• 대한임상검사과학회지
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• 제51권1호
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• pp.34-41
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• 2019

혈소판 응집은 혈관 손상의 경우 지혈 플러그 형성에 필수적이다. 그러나, 과도한 혈소판 응집은 혈전증, 죽상 동맥 경화증 및 심근 경색과 같은 순환기 장애를 일으킬 수도 있다. Scopoletin은 Scopolia 또는 Artemisia 속 식물의 뿌리에서 발견되는 성분으로, 항응고 및 항말라리아 작용을 가지는 것으로 알려져 있다. 본 연구는 collagen에 의해 유발된 혈소판 응집에 scopoletin이 미치는 영향을 조사하였다. Scopoletin은 활성화된 혈소판에서 생성되는 응집 유도 분자인 thromboxane $A_2$ ($TXA_2$) 및 세포 내 $Ca^{2+}$ 동원 ($[Ca^{2+}]_i$)의 하향 조절을 통해 항 혈소판 효과를 나타내었다. 한편, scopoletin은 세포 내 $Ca^{2+}$-길항제인 것으로 알려져 있는 cyclic adenosine monophosphate(cAMP)와 cyclic guanosine monophosphate (cGMP) 수치를 증가시켰다. 특히, scopoletin은 cGMP보다 cAMP 수준을 강력하게 증가함으로써 콜라겐에 의해 유발된 사람 혈소판 응집에서의 ${\alpha}IIb/{\beta}_3$에 대한 피브리노겐 결합을 억제하였다. 또한, scopoletin은 용량 의존적으로 collagen에 의해 증가된 adenosine trisphosphate (ATP)의 방출을 억제하였다. 이 결과는 혈소판 내 과립 분비를 통한 응집 증폭작용이 scopoletin에 의해 억제되었음을 의미한다. 따라서, 본 연구는 scopoletin이 강력한 항혈소판 효과를 가지며 혈소판-유래의 혈관 질환을 예방할 가능성이 크다는 것을 입증하였다.

• Archives of Pharmacal Research
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• 제29권5호
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• pp.375-383
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• 2006

The anti platelet effects of a novel guanidine derivative, KR-32570 ([5-(2-methoxy-5-chlorophenyl) furan-2-ylcarbonyl]guanidine), were investigated with an emphasis on the mechanisms underlying its inhibition of collagen-induced platelet aggregation. KR-32570 significantly inhibited the aggregation of washed rabbit platelets induced by collagen $(10{\mu}g/mL)$, thrombin (0.05 U/mL), arachidonic acid $(100{\mu}M)$, a thromboxane (TX) $A_2$ mimetic agent U46619 (9,11-dideoxy-9,11-methanoepoxy-prostaglandin $F_2,\;1{\mu}M$) and a $Ca^{2+}$ ATPase inhibitor thapsigargin $(0.5{\mu}M)$ ($IC_{50}$ values: $13.8{\pm}1.8,\;26.3{\pm}1.2,\;8.5{\pm}0.9,\;4.3{\pm}1.7\;and\;49.8{\pm}1.4{\mu}M$, respectively). KR-32570 inhibited the collagen-induced liberation of $[^3H]$arachidonic acid from the platelets in a concentration dependent manner with complete inhibition being observed at $50{\mu}M$. The $TXA_2$ synthase assay showed that KR-32570 also inhibited the conversion of the substrate $PGH_2$ to $TXB_2$ at all concentrations. Furthermore, KR-32570 significantly inhibited the $[Ca^{2+}]_i$ mobilization induced by collagen at $50{\mu}M$, which is the concentration that completely inhibits platelet aggregation. KR-32570 also decreased the level of collagen $(10{\mu}g/mL)$induced secretion of serotonin from the dense-granule contents of platelets, and inhibited the NHE-1-mediated rabbit platelet swelling induced by intracellular acidification. These results suggest that the antiplatelet activity of KR-32570 against collagen-induced platelet aggregation is mediated mainly by inhibiting the release of arachidonic acid, $TXA_2$ synthase, the mobilization of cytosolic $Ca^{2+}$ and NHE-1.

