Division of Cardiovascular and Diabetes Research, Leeds Institute for Genetics, Health and Therapeutics, Multidisciplinary Cardiovascular Research Centre, University of Leeds, Leeds, United Kingdom.
Blood. 2013 Jul 4;122(1):134-42. doi: 10.1182/blood-2013-04-494641. Epub 2013 May 22.
Diabetes is associated with hypofibrinolysis by mechanisms that are only partially understood. We investigated the effects of in vivo plasminogen glycation on fibrinolysis, plasmin generation, protein proteolytic activity, and plasminogen-fibrin interactions. Plasma was collected from healthy controls and individuals with type 1 diabetes before and after improving glycemia. Plasma-purified plasmin(ogen) functional activity was evaluated by chromogenic, turbidimetric, and plasmin conversion assays, with surface plasmon resonance employed for fibrin-plasminogen interactions. Plasminogen posttranslational modifications were quantified by mass spectrometry and glycation sites located by peptide mapping. Diabetes was associated with impaired plasma fibrin network lysis, which partly normalized upon improving glycaemia. Purified plasmin(ogen) from diabetic subjects had impaired fibrinolytic activity compared with controls (723 ± 16 and 317 ± 4 s, respectively; P < .01), mainly related to decreased fibrin-dependent plasmin generation and reduced protease activity (Kcat/KM 2.57 ± 1.02 × 10⁻³ and 5.67 ± 0.98 × 10⁻³ M⁻¹s⁻¹, respectively; P < .05). Nε-fructosyl-lysine residue on plasminogen was increased in diabetes compared with controls (6.26 ± 3.43 and 1.82 ± 0.95%mol, respectively; P < .01) with preferential glycation of lysines 107 and 557, sites involved in fibrin binding and plasmin(ogen) cleavage, respectively. Glycation of plasminogen in diabetes directly affects fibrinolysis by decreasing plasmin generation and reducing protein-specific activity, changes that are reversible with modest improvement in glycemic control.
糖尿病与纤溶作用降低有关，其机制尚未完全阐明。我们研究了体内纤溶酶原糖基化对纤溶、纤溶酶生成、蛋白水解活性和纤溶酶原-纤维蛋白相互作用的影响。采集健康对照者和 1 型糖尿病患者的血浆，分别在改善血糖前后进行采集。通过比色法、浊度法和纤溶酶转化试验评估血浆纤溶酶原功能活性，并采用表面等离子体共振法研究纤维蛋白-纤溶酶原相互作用。通过质谱法定量纤溶酶原的翻译后修饰，并通过肽图定位糖基化位点。糖尿病患者的血浆纤维蛋白网络溶解受损，改善血糖后部分恢复正常。与对照组相比，糖尿病患者的纯化纤溶酶原（723 ± 16 和 317 ± 4 s；P <.01）的纤维蛋白溶解活性受损，主要与纤维蛋白依赖性纤溶酶生成减少和蛋白酶活性降低有关（Kcat/KM 2.57 ± 1.02 × 10⁻³ 和 5.67 ± 0.98 × 10⁻³ M⁻¹s⁻¹；P <.05）。与对照组相比，糖尿病患者的纤溶酶原上 Nε-果糖基赖氨酸残基增加（6.26 ± 3.43 和 1.82 ± 0.95%mol；P <.01），其中赖氨酸 107 和 557 优先糖基化，这两个位点分别与纤维蛋白结合和纤溶酶原（ogen）裂解有关。糖尿病中纤溶酶原的糖基化直接通过减少纤溶酶生成和降低蛋白特异性活性来影响纤溶，这些变化可随血糖控制的适度改善而逆转。
