RESUMO
Lopap, found in the bristles of Lonomia obliqua caterpillar, is the first exogenous prothrombin activator that shows serine protease-like activity, independent of prothrombinase components and unique lipocalin reported to interfere with hemostasis mechanisms. To assess the action of an exogenous prothrombin activator reversing the anticoagulant and antihemostatic effect induced by low molecular weight heparin (LMWH), male New Zealand rabbits (N = 20, weighing 3.8-4.0 kg) allocated to 4 groups were anticoagulated with 1800 IU/kg LMWH (iv) over 2 min, followed by iv administration of saline (SG) or recombinant Lopap (rLopap) at 1 µg/kg (LG1) or 10 µg/kg (LG10), 10 min after the injection of LMWH, in a blind manner. Control animals (CG) were treated only with saline. The action of rLopap was assessed in terms of activated partial thromboplastin time (aPTT), prothrombin fragment F1+2, fibrinogen, and ear puncture bleeding time (BT) at 5, 10, 15, 17, 20, 30, 40, 60, and 90 min after initiation of LMWH infusion. LG10 animals showed a decrease of aPTT in more than 50% and BT near to normal baseline. The level of prothrombin fragment F1+2 measured by ELISA had a 6-fold increase with rLopap treatment (10 µg/kg) and was inversely proportional to BT in LMWH-treated animals. Thus, Lopap, obtained in recombinant form using E. coli expression system, was useful in antagonizing the effect of LMWH through direct prothrombin activation, which can be a possible strategy for the reversal of bleeding and anticoagulant events.
Assuntos
Animais , Masculino , Coelhos , Anticoagulantes/uso terapêutico , Coagulação Sanguínea/efeitos dos fármacos , Hemorragia/tratamento farmacológico , Antagonistas de Heparina/farmacologia , Heparina de Baixo Peso Molecular/antagonistas & inibidores , Protrombina/efeitos dos fármacos , Serina Endopeptidases/farmacologia , Hemorragia/induzido quimicamenteRESUMO
Lopap, found in the bristles of Lonomia obliqua caterpillar, is the first exogenous prothrombin activator that shows serine protease-like activity, independent of prothrombinase components and unique lipocalin reported to interfere with hemostasis mechanisms. To assess the action of an exogenous prothrombin activator reversing the anticoagulant and antihemostatic effect induced by low molecular weight heparin (LMWH), male New Zealand rabbits (N = 20, weighing 3.8-4.0 kg) allocated to 4 groups were anticoagulated with 1800 IU/kg LMWH (iv) over 2 min, followed by iv administration of saline (SG) or recombinant Lopap (rLopap) at 1 µg/kg (LG1) or 10 µg/kg (LG10), 10 min after the injection of LMWH, in a blind manner. Control animals (CG) were treated only with saline. The action of rLopap was assessed in terms of activated partial thromboplastin time (aPTT), prothrombin fragment F1+2, fibrinogen, and ear puncture bleeding time (BT) at 5, 10, 15, 17, 20, 30, 40, 60, and 90 min after initiation of LMWH infusion. LG10 animals showed a decrease of aPTT in more than 50% and BT near to normal baseline. The level of prothrombin fragment F1+2 measured by ELISA had a 6-fold increase with rLopap treatment (10 µg/kg) and was inversely proportional to BT in LMWH-treated animals. Thus, Lopap, obtained in recombinant form using E. coli expression system, was useful in antagonizing the effect of LMWH through direct prothrombin activation, which can be a possible strategy for the reversal of bleeding and anticoagulant events.
