Navegando por Palavras-chave "Tempo De Coagulação"

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    Expressão heteróloga, purificação e estudo de efeitos da rSculptina na coagulação sanguínea
    (Universidade Federal de São Paulo (UNIFESP), 2021) Viana, Juliana Da Silva [UNIFESP]; Tavassi, Ana Marisa Chudzinski [UNIFESP]; Universidade Federal de São Paulo
    Objective: To investigate the action of recombinant Sculptin as an anticoagulant and a inhibition of thrombin-induced platelet activation. Methods: E. coli strain BL21 (DE3) containing plasmid Sculptin-Pet28a was cultivated with IPTG induction. After expression, the culture was lysed and the supernatant subjected to affinity chromatography, another ion exchange and finally gel filtration. The purified protein profile was monitored during these steps using SDS-Page. Blood clotting time, partially activated thromboplastin time (aPTT), prothrombin time (PT) and thrombin time (TT) assays were evaluated in the presence of rSculptin using diagnostic kits. The evaluation of the effect of rSculptin on platelets was performed in a platelet aggregation assay. Furthermore, the action of the recombinant protein was evaluated ex vivo with aPTT and PT assays after animal treatment. Finally, an in vivo bleeding time test was performed. Results: The purified protein showed activity on aPTT, PT and TT evaluated in vitro, extending them in a dose response, but with greater sensitivity for TT. It also inhibited platelet aggregation when thrombin was used as an agonist. Furthermore, when rSculptin was administered to animals at a concentration of 1 mg/kg, it significantly prolonged the clotting time of the plasmas evaluated in the aPTT and PT assays, the same was not observed for the concentration of 0.1 mg/kg. Finally, in the preliminary in vivo bleeding time assay, the studied protein prolonged the clotting time of treated animals at the concentration evaluated (1 mg/kg). Conclusion: The results obtained demonstrate that rSculptin has action on the clotting times evaluated in animal plasma. Furthermore, the essays shows the maintenance of the biological activity of this protein on blood clotting after it been administered to animals. Although preliminary, the results obtained in the assessment of bleeding time suggest an anticoagulant potential of rSculptin in vivo.