hirudin and Brain-Edema

This study aimed to investigate the effects of thrombin released in hematoma after intracerebral hemorrhage (ICH) on the cerebral injury of perihematomal tissues and to evaluate the protection effect of hirudin on the cerebral injury after ICH. We used the autologous uncoagulated blood injection method to prepare the ICH rat model, and all rats were randomly divided into a normal group, an ICH group, or a hirudin group. At different time points, rat heads were cut to harvest brain sections. Immunohistochemical staining, histochemical staining, and hematoxylin and eosin staining were conducted for CD34, microglia, and neutrocytes. CD34-positive microvessels were most abundant in brain tissues of the sham-operation group. At 12 h after ICH, CD34 expression reduced and reached the minimum level at 72 h (P<0.01). At 6 h after ICH, microglia expression was visible and reached a peak at 48 h (P<0.01). At 12 h after ICH, neutrocyte infiltration was visible and the number was greatest at 48 h (P<0.01). The early application of hirudin after ICH could significantly reduce microglia and neutrocyte expression and could significantly slow down the CD34 decrease trend (P<0.01). However, hirudin application in the edematization stage after ICH did not significantly increase CD34- positive microvessel abundance (P>0.05). A thrombin-mediated inflammatory reaction is involved in the cerebral injury after ICH, and the early application of hirudin has a protective effect.

Topics: Animals; Antigens, CD34; Brain Edema; Cerebral Hemorrhage; Disease Models, Animal; Hirudins; Leukocytes; Male; Microglia; Microvessels; Rats; Rats, Sprague-Dawley; Thrombin

Edema formation has been linked to thrombin toxicity induced by blood clot at the acute stage of intracerebral hemorrhage. Thrombin induces cell toxicity in neuron, microglia and astrocyte. Aquaporin (AQP) 4 and 9 are proteins expressed on astrocyte in rat brain and involved in the brain water accumulation in brain edema. Recombinant hirudin (r-Hirudin) is a direct inhibitor of thrombin that can block the toxicitic effect of thrombin. In this study, we demonstrated that autologous whole blood infusion in caudate nucleus up-regulates the expression of AQP4 and AQP9 mRNAs and proteins. AQP4 and AQP9 mRNAs expression peaked at about 6 h after blood infusion. The AQP4 protein peaked at about 48 h while AQP9 at about 24 h after blood infusion. Thrombin induced up-regulation of AQP4 and AQP9 were inhibited by r-Hirudin administration and significantly decreased the expression of both AQPs. We further investigated the relationship between edema formation and expression of AQP4 and AQP9. The data presented here may be helpful in optimizing r-Hirudin as an anti-thrombin drug in the treatment of edema at the acute stage of ICH.

Topics: Animals; Aquaporin 4; Aquaporins; Brain Edema; Cerebral Hemorrhage; Gene Expression Regulation; Hirudins; Magnetic Resonance Imaging; Rats; Rats, Wistar; Recombinant Proteins; RNA, Messenger

Thrombin contributes to edema formation after intracerebral hemorrhage. Recent studies suggest that thrombin may also play a role in ischemic brain damage. In the present study, adult male Sprague-Dawley rats were anesthetized with pentobarbital. Middle cerebral artery (MCA) was occluded using the suture method. We found that brain thrombin activity was elevated after permanent MCA occlusion as was prothrombin messenger RNA expression. Intracerebral injection of a thrombin inhibitor, hirudin, reduced neurological deficits following cerebral ischemia. In contrast, intracerebral administration of exogenous thrombin (at a dose that is non-toxic to normal brain), markedly exacerbated brain edema after transient focal cerebral ischemia. These results indicate that extravascular thrombin inhibition may be a new therapeutic target for cerebral ischemia.

Topics: Animals; Antithrombins; Brain; Brain Edema; Brain Ischemia; Disease Susceptibility; Hirudins; Male; Osmolar Concentration; Prothrombin; Rats; Rats, Sprague-Dawley; RNA, Messenger; Severity of Illness Index; Thrombin