Assuntos
Anticoagulantes/uso terapêutico , Coagulação Sanguínea/efeitos dos fármacos , Hemorragia/tratamento farmacológico , Antagonistas de Heparina/farmacologia , Heparina de Baixo Peso Molecular/antagonistas & inibidores , Protrombina/efeitos dos fármacos , Serina Endopeptidases/farmacologia , Animais , Hemorragia/induzido quimicamente , Masculino , CoelhosRESUMO
AIM: Vein reconstruction using grafts may prevent sequelae of venous interruption or lesion. Autologous vein is sometimes unsuitable or absent for a vascular restoration. The aim of this study was to study glutaraldehyde-treated homologous vein graft as vein substitute and compare it with autologous vein as a substitute for a vena cava segment in rabbits. METHODS: Sixty rabbits were allocated into two groups: autologous vein graft (AG), and glutaraldehyde-treated homologous vein graft (HG). Each group was subdivided into three subgroups (N.=10) to be studied at: 24 hours, 14 days, and 28 days. The veins were treated in 0.19% glutaraldehyde, pH=7.4, for 1 hour and kept at 4 degrees C in saline with added gentamicin and amphotericin B. The animals received benzanthine penicillin on the day of graft implantation and heparin only during surgery. The grafts were implanted into the vena cava. Anastomosis was performed with interrupted sutures. Cavography was performed, after surgery, and at the time the animals were killed. Evaluation of the veins was made macroscopically and by light and scanning electron microscopy. RESULTS: Fibrosis was seen around the grafts at 14 and 28 days, with no difference in intensity between the groups. Cavography performed before euthanasia of the animals showed 4 partial thrombi in AG (2 at 24 hours and 2 at 14 days), 3 in HG (2 at 24 hours and 1 on day 14), and 4 occlusive thombi in HG (3 at 14 days and 1 at 28 days). Macroscopic examination did not show any thrombus in AG. In HG, two partial thrombi were confirmed at 24 hours and three occlusive thrombi at 14 days. There was no statistical difference in relation to patency between the two groups. At 14 and 28 days, the histological sections showed intimal hyperplasia of similar intensity and variable distribution in both groups. Evaluation by electron microscopy showed at 24 hours lesion areas characterized by absence of the endothelium on the graft surface, presence of inflammatory cells, and, at some sites, presence of mural thrombi in AG and HG. Both groups at 14 and 28 days showed endothelial cells covering the lesion area on the graft surface, this covering being larger in AG than in HG. CONCLUSIONS: In the studied model, both grafts behaved similarly in relation to patency and morphological characteristics. This suggests that the glutaraldehyde-treated graft can be a promising alternative for vein reconstruction, justifying further animal studies with the aim of using it in human surgery.
Assuntos
Bioprótese , Implante de Prótese Vascular/instrumentação , Prótese Vascular , Fixadores , Glutaral , Fixação de Tecidos/métodos , Veia Cava Inferior/transplante , Animais , Flebografia , Desenho de Prótese , Coelhos , Fatores de Tempo , Transplante Autólogo , Transplante Homólogo , Grau de Desobstrução Vascular , Veia Cava Inferior/diagnóstico por imagem , Veia Cava Inferior/patologia , Veia Cava Inferior/fisiopatologiaRESUMO
AIM: This study aimed at assessing the accuracy of ultrasound (US) in the diagnosis of recent deep vein thrombosis (DVT) in an experimental study in dogs. DESIGN: blinded and randomized experimental study. Twenty dogs were randomly divided in two groups: control group (CG) and thrombosis group (TG). US was performed in the pre- and postoperative period. Phlebography was performed immediately prior to the postoperative US. After the second US, a surgery was performed to detect whether thrombus was present or not. US results were compared to those of phlebography and surgical findings. RESULTS: In all dogs, inferior vena cava (IVC) was compressible. The relations of IVC diameter with the aorta were higher (P<0.005) in TG than in CG. Spectral Doppler in spontaneous breathing, tissue harmonic imaging, power Doppler and B flow showed sensitivity, specificity and accuracy of 1. Phlebography presented sensitivity of 90%, specificity of 80% and accuracy of 85%, when compared to surgical finding. CONCLUSIONS: For the diagnosis of recent DVT in the experimental model used, venous compressibility proved to be inefficient. The ratio of IVC diameter to aorta, when increased, suggests thrombosis. The use of new US technological advances increases accuracy. Phlebography was less accurate than US.
