RESUMO
Fibrinogen Bicêtre II is a dysfibrinogenemia in which there is a substitution of Lys for Asn at gamma 308. We have studied the polymerization of this abnormal fibrinogen by measurement of turbidity and have characterized clot structure by scanning electron microscopy, permeation, and viscoelastic measurements. The results of these studies demonstrate that this amino acid substitution has substantial effects on the structure and properties of the clot, resulting in clots made up of thick fibers and large pores with greatly reduced stiffness and increased slippage of protofibrils.
Assuntos
Fibrinogênios Anormais/química , Fibrinogênios Anormais/metabolismo , Fibrinogênios Anormais/genética , Humanos , MutaçãoRESUMO
The fibrino(geno)lytic protein designated hementerin contained in crude extracts of the salivary complex of Haementeria depressa leeches was purified to apparent homogeneity by gel filtration, ion exchange chromatography and preparative SDS-PAGE. It is a single-chain 80 kDa, PhMeSO2F-resistant, calcium-dependent, metalloproteinase, which specifically degrades fibrin(ogen) through a plasminogen-independent pathway. The amino terminal sequence of 8 residues shows 80% similarity with hementin, another fibrino(geno)lytic protein purified from Haementeria ghilianii leeches. However, their activities differ somewhat in terms of kinetics and with regard to the structure of the fibrin(ogen) fragments they may produce. Cleavage by hementerin of fibrinogen Aalpha, gamma and Bbeta chains, in that order, produces 270 kDa to 67 kDa fragments which differ from those produced by plasmin. Hementerin was also able to degrade cross-linked fibrin although at a lower rate as compared to fibrinogen. In conclusion, hementerin is a plasminogen-independent fibrino(geno)lytic metalloproteinase that degrades fibrinogen faster than fibrin, prevents blood coagulation and destroys fibrin clots in vitro.
Assuntos
Fibrinogênio/metabolismo , Fibrinolíticos/farmacologia , Sanguessugas/enzimologia , Metaloendopeptidases/farmacologia , Extratos de Tecidos/farmacologia , Animais , Fibrinolíticos/isolamento & purificação , Humanos , Cinética , Metaloendopeptidases/isolamento & purificação , Análise de Sequência , Extratos de Tecidos/isolamento & purificaçãoRESUMO
Fibrinolysis triggered by t-PA bound to fibrin is one of the main antithrombotic mechanisms. Defects in the fibrinolytic system-decreased tissue-type plasminogen activator (t-PA) activity and elevated levels of plasminogen activator inhibitor (PAI-1), in patients with SLE have been associated with an increased tendency to thrombosis. In the present study, 43 patients with SLE fulfilling the ACR criteria for the disease, were studied for the presence of autoantibodies to fibrin-bound t-PA, i.e. the physiological active form of this plasminogen activator. A solution of 200 IU/ml of t-PA was incubated with solid-phase fibrin prepared as previously described (Anal Biochem 1986; 153; 201-210). Sera diluted 1:50 were incubated with fibrin-bound t-PA, the plates were then washed, and bound immunoglobulins were detected using a polyvalent peroxidase-labeled goat anti-human Ig. Plates coated with fibrin alone were used as controls. Sera were considered positive when A490/630 obtained with normal human sera in two independent test was greater than the mean plus 2 SD. Eleven of 43 (26%) SLE sera demonstrated antibody reactivity against fibrin-bound t-PA. Within the anti-t-PA positive group there was a higher proportion of SLE patients with severe Raynaud's phenomenon and thrombotic events when compared to the anti-t-PA negative group: 36% vs 6% and 18% vs 6% respectively. These results suggest that autoantibodies to fibrin-bound t-PA could play a role in the pathogenesis of vascular disease in some SLE patients.