Phytochemical, Fluorescence Analysis, In vitro Antioxidant and In-vitro Thrombolytic Activity of Kabasura Kudineer Herbal Concoction

Objective: The purpose of the study is to carry out fluorescence analysis, phytochemical, in vitro antioxidant In-Vitro Thrombolytic Activity of Kabasura Kudineer Herbal Concoction. Methods: In the present study, fluorescence analysis, phytochemical, antioxidant In-Vitro Thrombolytic Activity of Kabasura Kudineer Herbal Concoction are carried out using standard procedures. Result: The fluorescence analysis under visible and ultraviolet light for Kabasura Kudineer herbal concoction treated with various chemical reagents shown different fluorescence effect. It showed a significant antioxidant activity in DPPH, Reducing Power and H2O2 scavenging methods. From our study, we also found that it showed 41.6% clot lysis activity respectively and they showed significant % of clot lysis effect with reference of Streptokinase (75.2%) and water (2.93%). Conclusion: The findings of the present study suggest that Kabasura Kudineer herbal concoction could be a potential source of natural antioxidant that could have greater importance as therapeutic agent in preventing has significant thrombolytic action. Additional studies are greatly essential for further drug development.

The oral thrombolysis for venous thrombosis (clinical trial)

Введение. Таргетная терапия тромбозов заключается в тромболитической терапии. В настоящий момент в подавляющем большинстве случаев терапия тромбозов у пациента сводится к назначению антикоагулянтов в расчете на механизм собственного фибринолиза/тромболизиса. Применение активаторов плазминогена эффективно, но сопряжено с опасностью тяжелых кровотечений и проводится только в условиях стационара. Очень перспективно для лечения тромбозов выглядит применение лекарственных препаратов на основе иммобилизированных субтилизинов, которые имеют прямое тромболитическое действие при пероральном приеме. В России зарегистрирован лекарственный препарат на основе иммобилизированных субтилизинов Тромбовазим, который имеет показание лечение хронической венозной недостаточности. Основой фармакодинамики препарата является тромболитическое действие. Цель исследования: показать, что включение лекарственного препарата Тромбовазим в комплексную терапию тромбозов венозного русла нижних конечностей улучшает результаты лечения. Материалы и методы. Проведено многоцентровое рандомизированное плацебоконтролируемое двойное слепое клиническое исследование. В настоящий момент лекарственный препарат Тромбовазим зарегистрирован к медицинскому применению в дозе 1600 ЕД/сут. Клиническое исследование было проведено по двум протоколам: Протокол 1 включение Тромбовазима в комплексную терапию тромбоза в суточной дозе 1600 ЕД Протокол 2 включение Тромбовазима в комплексную терапию тромбоза в более высоких дозах 3200 и 4800 ЕД/сут. Результаты. У пациентов, принимающих лекарственный препарат Тромбовазим в дозе 1600 3200 ЕД/сут, увеличение кровотока либо реканализацию вены наблюдали в 1,6 раза чаще (на 60 больше), чем среди пациентов, получающих плацебо. Эффективная тромболитическая доза препарата Тромбовазим 1600 3200 ЕД/сут. Заключение. Лекарственный препарат на основе иммобилизированных субтилизинов Тромбовазим обладает тромболитическим действием при пероральном применении. Тромбовазим в составе комплексной терапии больных с тромбозом венозных сосудов быстро восстанавливает кровоток в зоне скомпрометированного кровообращения. Introduction. The target therapy of thrombosis consists of thrombolytic therapy. At present in most of cases, the therapy of thromboses in patient is reduced to the prescribing of anticoagulants based on mechanism of own fibrinolysis/thrombolysis. The use of plasminogen activators is effective, but is associated with the risk of severe bleeding and is carried out only in hospital. The administration of drugs that based on immobilized subtilisins looks very promising for thromboses treatment. The immobilized subtilisins have a direct thrombolytic effect under oral administration. The drug Trombovazim is based on immobilized subtilisins and has been registered in Russia. Trombovazim has an indication treatment of chronic venous insufficiency thrombolytic action is the basis of its pharmacodynamics. Aim: to show that Trombovazim in complex therapy of venous thromboses of the lower extremities improves the treatment outcomes. Materials and methods. A multicenter randomized doubleblind placebocontrolled clinical trial was carried out. At present Trombovazim is registered for medical use at a dose of 1600 U/day. The clinical study was undertaken according to 2 protocols: Рrotocol 1 Trombovazim inclusion in complex therapy of thrombosis at a dose of 1600 U/day Рrotocol 2 Trombovazim inclusion in complex therapy of thrombosis in higher doses 3200 and 4800 U/day. Results. In patients taking Trombovazim at a dose of 1600 3200 U/day increased blood flow or vein recanalization was observed 1.6 times more often (60 more) than in patients receiving placebo. The effective thrombolytic dose of Trombovazim is 1600 3200 U/day. Conclusion. Trombovazim has a thrombolytic action under oral administration. Thrombovazim in complex therapy of patients with venous thrombosis quickly restores the blood flow in a zone of compromised blood circulation.

