背景:人血清白蛋白(HSA)是最丰富的血浆蛋白,对体内糖基化敏感。糖尿病(DM)患者的慢性高血糖状况会诱导非酶Maillard反应,使血浆蛋白变性并形成晚期糖基化终产物(AGEs)。HSA-AGE是DM患者中普遍存在的错误折叠蛋白,并且与因子XII活化和下游促炎激肽释放酶-激肽系统活性相关,而没有任何相关的内在途径的促凝血活性。
目的:本研究旨在确定HSA-AGE与糖尿病病理生理学的相关性。
方法:从DM患者和正常血糖志愿者获得的血浆中探测FXII的激活,前激肽释放酶(PK),并通过免疫印迹裂解高分子量激肽原。通过显色测定法测定组成型血浆激肽释放酶活性。FXII的活化和动力学调制,PK,FXI,FIX,和FX通过体外产生的HSA-AGE进行了显色试验,血浆凝血试验,和使用全血的体外流动模型。
结果:从DM患者获得的血浆含有增加的血浆AGEs,激活FXIIa,并得到裂解的裂解的高分子量激肽原。鉴定了升高的组成型血浆激肽释放酶酶活性,与糖化血红蛋白水平呈正相关,这是这种现象的第一个证据。HSA-AGE,在体外产生,触发FXIIa依赖性PK激活,但通过抑制血浆中FXIa和FIXa依赖性FX激活来限制内源性凝血途径激活。
结论:这些数据表明HSA-AGEs通过FXII和激肽释放酶-激肽系统激活在DM的病理生理学中的促炎作用。FXII活化的促凝血作用通过由HSA-AGEs抑制FXIa和FIXa依赖性FX活化而丧失。
Human serum albumin (HSA) is the most abundant plasma protein and is sensitive to glycation in vivo. The chronic hyperglycemic conditions in patients with diabetes mellitus (DM) induce a nonenzymatic Maillard reaction that denatures plasma proteins and forms advanced glycation end products (AGEs). HSA-AGE is a prevalent misfolded protein in patients with DM and is associated with factor XII activation and downstream proinflammatory kallikrein-kinin system activity without any associated procoagulant activity of the intrinsic pathway.
This study aimed to determine the relevance of HSA-AGE toward diabetic pathophysiology.
The plasma obtained from patients with DM and euglycemic volunteers was probed for activation of FXII, prekallikrein (PK), and cleaved high-molecular-weight kininogen by immunoblotting. Constitutive plasma kallikrein activity was determined via chromogenic assay. Activation and kinetic modulation of FXII, PK, FXI, FIX, and FX via in vitro-generated HSA-AGE were explored using chromogenic assays, plasma-clotting assays, and an in vitro flow model using whole blood.
Plasma obtained from patients with DM contained increased plasma AGEs, activated FXIIa, and resultant cleaved cleaved high-molecular-weight kininogen. Elevated constitutive plasma kallikrein enzymatic activity was identified, which positively correlated with glycated hemoglobin levels, representing the first evidence of this phenomenon. HSA-AGE, generated in vitro, triggered FXIIa-dependent PK activation but limited the intrinsic coagulation pathway activation by inhibiting FXIa and FIXa-dependent FX activation in plasma.
These data indicate a proinflammatory role of HSA-AGEs in the pathophysiology of DM via FXII and kallikrein-kinin system activation. A procoagulant effect of FXII activation was lost through the inhibition of FXIa and FIXa-dependent FX activation by HSA-AGEs.