The Inhibition Of Thrombin-induced Brain Injury In The Rats

Hemorrhagic stroke is the major cause of death and disability in the world. However, both experimental and clinical studies have shown that moderte intracerebral hemorrhage (ICH) itself does not induce significant increase of intracranial pressure, so brain injury by edema surrounding hematoma and the secondary neuron apoptosis are the main causes affecting the prognosis of patients; therefore, they became the most interested recently. Thrombin, which activates the formation of fibrin, has been suggested as a key contributor to the formation of brain edema and apoptosis of neuron following ICH. The underlying mechanisms remain unclear.ObjectiveThe aim of this study was to investigate the action of thrombin in the formation of brain edema and neuron apoptosis following ICH. And the inhibition of thrombin-induced brain injury was further determined by acministration of thormbotic inhibitor, huridin, and a calcium antagonist, nimodipine in SD rat.MethodsDifferent doses of thrombin were injected into nucleus caudatus of SD rats using a stereotaxic technique: 1. To examine brain edema using a "dry-wet-weighing technique" andhistological method (HE staining).2. To examine the changes of cytoskeleton (MAP-2, NF200, GFAP) in thrombin-indueced brain injury (especially the areas of dendrites, axons and cytoprocess) and to examine whether these injuries could be reversed by immunohistochemistry;3. To examine neuron cell death following intracerebral hemorrhages and the effects of a thrombin inhibitor, huridin, and a selective calcium channel blocker, nimodipine , on the thrombin-induced edema and cell death by TUNEL assay and electronic microscope examination.4. SD rats were divided randomly into 6 groups (5 rats/group): saline only; low dose of thrombin (lu); high dose of thrombin (5u); low dose of thrombin (lu) with low dose of huridin (lu); high dose of thrombin (5u) with high dose of hurdin (5u); nimodipine treated. Five time points, 4hr, 24hr, 48hr, 3d, 7d, were used in each group. Nimodipine (50ug/100g of body weight) was injected intraperitoneally from day 0 to day 6 every day.Results1. This study showed that Thrombin-induced brain edema was dose-dependent. High dose of thrombin resulted severe cerebral edema as early as 4hrs after injection and the maximum edema occured at 24-48 hr. The edema would gradually decrease and could be reversedwithin 3-7 day. However, Low dosage did not induce obvious brain edema.2. Thrombin-induced edema could be inhibited by a specific inhibitor of thrombin, huridin, and a calcium-ion antagonist, nimodipine. Therefore, these agents might inhibit thrombin-induced edema following ICH.3. High dose of thrombin caused cytoskeleton change at an early stage (4hr.), and reversible or irreversible injuries at 24-48 hr. after injection. And neuron necrosis, irreversible injury occurred within 3-7 days after injection. In contrast, low dose of thrombin did not have these effects. Huridin could reverse but nimodipine, the calcium-ion antagonist, might attenuate the effects of thrombin.4. At high dose of thrombin, neural cell apoptosis started at 4 hours after injection and peaked at 24-48 hours. However, low dose of thrombin did not have this effect. Huridin could reverse but nimodipine decrease the effects of thrombin.ConclusionsIn summary, the results of current study demonstrate that injection of thrombin cause brain edema, necrosis and cell death in a dose-dependent manner in rats. The effects occur as early as 4 hours after injection, and reach its peak at 24-48 hours. Early treatments with thrombin inhibtor (huridin) or calcium antagonist (nimodipine) could greatly reduce thebrain damage. Therefore, this study provide us possible therapeutic agents in the treatment of intracerebral hemorrhages.