Thrombin generation and inhibition in the mediation of ischemic neuronal death

Abstract

Thrombin is a serine protease with dual effects on cell death and survival: It mediates apoptotic cell death at high concentrations in cultured astrocytes and hippocampal neurons and induces neuroprotection at low concentrations in the same cells. Recent evidence indicates that thrombin plays a role in ischemic cell death, as rats subjected to middle cerebral artery occlusion were protected by the intracerebral injection of hirudin, a selective thrombin inhibitor. We are using an in vitro approach to further characterise the mechanisms involved. Organotypic hippocampal slice cultures were subjected to a 30 minute oxygen (10%) and glucose (1 mmol/L) deprivation (OGD). 24 hours after OGD, there was a marked increase in thrombin immunoreactivity on Western blots, showing for the first time that OGD leads to the activation of prothrombin into thrombin. We could also demonstrate that this ischemia induced activation of thrombin plays an important role in ischemic neuronal death, as both recombinant hirudin and protease nexin-1, an endogenous cerebral thrombin inhibitor, significantly prevented neuronal death, when administered after OGD. Using this in vitro approach we are currently characterising the role of thrombin as a mediator of ischemic neuronal death.