Activation of microglial cells by thrombin:: Past, present, and future

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.