• 대한본초학회지
• /
• 제26권4호
• /
• pp.125-132
• /
• 2011

Objectives : An aim of study is to investigate effects of curculiginis rhizoma in vitro (factor Xa (FXa) inhibitor assay, prothrombinase assay, prothrombin time (PT) assay, activated partial thromboplastin time (aPTT) assay) and in vivo experiment (blood clotting time, thromboxane B2 content assay in serum and weight of thrombus by AV-shunt rat model). Methods : We gained a human serum and used serum in vitro study such as factor X activity (FXa) inhibition, prothrombinase inhibition, prothrombin time (PT) and activated partial thromboplastin time. Fifteen SD rats were divided into three groups (intact control group and two experimental group treated with extract of Curculiginis Rhizoma(ECR)). Rats were orally administrated DW (intact control group), 600 mg/kg concertration of ECR and 200 mg/kg concertration of ECR. After one hour, we anesthetized rats and made arteriovenous (AV) shunt rat models to study weights of thrombus, took a hole blood to study content of thromboxane B2 and blood clotting time. Results : In vitro, ECR increased a inhibitory activity of FXa, prothrombinase and aPTT compared than intact control group. Especially ECR made significant increase of FXa and prothrombinase inhibitory activity (p<0.05, p<0.01). And PT were increased in ECR control group compared with intact control group. In vivo, a blood clotting time of experiment group treated with ECR 600 mg/kg were significantly increased compared with that of intact control group (p<0.05) and content of thromboxane B2 was significantly decreased in group treated with ECR 600 mg/kg in seum. The weight of thrombus were significantly reduced in group treated with ECR 600 mg/kg compared with intact control group (p<0.05). But in vivo experiment study, those of group treated with ECR 200 mg/kg were reduced compared with those of intact control group without statistical significance. Conclusions : ECR has a antithromboic activity in internal course with inhibitory activity of FXa and prothrombinase in vitro, it required to research more study for effective compounds.

• 생명과학회지
• /
• 제13권4호
• /
• pp.463-472
• /
• 2003

제5인자와 지질막 phosphatidylserine과의 상호작용은 prothrombinase 복합체의 활성을 조절하는데 중요하다. 본 연구에서 제5인자의 지질 결합부위에 위치한 Trp2063과 Trp2064를 동시에 돌연변이 시킨 재조합 제5인자를 과발현 시키고 정제하였다. 돌연변이된 제5인자는 1-10%의 phosphatidylserine을 포함하는 지질막에서 아주낮은 활성을 보였다. surface plasmon resonance에 의해서 지질막과의 결합을 측정한 결과 돌연변이된 제5인자가 본래의 제5인자보다 고정된 지질막에의 결합이 현저하게 떨어지는 것을 관찰하였다. 제5인자가 phosphatidylserine을 포함하는 지질막에 높은 친화력으로 결합하기 위해서는 Trp2063과 Trp2064가 필수적이고 이러한 상호작용은 생리적인 phosphatidylserine 농도를 포함하는 지질막 위에서 prothrombinase 복합체의 형성에 필요하다는 결론을 내렸다.

• 대한본초학회지
• /
• 제26권4호
• /
• pp.139-147
• /
• 2011

Objectives : The aim of this study is to research an anti-thrombus effect by Diospyros Kaki Calyx. Methods : The healthy human plasma were gained and used in vitro study such as factor X activity (FXa) inhibition, prothrombinase inhibition, prothrombin time (PT) and activated partial thromboplastin time. Fifteen SD rats were divided into three groups ; intact control group (orally administrated with distilled water 5ml/kg) and two experimental group treated with extract of diospyros kaki calyx (EKC). Experimental rats were orally 600 mg/kg concentration of EKC and 200 mg/kg concentration of EKC. After an hour from administration, we anesthetized rats and made arteriovenous (AV) shunt rat models to study weight of thrombus, took whole blood to study content of thromboxane B2 and blood clotting time. Results : In vitro, EKC significantly increased inhibitory activity of FXa, prothrombinase compared with intact control group ($^*P$ <0.05). PT and aPTT were increased in EKC treated (600 mg/kg) group compared with intact control group ($^*P$ <0.05). In vivo, blood clotting time of experiment group treated with EKC 600 mg/kg were significantly increased compare with that of intact control group (p<0.05) and content of thromboxane B2 was significantly decreased in group treated with EKC 600 mg/kg in serum. The weight of thrombus were significantly reduced in group treated with EKC 600 mg/kg compared with intact control group (p<0.05). But in vivo experiment study, those parameters of group treated with EKC 200 mg/kg were relatively decreased compared with those of intact control group without statistical significance. Conclusions : EKC has an antithrombic activity because of inhibition internal course such as FXa and prothrombin. And EKC inhibited a hole blood clotting in vivo experiment by low content of thromboxane B2.

• BMB Reports
• /
• 제40권5호
• /
• pp.832-838
• /
• 2007

A novel inhibitory protein against blood coagulation factor Va (FVa) was purified from muscle protein of granulated ark (Tegillarca granosa, order Arcoida, marine bivalvia) by consecutive FPLC method using anion exchange and gel permeation chromatography. In the results of ESI-QTOF tandem mass analysis and database research, it was revealed that the purified T. granosa anticoagulant protein (TGAP) has 7.7 kDa of molecular mass and its partial sequence, HTHLQRAPHPNALGYHGK, has a high identity (64%) with serine/threonine kinase derived from Rhodopirellula baltica (order Planctomycetales, marine bacteria). TGAP could potently prolong thrombin time (TT), corresponding to inhibition of thrombin (FIIa) formation. Specific factor inhibitory assay showed that TGAP inhibits FVa among the major components of prothrombinase complex. In vitro assay for direct-binding affinity using surface plasmon resonance (SPR) spectrometer indicated that TGAP could be directly bound with FVa. In addition, the binding affinity of FVa to FII was decreased by addition of TGAP in dose-dependant manner ($IC_{50}$ value = 77.9 nM). These results illustrated that TGAP might interact with a heavy chain of FVa ($FVa_H$) bound to FII in prothrombin complex. The present study elucidated that non-cytotoxic T. granosa anticoagulant protein (TGAP) bound to FVa can prolong blood coagulation time by inhibiting conversion of FII to FIIa in blood coagulation cascade. In addition, TGAP did not significantly (P < 0.05) show fibrinolytic activity and cytotoxicity on venous endothelial cell line (ECV 304).