Assuntos
Ultrassonografia Doppler , Veia Cava Inferior/diagnóstico por imagem , Trombose Venosa/diagnóstico por imagem , Animais , Modelos Animais de Doenças , Cães , Diagnóstico Precoce , Feminino , Masculino , Flebografia , Valor Preditivo dos Testes , Sensibilidade e Especificidade , Fatores de Tempo , Ultrassonografia Doppler em Cores , Veia Cava Inferior/cirurgia , Trombose Venosa/cirurgiaRESUMO
AIM: The authors assess a modified Greenfield filter (GF) for the long-term patency, filter tilting and histopathologic alterations of the inferior vena cava (IVC). METHODS: Adult sheep (n=7) underwent modified GF placement in the IVC. Cavograms were obtained every 3 months and pulmonary angiography at 12 months. Histopathologic and scanning electron microscopy (SEM) analyses were performed on the IVC explanted at 12 months. RESULTS: Cavograms showed that all IVC were patent at the end of the study. Filter tilting occurred in 2/7 animals and extrusion of struts was not observed. Macroscopic examination at explantation showed minimal venous wall thickening. Microscopic examination showed minimal IVC fibrosis and intimal hyperplasia. SEM showed endothelium on the IVC surface at the filter implantation site and a presumed endothelial layer covering partially or totally the struts. The interface filter-IVC was covered by deposits of leucocytes and platelets. No signs of pulmonary embolism were found in all pulmonary angiograms of both groups. CONCLUSION: The modified filter presented good biocompatibility, stability and absence of thrombogenicity at 12 months. It presented low tendency to tilting and extrusion of struts. The long-term histopathologic alterations in vena caval wall were minimal and the appearance of the studied filters in the IVC was similar to stents placed in the arterial system.
Assuntos
Filtros de Veia Cava , Veia Cava Inferior/patologia , Animais , Materiais Biocompatíveis , Feminino , Masculino , Microscopia Eletrônica de Varredura , Desenho de Prótese , Ovinos , Grau de Desobstrução VascularRESUMO
AIM: Duplex scanning has been used in the evaluation of the aorta and proximal arteries of the lower extremities, but has limitations in evaluating the arteries of the leg. The utilization of ultrasonographic contrast (USC) may be helpful in improving the quality of the image in these arteries. The objective of the present study was to verify whether the USC increases the diagnostic accuracy of patency of the leg arteries and if it diminishes the time needed to perform duplex scanning. METHODS: Twenty patients with critical ischemia (20 lower extremities) were examined by standard duplex scanning, duplex scanning with contrast and digital subtraction arteriography (DSA). The 3 arteries of the leg were divided into 3 segments, for a total of 9 segments per limb. Each segment was evaluated for patency in order to compare the 3 diagnostic methods. Comparison was made between standard duplex scanning and duplex scanning with contrast in terms of quality of the color-coded Doppler signal and of the spectral curve, and also of the time to perform the exams. RESULTS: Duplex scanning with contrast was similar to arteriography in relation to patency diagnosis (p>0.3) and even superior in some of the segments. Standard duplex scanning was inferior to arteriography and to duplex scanning with contrast (p<0.001). There were improvements of 70% in intensity of the color-coded Doppler signal and 76% in the spectral curve after the utilization of contrast. The time necessary to perform the examinations was 23.7 minutes for standard duplex scanning and 16.9 minutes for duplex scanning with contrast (p<0.001). CONCLUSIONS: The use of ultrasonographic contrast increased the accuracy of the diagnosis of patency of leg arteries and diminished the time necessary for the execution of duplex scanning.
