RESUMO
When activated in vitro, thrombin-activatable fibrinolysis inhibitor (TAFI) slows clot lysis by cleaving the C-terminal lysine and arginine residues from partially degraded fibrin. An inhibitor of carboxypeptidase isolated from potato (CPI) reverses prolongation of clot lysis by inhibiting activated TAFI. We investigated in vivo effect of TAFI inhibition on tissue-type plasminogen activator (t-PA)-induced clot lysis using CPI in a rabbit jugular vein thrombolysis model. It was found necessary to further purify the CPI preparations from commercial sources by HPLC chromatography to remove endotoxin and anti-plasmin activity that would affect the endogenous fibrinolytic system. The effect of intravenous administration of the purified CPI with t-PA was determined by measuring thrombus weight at the end of 90 minutes in six groups of animals. In the control group receiving saline, the median thrombus weight was 116 mg. In the group that received CPI only (0.5 mg/kg bolus injection followed by 0.3 mg/kg/h infusion), the median thrombus weight was 121 mg. In the group that received t-PA at a dose of 10 microg/kg bolus followed by 67 microg/kg/h infusion, the median thrombus weight decreased to 86 mg. When CPI was coadministered with the same regimen of t-PA, the median value further decreased to 58 mg. When animals were given three times higher the dose of t-PA (30 microg/kg bolus followed by 200 microg/kg/h infusion) in the absence or presence of CPI, median thrombus weights were 56 mg and 0 mg, respectively. Our results demonstrate that systemic coadministration of the purified CPI improves clot lysis induced by t-PA.
Assuntos
Carboxipeptidases/antagonistas & inibidores , Fibrina/metabolismo , Fibrinólise/efeitos dos fármacos , Proteínas de Plantas/uso terapêutico , Trombina/fisiologia , Terapia Trombolítica , Ativador de Plasminogênio Tecidual/uso terapêutico , Trombose Venosa/tratamento farmacológico , Animais , Carboxipeptidase B2 , Carboxipeptidases/fisiologia , Cromatografia Líquida de Alta Pressão , Avaliação Pré-Clínica de Medicamentos , Sinergismo Farmacológico , Endotélio Vascular/fisiopatologia , Ativação Enzimática/efeitos dos fármacos , Fibrinogênio/metabolismo , Infusões Intravenosas , Proteínas de Plantas/administração & dosagem , Proteínas de Plantas/isolamento & purificação , Proteínas de Plantas/farmacologia , Plasminogênio/metabolismo , Inibidores de Proteases , Coelhos , Solanum tuberosum/enzimologia , Ativador de Plasminogênio Tecidual/farmacologia , alfa 2-Antiplasmina/metabolismoRESUMO
We have identified a novel human gene, kiap (kidney inhibitor of apoptosis protein) that encodes a single BIR domain and a RING zinc finger domain. kiap has been assigned to the q13.3 region of human chromosome 20 by fluorescent in situ hybridization analysis. Northern blot analysis indicates that KIAP is expressed mainly in placenta, lymph node and fetal kidney. In this report, we show that overexpression of KIAP blocks apoptosis induced by menadione or by overexpression of BAX. In addition, we show that overexpression of KIAP enhances apoptosis induced by etoposide, and, that KIAP fails to block apoptosis induced by overexpression of Fas. Thus, KIAP, a new member of the inhibitor of apoptosis protein (IAP) family, has pleiotropic effects on apoptosis induced by various stimuli.
Assuntos
Apoptose/fisiologia , Proteínas de Bactérias/genética , Proteínas de Insetos , Rim/fisiologia , Proteínas , Sequência de Aminoácidos , Proteínas de Bactérias/classificação , Proteínas de Bactérias/isolamento & purificação , Proteínas de Bactérias/metabolismo , Sequência de Bases , Mapeamento Cromossômico , Cromossomos Humanos Par 20 , Clonagem Molecular , DNA Complementar/análise , Bases de Dados Factuais , Humanos , Proteínas Inibidoras de Apoptose , Cariotipagem , Rim/citologia , Dados de Sequência Molecular , Filogenia , Estrutura Terciária de Proteína , RNA Mensageiro/metabolismo , Homologia de Sequência de Aminoácidos , Distribuição Tecidual , Proteínas Virais/química , Receptor fas/biossíntese , Receptor fas/fisiologiaRESUMO
Thrombomodulin is an endothelial surface thrombin receptor. Thrombin bound to thrombomodulin loses all procoagulant activity and instead activates the protein C anticoagulant pathway. We developed a recombinant thrombomodulin analog and compared the effects of recombinant thrombomodulin (100 micrograms/ea), saline (controls), recombinant hirudin (1.0 mg/kg), and heparin (100 units/kg) on thrombus formation, activated partial thromboplastin time, and tail transection bleeding time in a rat model of stasis-induced venous thrombosis. Results showed that thrombus was detected in the vena cava in six of the six rats treated with saline solution, in zero of the six rats treated with recombinant thrombomodulin (p less than 0.05), in one of six rats treated with recombinant hirudin (p less than 0.05), and in zero of six rats treated with heparin (p less than 0.05). The activated partial thromboplastin time in rats receiving recombinant thrombomodulin was slightly longer than controls (22 +/- 8 vs 37 +/- 6, p less than 0.05). The bleeding times in rats receiving recombinant thrombomodulin were approximately twice as long as controls (215 +/- 68 vs 545 +/- 173, p = 0.037). In all rats treated with recombinant hirudin or heparin, activated partial thromboplastin times were greater than 120 seconds and bleeding times were greater than 1200 seconds. We conclude that recombinant thrombomodulin inhibits venous thrombosis in a rat model with less prolongation of activated partial thromboplastin time and bleeding time than heparin or hirudin.