Essential Oil from Lippia origanoides (Verbenaceae): Haemostasis and Enzymes Activity Alterations

Background: The search for natural inhibitors of snake venom toxins is essential to supplement or even replace the serum therapy. The aim of this work was to evaluate the pharmacological properties of essential oil from Lippia origanoides Kunth. (Verbenaceae). Methods: The oil was extracted by hydrodistillation and the constituents were identified and quantified by GC-MS and GC-FID. The essential oil from L. origanoides was evaluated in hemolysis tests, on the activities of phospholipases A2 and serine proteases and in coagulation and thrombolysis induced by different snake venoms. Results: The major constituents of essential oil were carvacrol, p-cymene, γ-terpinene, and thymol. The oil inhibited approximately 10 % of the phospholipase A2 activity induced by Bothrops atrox, Bothrops jararaca, Bothrops jararacussu and Bothrops moojeni venoms and was not cytotoxic against erythrocytes. However, previous incubation of the oil with B. jararacussu, B. moojeni, and Crotalus durissus terrificus (C.d.t.) venoms resulted in potentiation of hemolytic activity (30 % and 50 % for 0.6 µL mL-1 and 1.2 µL mL-1, respectively). The essential oil presented a procoagulant effect on human citrated plasma, potentiated the thrombolytic action of proteases and phospholipases A2 present in B. jararacussu venom, and serine protease activity induced by B. jararaca and Lachesis muta venoms. When pre-incubated with the C.d.t. venom, however, prothrombotic activity was observed. Conclusion: The results obtained in this work amplify the pharmacological characterization of the essential oil from L. origanoides. However, new studies are fundamental to define the action mechanisms and determine pharmaceutical applications.

The mouse dorsal skinfold chamber as a model for the study of thrombolysis by intravital microscopy

SummaryAlthough intravital microscopy models of thrombosis in mice have contributed to dissect the mechanisms of thrombus formation and stability, they have not been well adapted to study long-term evolution of occlusive thrombi. Here, we assessed the suitability of the dorsal skinfold chamber (DSC) for the study of thrombolysis and testing of thrombolytic agents by intravital microscopy. We show that induction of FeCl3-induced occlusive thrombosis is achievable in microvessels of DSCs, and that thrombi formed in DSCs can be visualised by intravital microscopy using brightfield transmitted light, or fluorescent staining of thrombus components such as fibrinogen, platelets, leukocytes, and von Willebrand factor. Direct application of control saline or recombinant tissue-plasminogen activator (rtPA) to FeCl3-produced thrombi in DSCs did not affect thrombus size or induce recanalisation. However, in the presence of hirudin, rtPA treatment caused a rapid dose-dependent lysis of occlusive thrombi, resulting in recanalisation within 1 hour after treatment. Skin haemorrhage originating from vessels located inside and outside the FeCl3-injured area was also observed in DSCs of rtPA-treated mice. We further show that rtPA-induced thrombolysis was enhanced in plasminogen activator inhibitor-1-deficient (PAI-1−/−) mice, and dropped considerably as the time between occlusion and treatment application increased. Together, our results show that by allowing visualization and measurement of thrombus lysis and potential bleeding complications of thrombolytic treatments, the DSC provides a model for studying endogenous fibrinolysis and for first-line screening of thrombolytic agents. Furthermore, using this system, we found that thrombin and clot aging impair the thrombolytic action of rtPA towards FeCl3-produced thrombi.