• 대한치과보철학회지
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• 제50권3호
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• pp.149-155
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• 2012

연구 목적: 이 논문은 임플란트 고정체가 골융합이 이루어지기전 의원성 동요가 있을 경우 골결합에 어떤 영향을 미치는지를 알아 보고자 한다. 연구 재료 및 방법: 실험에 사용한 임플란트는 직경 3.73 mm, 길이 4 mm의 순수한 타이타늄과 RBM ($MegaGen^{(R)}$: Ca-P) 처리된 임플란트(Grade IV)를 사용하였다. 몸무게 3.5kg 이상의 토끼(Female, New Zealand White)의 한쪽 경골에 2개씩 양쪽 다리에 임플란트를 식립하여 모두 80개의 임플란트가 식립되었다. 비틀림 제거력(Removal torque)의간격에 따라 그룹 I (6주), 그룹 II (4일+6주), 그룹 III (4일+1주+6주), 그룹 IV (1주+6주), 그룹 V (1주+1주+6주), 그룹 VI (2주+6주), 그룹 VII (2주+1주+6주), 그룹 VIII (3주+6주), 그룹 IX (3주+1주+6주), 그룹 X (10주)으로 10개의 그룹으로 나누었다. 본 실험에서 그룹 I과 그룹X가 대조군이며, 비틀림 제거력은 digital torque gauze (Mark-10, USA)를 사용하여 6주와 10주에 측정하였다. 실험군에서는 마지막 비틀림 제거력을 측정하기 전에 의원성 동요를 가하여 한 번 혹은 두 번 비틀림 제거력을 측정하여 그 수치를 기록하였다. 그 후, 대조군을 제외하고 비틀림 제거력을 측정한 임플란트는 가급적 원래의 위치로 돌려 놓고 봉합을 하였다. 모든 실험군은 마지막 비틀림 제거력 측정전까지 6주간의 치유기간을 주었으며, 마지막 비틀림 제거력은 실험군 첫 번째 또는 두 번째 비틀림 제거력값과 비교하여 결과를 분석하였다. 결과: 마지막 비틀림 제거력 측정값에서 그룹X (10주)의 값이 대조군인 그룹 I (6주)의 값보다 높았으나, 통계적으로는 유의하지 않았다. 실험군과 대조군 사이의 통계적으로 유의한 차이를 보이지 않았다(P>.05). 첫 번째 비틀림 제거력 측정치에서, 실험군(4일혹은1주)에서 다른 실험군(2주혹은3주)의 수치보다 낮은 값을 보였다. 치유기간에 따른 각 실험군의 비교에서, 최종 비틀림 제거력 값이 첫 번째 비틀림 제거력 값 보다 현저히 높은 값을 보였다. 결론: 골유착이 형성되기 전 한 번 또는 두 번 의원성으로 동요는, 만약 충분한 치유기간을 가지게 될 경우 임플란트의 골유착에 영향을 주지 않는다.

• Biomolecules & Therapeutics
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• 제7권3호
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• pp.227-233
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• 1999

The effects of 2-bromo-3-(3,5-tert-butyl-4-hydroxylphenyl)-1,4-naphthalenedione(TPN2), a synthetic vitamin K derivative, on platelet aggregation and its action mechanisms were investigated in rat platelet. TPN2 inhibited the platelet aggregation induced by collagen($10\mu\textrm{g}$/ml), thrombin(0.1 U/ml), A23187($10\mu\textrm{M}$) and arachidonic acid($100\mu\textrm{M}$) in concentration-dependent manner with $IC_{50}$ values of 6.5$\pm$1.3, 59.3$\pm$4.5, 13.0$\pm$2.37 and 2.9$\pm$$1.0\mu\textrm{M}$, respectively. Collagen-induced serotonin release was significantly reduced by TPN2. The elevation of intracellular free $Ca^{2+}$ concentration ([$Ca^{2+}$]i) by collagen stimulation was greatly decreased by the pretreatment of TPN2, which was due to the inhibition of calcium release from intracellular store and influx from outside of the cell. TPN2 also significantly reduced the thromboxane $A_2$($TXA_2$) formation in a concentration-dependent manner. The collagen-induced arachidonic acid (AA) release in [$^3H$]-AA incorporated platelet, an indicative of the phospholipase $A_2$ activity, was decreased by TPN2 pretreatment. TPN2 significantly inhibited the activity of thromboxane synthase, but did not affect the cyclooxygenase activity. From these results. it is suggested that TPN2 exert its antiplatelet activity through the inhibition of the intra-cellular $Ca^{2+}$ mobilization and the decrease of the $TXA_2$ synthesis.

