulmonary bypass pump. The patient's postoperative course was uneventful without signs of bleeding or any other complications. We used anticoagulants at the postoperative first day. At the postoperative 26th day, we performed abdominal Doppler sonography and we confirmed that the graft patency was good. The patient was discharged with SVC obstructive symptoms but we noticed relief of SVC obstructive symptoms in the course of follow-up.

• Archives of Pharmacal Research
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• v.28 no.7
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• pp.816-822
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• 2005

Catharsius protease-2 (CPM-2) was isolated from the body of dung beetles, Catharsius molossus, using a three step purification process (ammonium sulfate fractionation, gel filtration on Bio-Gel P-60, and affinity chromatography on DEAE Affi-Gel blue). The purified CPM-2, having a molecular weight of 24 kDa, was assessed homogeneously by SDS-polyacrylamide gel electrophoresis. The N-terminal amino acid sequence of CPM-2 was composed of X Val Gin Asp Phe Val Glu Glu lie Leu. CPM-2 was inactivated by $Cu^{2+}\;and\;Zn^{2+}$ and strongly inhibited by typical serine proteinase inhibitors such as TLCK, soybean trypsin inhibitor, aprotinin, benzamidine, and ${\alpha}_1$-antitrypsin. However, EDTA, EGTA, cysteine, $\beta$-mercaptoethanol, E64, and elastatinal had little effect on enzyme activity. In addition, antiplasmin and antithrombin III were not sensitive to CPM-2. Based on the results of a fibrinolytic activity test, CPM-2 readily cleaved $A{\alpha}-$ and $B{\beta}$-chains of fibrinogen and fibrin, and y-chain of fibrinogen more slowly. The nonspecific action of the enzyme resulted in extensive hydrolysis, releasing a variety of fibrinopeptides of fibrinogen and fibrin. Polyclonal antibodies of CPM-2 were reactive to the native form of antigen. The ELISA was applied to detect quantities, in nanograms, of the antigen in CPM-2 protein.