Silence K, Collen D, Lijnen H R
Center for Thrombosis and Vascular Research, University of Leuven, Belgium.
Blood. 1993 Aug 15;82(4):1175-83.
The effects of alpha 2-antiplasmin and fibrin on the activation of plasminogen by recombinant staphylokinase (STAR) were studied in an effort to elucidate further the molecular basis of the fibrin-specificity of this fibrinolytic agent. In purified systems consisting of 1.5 mumol/L intact or low-M(r) plasminogen and 3 mumol/L alpha 2-antiplasmin, at 37 degrees C and in the absence of fibrin, STAR did not induce plasminogen activation and plasmin-alpha 2-antiplasmin complex (PAP) formation. Addition of a purified fibrin clot (30% vol at a concentration of 3 mg/mL) to mixtures containing intact plasminogen caused approximately 40% plasminogen activation within 2 hours, whereas in mixtures containing low-M(r) plasminogen, no activation was observed. In contrast, 10 nmol/L streptokinase (SK) induced 74% to 100% plasminogen activation within 2 hours in mixtures containing either intact or low-M(r) plasminogen, in both the absence and the presence of fibrin. In citrated human plasma in the absence of fibrin, 30 nmol/L STAR did not induce measurable plasminogen activation and PAP formation (< 1.5% within 2 hours), whereas addition of a plasma clot (12% vol) resulted in complete clot lysis and conversion of 19% +/- 8% of the plasminogen to PAP within 2 hours. Addition of a second plasma clot produced 23% +/- 2% additional plasminogen activation. Equipotent concentrations for plasma clot lysis of SK (100 nmol/L) induced 54% +/- 11% plasminogen activation in the absence and 49% +/- 16% in the presence of fibrin. Addition of 50 mmol/L 6-aminohexanoic acid (6-AHA) abolished the effect of fibrin on plasminogen activation with STAR, but not on activation with SK. In alpha 2-antiplasmin-depleted human plasma in the absence of fibrin, 30 nmol/L STAR did not induce fibrinogen breakdown (> 90% residual fibrinogen after 6 hours), whereas 30 nmol/L preformed plasmin-STAR complex induced extensive fibrinogen degradation (70% within 20 minutes). Thus, in the absence of fibrin, alpha 2-antiplasmin inhibits the activation of plasminogen by STAR, by preventing generation of active plasmin-STAR complex. Fibrin stimulates plasminogen activation by STAR via mechanisms involving the lysine-binding sites of plasminogen, probably by facilitating the generation of plasmin-STAR complex and by delaying its inhibition at the clot surface.
为了进一步阐明这种纤溶药物纤维蛋白特异性的分子基础，研究了α2 - 抗纤溶酶和纤维蛋白对重组葡萄球菌激酶（STAR）激活纤溶酶原的影响。在由1.5 μmol/L完整或低分子量纤溶酶原和3 μmol/Lα2 - 抗纤溶酶组成的纯化系统中，于37℃且无纤维蛋白存在的情况下，STAR未诱导纤溶酶原激活及纤溶酶 - α2 - 抗纤溶酶复合物（PAP）形成。向含有完整纤溶酶原的混合物中加入纯化的纤维蛋白凝块（浓度为3 mg/mL时体积占30%），在2小时内导致约40%的纤溶酶原激活，而在含有低分子量纤溶酶原的混合物中未观察到激活现象。相比之下，10 nmol/L链激酶（SK）在含有完整或低分子量纤溶酶原的混合物中，无论有无纤维蛋白存在，在2小时内均诱导74%至100%的纤溶酶原激活。在无纤维蛋白的枸橼酸化人血浆中，30 nmol/L STAR未诱导可测量的纤溶酶原激活及PAP形成（2小时内<1.5%），而加入血浆凝块（体积占12%）导致完全凝块溶解，并在2小时内将19%±8%的纤溶酶原转化为PAP。加入第二个血浆凝块可产生额外23%±2%的纤溶酶原激活。SK（100 nmol/L）使血浆凝块溶解的等效浓度在无纤维蛋白时诱导54%±11%的纤溶酶原激活，在有纤维蛋白时诱导49%±16%的纤溶酶原激活，并在有纤维蛋白时诱导49%±16%的纤溶酶原激活。加入50 mmol/L 6 - 氨基己酸（6 - AHA）消除了纤维蛋白对STAR激活纤溶酶原的作用，但未消除对SK激活的作用。在无纤维蛋白的α2 - 抗纤溶酶缺陷型人血浆中，30 nmol/L STAR未诱导纤维蛋白原降解（6小时后>90%残留纤维蛋白原），而30 nmol/L预先形成的纤溶酶 - STAR复合物诱导广泛的纤维蛋白原降解（2分钟内70%）。因此，在无纤维蛋白时，α2 - 抗纤溶酶通过阻止活性纤溶酶 - STAR复合物的产生来抑制STAR对纤溶酶原的激活。纤维蛋白通过涉及纤溶酶原赖氨酸结合位点的机制刺激STAR激活纤溶酶原，可能是通过促进纤溶酶 - STAR复合物的产生以及通过延迟其在凝块表面的抑制作用。