In humans, intracerebral hemorrhage (ICH) causes marked perihematomal edema formation and neurological deficits. A rat ICH model, involving infusion of autologous blood into the caudate, has been used extensively to study mechanisms of edema formation, but an examination of behavioral outcome would improve its preclinical utility and provide a more rigorous assessment of the pathological cascade of events over time. The purpose of this study was to use a battery of sensorimotor function tests to examine the neurological effects of ICH in the rat and to examine which components of the hematoma are involved in generating those effects.. The behavioral tests used were forelimb placing, preference for forelimb use for weight shifts during vertical exploration of a cylindrical enclosure, and a corner turn test. Rats were tested from day 1 to day 28 after injection of autologous whole blood; injection of blood plus hirudin (thrombin inhibitor), packed red blood cells, thrombin, or saline; or needle placement only.. The battery of tests indicated that there were marked neurological deficits by day 1 after ICH, with progressive recovery of function over 4 weeks. The forelimb placing score paralleled changes in edema. Injection of thrombin caused and injection of hirudin reduced the ICH-induced neurological deficits. Injection of packed red blood cells, which causes delayed edema formation, induced delayed neurological deficits. These tests allow continuous monitoring of neurological deficits after rat ICH and assessment of therapeutic interventions. The time course of the neurological deficit closely matched the time course of cerebral edema for both ICH and injection of blood components. There was marked recovery of function after ICH, which may be amenable to therapeutic manipulation.

Topics: Animals; Behavior, Animal; Brain; Brain Chemistry; Brain Edema; Cerebral Hemorrhage; Diagnostic Techniques, Neurological; Disease Models, Animal; Disease Progression; Drug Administration Routes; Fibrinolytic Agents; Forelimb; Hirudins; Male; Rats; Rats, Sprague-Dawley; Recovery of Function; Thrombin; Water

Previous work from this laboratory has shown that injection of thrombin into rat basal ganglia causes brain edema. This study investigates the effect on rat brain of thrombin-soaked gelatin sponge (used for intraoperative hemostasis in clinical situations) at a concentration similar to that used in humans. Three models were developed to evaluate this effect. In the first model, a gelatin sponge soaked with vehicle or thrombin (100 U/cm3) was placed on the intact pia of the right frontal lobe in rats without cortical lesions. In the second model, frontal cortex was excised (3 mm3) and the exposed brain was cauterized with electrocoagulation. Gelatin sponge was soaked with vehicle or thrombin (1000, 100, 10, or 1 U/cm3) and placed in the lesion site. In the third model, hirudin, a specific thrombin antagonist, was added to the thrombin-soaked gelatin sponge and placed in a similar cortical lesion to determine if the observed effects were specific to thrombin. The dose-response range for thrombin was determined qualitatively by magnetic resonance (MR) imaging and quantitatively by brain edema formation 24 hours after exposure. We found no edema in the cortically intact rats. The rats given cortical lesions developed significant edema when subjected to 1000, 100, and 10 U/cm3 thrombin as seen on MR imaging and at 100 and 10 U/cm3 thrombin as revealed by wet/dry weight and ion studies of brain tissue. Topical hirudin prevented thrombin-induced edema. It is concluded that thrombin-soaked gelatin sponges cause or enhance significant brain edema in rats at concentrations typically used for human neurosurgery.

Topics: Administration, Topical; Animals; Brain; Brain Edema; Dose-Response Relationship, Drug; Gelatin Sponge, Absorbable; Hirudins; Magnetic Resonance Imaging; Male; Osmolar Concentration; Rats; Rats, Sprague-Dawley; Thrombin

Purified thrombin from an exogenous source is a hemostatic agent commonly used in neurosurgical procedures. The toxicity of thrombin in the brain, however, has not been examined. This study was performed to assess the effect of thrombin on brain parenchyma, using the formation of brain edema as an indicator of injury. Ten microliters of test solution was infused stereotactically into the right basal ganglia of rats. The animals were sacrificed 24 hours later, and the extent of brain edema and ion content were measured. Concentrations of human thrombin as low as 1 U/microliter resulted in a significant increase in brain water content. Rats receiving 10 U/microliters had a mortality rate of 33% compared to no mortality in the groups receiving smaller doses. Thrombin-induced brain edema was inhibited by a specific and potent thrombin inhibitor, hirudin. A medical grade of bovine thrombin commonly used in surgery also caused brain edema when injected at a concentration of 2 U/microliters. Edema formation was prevented by another highly specific thrombin inhibitor, N alpha-(2-Naphthalenesulfonylglycyl)-4-DL-phenylalaninepiperidid e (alpha-NAPAP). Thrombin-induced brain edema was accompanied by increases in brain sodium and chloride contents and a decrease in brain potassium content. Changes in brain ions were inhibited by both hirudin and alpha-NAPAP, corresponding to the inhibition of brain water accumulation. This study shows that thrombin causes brain edema when infused into the brain at concentrations as low as 1 U/microliter, an amount within the range of concentrations used for topical hemostasis in neurosurgery.

Topics: Animals; Brain; Brain Edema; Chlorides; Dipeptides; Hirudins; Injections, Spinal; Magnetic Resonance Imaging; Male; Potassium; Protease Inhibitors; Rats; Rats, Sprague-Dawley; Sodium; Thrombin; Water