• Journal of Ginseng Research
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• 제41권4호
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• pp.548-555
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• 2017

Background: Korean Red Ginseng has been used for several decades to treat many diseases, enhancing both immunity and physical strength. Previous studies have documented the therapeutic effects of ginseng, including its anticancer, antiaging, and anti-inflammatory activities. These activities are mediated by ginsenosides present in the ginseng plant. Ginsenoside Rg3, an effective compound from red ginseng, has been shown to have antiplatelet activity in addition to its anticancer and anti-inflammatory activities. Platelets are important for both primary hemostasis and the repair of the vessels after injury; however, they also play a crucial role in the development of acute coronary diseases. We prepared ginsenoside Rg3-enriched red ginseng extract (Rg3-RGE) to examine its role in platelet physiology. Methods: To examine the effect of Rg3-RGE on platelet activation in vitro, platelet aggregation, granule secretion, intracellular calcium ($[Ca^{2+}]_i$) mobilization, flow cytometry, and immunoblot analysis were carried out using rat platelets. To examine the effect of Rg3-RGE on platelet activation in vivo, a collagen plus epinephrine-induced acute pulmonary thromboembolism mouse model was used. Results: We found that Rg3-RGE significantly inhibited collagen-induced platelet aggregation and $[Ca^{2+}]_i$ mobilization in a dose-dependent manner in addition to reducing ATP release from collagen-stimulated platelets. Furthermore, using immunoblot analysis, we found that Rg3-RGE markedly suppressed mitogen-activated protein kinase phosphorylation (i.e., extracellular stimuli-responsive kinase, Jun N-terminal kinase, p38) as well as the PI3K (phosphatidylinositol 3 kinase)/Akt pathway. Moreover, Rg3-RGE effectively reduced collagen plus epinephrine-induced mortality in mice. Conclusion: These data suggest that ginsenoside Rg3-RGE could be potentially be used as an antiplatelet therapeutic agent against platelet-mediated cardiovascular disorders.

• Journal of Ginseng Research
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• 제35권2호
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• pp.209-218
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• 2011

Ginseng has been used as a general tonic agent to invigorate the human body as an adaptogenic agent. In a previous report, we have shown that ginseng contains a novel glycolipoprotein called gintonin. The main function of gintonin is to transiently enhance intracellular free $Ca^{2+}$ $[Ca^{2+}]_i$ levels in animal cells. The previous method for gintonin isolation included multiple steps using organic solvents. In the present report, we developed a simple method for the preparation of crude gintonin from ginseng root as well as stem and leaf, which produced a higher yield of gintonin than the previous one. The yield of gintonin was 0.20%, 0.29%, and 0.81% from ginseng root, stem, and leaf, respectively. The apparent molecular weight of gintonin isolated from stem and leaf through sodium dodecyl sulfate polyacrylamide gel electrophoresis was almost same as that from root but the compositions of amino acids, carbohydrates or lipids differed slightly between them. We also examined the effects of crude gintonin from ginseng root, stem, and leaf on endogenous $Ca^{2+}$-activated $Cl^-$ channel (CaCC) activity of Xenopus oocytes through mobilization of $[Ca^{2+}]_i$. We found that the order of potency for the activation of CaCC was ginseng root > stem > leaf. The $ED_{50}$ was $1.4{\pm}1.4$, $4.5{\pm}5.9$, and $3.9{\pm}1.1$ mg/mL for root, stem and leaf, respectively. In the present study, we demonstrated for the first time that in addition to ginseng root, ginseng stem and leaf also contain gintonin. Gintonin can be prepared from a simple method with higher yield of gintonin from ginseng root, stem, and leaf. Finally, these results demonstrate the possibility that ginseng stem and leaf could also be utilized for ginstonin preparation after a simple procedure, rather than being discarded.

