Möller Thomas, Weinstein Jonathan R, Hanisch Uwe-Karsten
Department of Neurology, School of Medicine, University of Washington, Seattle, Washington 98195, USA.
Semin Thromb Hemost. 2006 Apr;32 Suppl 1:69-76. doi: 10.1055/s-2006-939556.
In addition to its role in the coagulation cascade, the serine proteinase thrombin (factor IIa) activates cell surface proteinase-activated receptors (PARs) both within and outside the vascular system. PARs are expressed in the central nervous system and mediate thrombin-induced cellular responses in a variety of neural cell types, including microglial cells. Microglial activation by thrombin was reported to induce proliferation, cytokine release, and intracellular calcium signaling. Recently, additional experiments questioned whether these effects are mediated either by thrombin's proteolytic activity or by thrombin itself. Analysis of commercially available plasma-derived thrombin frequently used in the earlier studies showed that cyto/chemokine release-activating properties were not residing with thrombin but were with high molecular weight contaminant(s). In the absence of such contamination, no microglial activation was seen. We compared commercial-grade plasma-derived thrombin to pharmaceutical-grade recombinant thrombin devoid of any measurable contamination. The pharmaceutical-grade thrombin displayed a much more limited profile of microglia-activating properties, triggering only intracellular calcium signals and small changes in surface antigen expression. The signals induced by the pharmaceutical-grade thrombin were completely abolished by proteolytic inhibition, indicating that they are proteolysis-dependent, are most likely PAR mediated, and reflect thrombin's true microglia-activating potential. Prior reports using nonpharmaceutical-grade thrombin need to be reinterpreted critically given these new findings.
除了在凝血级联反应中的作用外，丝氨酸蛋白酶凝血酶（因子IIa）还可激活血管系统内外的细胞表面蛋白酶激活受体（PARs）。PARs在中枢神经系统中表达，并介导凝血酶在多种神经细胞类型（包括小胶质细胞）中诱导的细胞反应。据报道，凝血酶激活小胶质细胞可诱导增殖、细胞因子释放和细胞内钙信号传导。最近，更多实验对这些效应是由凝血酶的蛋白水解活性还是由凝血酶本身介导提出了质疑。对早期研究中常用的市售血浆来源凝血酶的分析表明，细胞因子/趋化因子释放激活特性并非存在于凝血酶中，而是存在于高分子量污染物中。在没有这种污染的情况下，未观察到小胶质细胞激活。我们将商业级血浆来源凝血酶与无任何可检测污染物的药用级重组凝血酶进行了比较。药用级凝血酶显示出的小胶质细胞激活特性更为有限，仅触发细胞内钙信号和表面抗原表达的微小变化。药用级凝血酶诱导的信号被蛋白水解抑制完全消除，表明它们依赖蛋白水解，很可能由PAR介导，并反映了凝血酶真正的小胶质细胞激活潜力。鉴于这些新发现，需要对先前使用非药用级凝血酶的报告进行严格重新解读。