Assuntos
Angiografia Digital , Meios de Contraste/administração & dosagem , Isquemia/diagnóstico , Perna (Membro)/irrigação sanguínea , Ultrassonografia Doppler Dupla , Adulto , Idoso , Idoso de 80 Anos ou mais , Artérias/diagnóstico por imagem , Artérias/fisiopatologia , Feminino , Humanos , Aumento da Imagem , Isquemia/diagnóstico por imagem , Isquemia/fisiopatologia , Perna (Membro)/diagnóstico por imagem , Masculino , Pessoa de Meia-Idade , Grau de Desobstrução VascularRESUMO
AIM: Autologous vein (AV) is sometimes not suitable or present for a vascular restoration. Homologous vein preserved in glutaraldehyde may be an alternative to AV, but little is yet known about this graft and its healing process after implantation in arteries. The purpose of this study was to compare the initial healing process of glutaraldehyde-tanned homologous venous grafts (group 1) with fresh autologous venous grafts (group 2), at 4 or 15 days. METHODS: Forty Norfolk rabbits were allocated in 2 groups of 20 animals each. The grafts was interposed in the infrarenal aorta of the rabbit. Anastomotic tensile strength (TS), hydroxyproline (HP) determination, and histology (HA) were performed. RESULTS: TS increased in both groups, from the 4th to 15th day, (p<0.01) in both proximal (G1: from 364.5+/-98.3 g to 491.8+/-107.3 g; G2: from 366.26+/-85.15 g to 518.46+/-82.79 g) and distal anastomosis (G1: from 363.53+/-96.26 g to 507.32+/-91.01 g; G2: from 352.30+/-102.41 g to 528.67+/-48.58 g), with no difference between the groups. HP did not change (p>0.10) in this same period and was similar in both groups, in the proximal (G1: from 677.99+/-153.98 microg/100 mg to 914.92+/-459.83 microg/100 mg; G2: from 668.65+/-170.28 microg/100 mg to 669.46+/-319.80 ug/100 mg) as well as in the distal anastomosis (G1: from 740.07+/-213.53 microg/100 mg to 923.52+/-270.57 microg/100 mg; G2: from 737.66+/-266.76 microg/100 mg to 707.68+/-171.25 microg/100 mg). Initial inflammatory and reparative features of the anastomosis were similar in both groups. CONCLUSION: We can conclude that the healing process of the glutaraldehyde-tanned homologous vein graft was similar to that of the fresh autologous venous graft.
Assuntos
Aorta/fisiopatologia , Aorta/transplante , Bioprótese , Prótese Vascular , Veias/transplante , Cicatrização/fisiologia , Anastomose Cirúrgica , Animais , Hidroxiprolina/farmacologia , Modelos Animais , Modelos Cardiovasculares , Complicações Pós-Operatórias/diagnóstico por imagem , Complicações Pós-Operatórias/etiologia , Complicações Pós-Operatórias/fisiopatologia , Coelhos , Radiografia , Resistência à Tração/fisiologia , Fatores de Tempo , Transplante Autólogo , Transplante Homólogo , Grau de Desobstrução Vascular/fisiologia , Veias/fisiopatologiaRESUMO
The venom of Lonomia obliqua caterpillar may induce a hemorrhagic syndrome in humans, and blood incoagulability by afibrinogenemia when intravenously injected in laboratory animals. The possible antithrombotic and thrombolytic activities of L. obliqua caterpillar bristle extract (LOCBE) were evaluated in this study. The minimal intravenous dose of the extract necessary to induce afibrinogenemia and anticoagulation was 3.0 and 10.0 microg protein/kg body weight for rabbits and rats, respectively. In rabbits, this dose induced total blood incoagulability for at least 10 h and did not reduce the weight of preformed venous thrombi, in contrast to streptokinase (30,000 IU/kg). In rats, pretreatment with 5.0 and 10.0 microg/kg LOCBE prevented the formation of thrombi induced by venous stasis or by injury to the venous endothelium. The dose of 5.0 microg/kg LOCBE did not modify blood coagulation assay parameters but increased bleeding time and decreased plasma factor XIII concentration. When the extract was administered to rats at the dose of 10.0 microg/kg, the blood was totally incoagulable for 6 h. These data show that LOCBE was effective in preventing experimental venous thrombosis in rats, justifying further studies using purified fractions of the extract to clarify the mechanisms of this effect.