• 대한치과보철학회지
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• 제52권2호
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• pp.105-112
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• 2014

연구 목적: 이 연구의 목적은 임플란트 식립 초기 수회의 의원성 동요가 최종 골결합에 미치는 영향을 확인하는 것이다. 연구 재료 및 방법: 순수한 티타늄(Grade IV) 으로 직경 3.75 mm, 길이 8 mm의 실험용 임플란트를 제작한 후, 하방 4 mm에 RBM 표면처리($MegaGen^{(R)}$: Ca-P)를 하였다. 임플란트의 하부만 골에 식립되었으며 비침하시켰다. 수술은 3.5 kg이상의 토끼의 좌우 경골의 단층 치밀골에 각각 2개씩 130개를 통상적인 방법으로 식립하였다(Female, New Zealand White). 비틀림 제거 간격에 따라, 다음과 같은 13개의 군으로 나누었다; Group I (1일), Group II (1일 + 2일), Group III (1일 + 2일 + 3일), Group IV (1일 + 2일 + 3일 + 4일), Group V (2일), Group VI (2일 + 4일), Group VII (2일 + 4일 + 6일), Group VIII (2일 + 4일 + 6일 + 8일), Group IX (4일), Group X (4일 + 7일), Group XI (4일 + 7일 + 10일), Group XII (4일 + 7일 + 10일 + 14일), 그리고 대조군. 대조군은 식립 후 8주의 치유기간을 준 후 최종 비틀림 제거력을 측정하였다(Mark-10, USA). 실험군은 각각의 조건에 맞추어 1회에서 4회에 걸쳐 비틀림 제거력을 측정하였다. 비틀림 제거력을 측정한 이후에 대체적으로 임플란트를 측정 전의 위치로 재위치시켰다. 모든 실험군에서 임플란트 식립 후 8주간의 치유기간을 준 다음 최종 비틀림 제거력을 측정하여 대조군, 각 실험군의 1, 2, 3, 4번째 비틀림 제거력과 비교하였다. 결과: 실험군 간에 최종 비틀림 제거력을 비교한 결과에서, XII군을 제외한 실험군의 비틀림 제거력은 대조군과 유의할만한 차이를 보이지는 않았다. 그리고 실험 I군과 II군의 값은 VI, VIII, X, XI, XII군의 값보다 유의하게 높게 나타났다. 그리고 III, IV, V군은 XI, XII군의 값보다 유의하게 높게 나타났다. 각 실험군간 비틀림 제거력 비교에서, 최종 비틀림 제거력은 VIII, X, XI, XII군을 제외한 모든 군에서 높게 나타났다. 결론: 충분한 치유기간이 주어진다면, 토끼에게 임플란트 식립 후 매우 이른 초기에 지대주에 가해지는 약간의 동요는 임플란트 골결합에 영향을 미치지 않았다.

• The Korean Journal of Physiology and Pharmacology
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• 제3권1호
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• pp.83-91
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• 1999

Angiotensin II (ANG II) has a biphasic effect on $Na^+$ transport in proximal tubule: low doses of ANG II increase the $Na^+$ transport, whereas high doses of ANG II inhibit it. However, the mechanisms of high dose ANG II-induced inhibition on $Na^+$ uptake are poorly understood. Thus the aim of the present study was to investigate signal transduction pathways involved in the ANG II-induced inhibition of $Na^+$ uptake in the primary cultured rabbit renal proximal tubule cells (PTCs) in hormonally defined serum-free medium. ANG II $(10^{-9}\;M)-induced$ inhibition of $Na^+$ uptake was blocked by losartan $(10^{-8}\;M,\;AT_1\;antagonist),$ but not by PD123319 $(10^{-8}\;M,\;AT_2\;antagonist)$ (P<0.05). ANG II-induced inhibition of $Na^+$ uptake was also completely abolished by neomycin $(10^{-4}\;M,$ PLC inhibitor), W-7 $(10^{-4}\;M,$ calmodulin antagonist), and $AACOCF_3\;(10^{-6}\;M,\;PLA_2\;inhibitor)$ (P<0.05). ANG II significantly increased $[^3H]arachidonic$ acid (AA) release compared to control. The ANG II-induced $[^3H]AA$ release was blocked by losartan, $AACOCF_3,$ neomycin, and W-7, but not by PD123319. ANG II-induced $[^3H]AA$ release in the presence of extracellular $Ca^{2+}$ was greater than in $Ca^{2+}-free$ medium, and it was partially blocked by TMB-8 $(10^{-4}\;M,$ intracelluar $Ca^{2+}$ mobilization blocker). However, in the absence of extracellular $Ca^{2+},$ it was completely blocked by TMB-8. In addition, econazole $(10^{-6}\;M,$ cytochrome P-450 monooxygenase inhibitor) and indomethacin $(10^{-6}\;M,$ cyclooxygenase inhibitor) blocked ANG II-induced inhibition of $Na^+$ uptake, but NGDA $(10^{-6}\;M,$ lipoxygenase inhibitor) did not affect it. In conclusion, $PLA_2-mediated$ AA release is involved in ANG II-induced inhibition of $Na^+$ uptake and is modulated by $[Ca^{2+}]_i$ in the PTCs.

• Journal of Microbiology and Biotechnology
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• 제7권4호
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• pp.229-236
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• 1997

A gene, $phbC_{2.4.1}$ encoding poly-3-hydroxybutyric acid (PHB) synthase of Rhodobacter sphaeroides 2.4.1 was cloned by employing heterologous expression in Escherichia coli. R. sphaeroides chromosomal DNA partially digested with MboI was cloned in pUC19 followed by mobilization into E. coli harbouring $phbA,B_{AC}$ in pRK415, which code for ${\beta}$-ketothiolase and acetoacetyl CoA reductase of Alcaligenes eutrophus, respectively. Two E. coli clones carrying R. sphaeroides chromosomal fragment of $phbC_{2.4.1}$ in pUC19 were selected from ca. 10,000 colonies. The PHB-producing colonies had an opaque white appearance due to the intracellular accumulation of PHB. The structure of PHB produced by the recombinant E. coli as well as from R. sphaeroides 2.4.1 was confirmed by [$H^{+}$]-nuclear magnetic resonance (NMR) spectroscopy. Restriction analysis of the two pUC19 clones revealed that one insert DNA fragment is contained as a part of the other cloned fragment. An open reading frame of 601 amino acids of $phbC_{2.4.1}$ with approximate M.W. of 66 kDa was found from nucleotide sequence determination of the 2.8-kb SaiI-PstI restriction endonuclease fragment which had been narrowed down to support PHB synthesis through heterologous expression in the E. coli harbouring $phbA,B_{AC}$. The promoter (s) of the $phbC_{2.4.1}$ were localized within a 340-bp DNA region upstream of the $phbC_{2.4.1}$ start codon according to heterologous expression analysis.

• Journal of Applied Biological Chemistry
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• 제62권1호
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• pp.93-100
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• 2019

The root of Panax ginseng is used in ethnomedicine throughout eastern Asia and various recent studies have proved that Panax ginseng has inhibitory effects on cardiovascular disease. Each factor causing cardiovascular disease is known to have a very complex process which is achieved by a diverse number of mechanisms. Among these factors, platelets are the most important because they directly participate in thrombogenesis. Therefore, inhibiting the activity of platelets is an essential element for prevention of cardiovascular diseases. Our previous study showed the antiplatelet effects of Korean red ginseng extract and two of its components, ginsenoside Rg3 and ginsenoside Ro. However, the inhibitory mechanism of other ginsenosides remains unclear. Therefore, we investigated the inhibitory mechanism of ginsenoside F4 (G-F4) from Korean red ginseng on the regulation of signaling molecules involved in human platelet aggregation. With the use of G-F4, collagen-induced human platelet aggregation was inhibited in a dose-dependent manner, and it suppressed collagen-induced elevation of $[Ca^{2+}]_i$ mobilization through elevated phosphorylation of inositol 1, 4, 5-triphosphate receptor I ($Ser^{1756}$). In addition, G-F4 inhibited fibrinogen binding to ${\alpha}IIb/{\beta}_3$ during collagen-induced human platelet aggregation. Thus, in the present study, G-F4 showed an inhibitory effect on human platelet activation, suggesting its potential use as a new natural medicine for preventing platelet-mediated cardiovascular diseases.