Assuntos
Anticoagulantes/farmacologia , Venenos de Artrópodes/farmacologia , Coagulação Sanguínea/efeitos dos fármacos , Fibrinolíticos/farmacologia , Trombose Venosa/prevenção & controle , Animais , Anticoagulantes/uso terapêutico , Venenos de Artrópodes/uso terapêutico , Tempo de Sangramento , Fator XIII/análise , Fibrinolíticos/uso terapêutico , Veias Jugulares/efeitos dos fármacos , Masculino , Coelhos , Ratos , Ratos Wistar , Veias Cavas/efeitos dos fármacosRESUMO
The venom of Lonomia obliqua caterpillar may induce a hemorrhagic syndrome in humans, and blood incoagulability by afibrinogenemia when intravenously injected in laboratory animals. The possible antithrombotic and thrombolytic activities of L. obliqua caterpillar bristle extract (LOCBE) were evaluated in this study. The minimal intravenous dose of the extract necessary to induce afibrinogenemia and anticoagulation was 3.0 and 10.0 æg protein/kg body weight for rabbits and rats, respectively. In rabbits, this dose induced total blood incoagulability for at least 10 h and did not reduce the weight of preformed venous thrombi, in contrast to streptokinase (30,000 IU/kg). In rats, pretreatment with 5.0 and 10.0 æg/kg LOCBE prevented the formation of thrombi induced by venous stasis or by injury to the venous endothelium. The dose of 5.0 æg/kg LOCBE did not modify blood coagulation assay parameters but increased bleeding time and decreased plasma factor XIII concentration. When the extract was administered to rats at the dose of 10.0 æg/kg, the blood was totally incoagulable for 6 h. These data show that LOCBE was effective in preventing experimental venous thrombosis in rats, justifying further studies using purified fractions of the extract to clarify the mechanisms of this effect
Assuntos
Animais , Ratos , Coelhos , Masculino , Anticoagulantes , Venenos de Artrópodes , Coagulação Sanguínea , Fibrinolíticos , Trombose Venosa , Anticoagulantes , Venenos de Artrópodes , Tempo de Sangramento , Fator XIII , Fibrinolíticos , Veias Jugulares , Ratos Wistar , Veias CavasRESUMO
BACKGROUND AND OBJECTIVES: Thrombin activatable fibrinolysis inhibitor (TAFI) plays an important role in hemostasis, functioning as a potent fibrinolysis inhibitor. TAFI gene variations may contribute to plasma TAFI levels and thrombotic risk. DESIGN AND METHODS: We sequenced a 2083-bp region of the 5'-regulatory region of the TAFI gene in 127 healthy subjects searching for variations, and correlated identified polymorphisms with plasma TAFI levels. TAFI polymorphisms were examined as risk factors for venous thrombosis by determining their prevalence in 388 patients with deep venous thrombosis (DVT) and in 388 controls. RESULTS: Seven novel polymorphisms were identified: -152 A/G, -438 A/G, -530 C/T, -1053 T/C, -1102 T/G, -1690 G/A, and -1925 T/C. -152 A/G, -530 C/T and -1925 T/C were found to be in strong linkage disequilibrium, as were the -438 A/G, -1053 T/C, -1102 T/G and -1690 G/A. Plasma TAFI levels were higher in -438GG/-1053CC/-1102GG/-1690AA homozygotes than in -438AG/-1053TC/-1102TG/-1690GA heterozygotes, and -438AA/-1053TT/-1102TT/-1690GG homozygotes had the lowest TAFI levels (p=0.0003). TAFI concentrations in -152AA/-530CC/-1925TT homozygotes were somewhat higher but not significantly different from levels observed for -152AG/-530CT/-1925TC heterozygotes. Taken in combination, -438AG/-1053TC/-1102TG/-1690GA and -438AA/-1053TT/-1102TT/-1690GG yielded an OR for DVT of 0.8 (95%CI: 0.6-1). In subjects aged <35 years the OR was 0.7 (95%CI: 0.5-1.1). The OR for -152AG/-530CT/-1925TC was 1 (95%CI: 0.5-2.2) in the whole group of patients and controls, whereas in subjects aged <35 years the OR was 0.1 (95%CI: 0.02-0.9). INTERPRETATION AND CONCLUSIONS: Polymorphisms in the TAFI promoter determine plasma antigen levels and may influence the risk of venous thrombophilia.