n-acetylheparin and Brain-Edema

Background and Purpose- Early erythrolysis in the hematoma contributes to brain injury after intracerebral hemorrhage (ICH). This study investigated the effects of N-acetylheparin, a complement inhibitor, and aurin tricarboxylic acid, a membrane attack complex inhibitor, on early erythrolysis, brain iron deposition, and brain injury in aged rats. Methods- There were 3 parts in the study. First, aged (18 months old) male Fischer 344 rats had an ICH. The time course of erythrolysis in the hematoma was determined by T2* weighted magnetic resonance imaging, and the expression of CD163 was examined. Second, aged rats had an ICH with N-acetylheparin or vehicle. Rats were euthanized at days 1, 3, and 28 after magnetic resonance imaging (T2-, T2*-weighted, and T2* array) and behavioral tests. Brains were used for immunohistochemistry. Third, aged rats had an ICH with avaurin tricarboxylic acid or vehicle. The rats had magnetic resonance imaging and behavioral tests and were euthanized at day 3. Brains were used for immunohistochemistry. Results- Early erythrolysis occurred within the clot in aged F344 rats. There were increased numbers of CD163-positive cells after ICH. Almost all perihematomal CD163-positive cells were microglia/macrophages, while positive neurons were found more distant from the hematoma. Coinjection of N-acetylheparin attenuated erythrolysis, iron accumulation, CD163 expression, microglia activation, brain swelling, and neuronal death in the acute phase, as well as reducing brain atrophy and neurological deficits in the chronic phase. Coinjection of aurin tricarboxylic acid also reduced erythrolysis and ICH-induced brain injury. Conclusions- Inhibiting complement activation resulted in less erythrolysis and brain injury after ICH.

Topics: Aging; Animals; Antigens, CD; Antigens, Differentiation, Myelomonocytic; Aurintricarboxylic Acid; Brain Edema; Brain Injuries; Complement Inactivating Agents; Erythrocytes; Hemolysis; Heparin; Intracranial Hemorrhages; Ischemic Attack, Transient; Macrophage Activation; Male; Microglia; Rats; Rats, Inbred F344; Receptors, Cell Surface

The present study examined whether thrombin activates the complement cascade in the brain and whether N-acetylheparin, an inhibitor of complement activation, attenuates brain injury induced by thrombin. There were three sets of studies. In the first set, rats had an intracerebral infusion of either five-unit thrombin or a needle insertion. Brains were sampled at 24 hours for Western blot analysis and immuno-histochemistry. In the second set, rats received either five-unit thrombin+saline, five-unit thrombin+25 microg N-acetylheparin or five-unit thrombin+100 microg N-acetylheparin infusion. Brains were sampled 24 hours later for water content measurement. In the third set, rats received either five-unit thrombin+saline or five-unit thrombin+ 100 microg N-acetylheparin. Behavioral tests sensitive to unilateral striatal damage were carried out for two weeks. Western blotting demonstrated that complement C9 and clusterin levels increase 24 hours after thrombin infusion (P < 0.01). Both C9 and clusterin positive cells were found around the injection site. High-dose (100-microg) but not low-dose (25-microg) N-acetylheparin attenuated thrombin-induced brain edema (81.5 +/- 0.4% vs. 83.7 +/- 0.3% in the vehicle, P < 0.05). Behavior was also significantly improved by N-acetylheparin (P < 0.05). In conclusion, thrombin-induced edema formation and neurological deficits were both reduced by N-acetylheparin. This suggests that inhibition may be a novel treatment for the thrombin-induced brain injury that occurs in intracerebral hemorrhage.

Topics: Animals; Behavior, Animal; Brain Edema; Complement Activation; Complement Inactivating Agents; Dose-Response Relationship, Drug; Drug Combinations; Heparin; Male; Nervous System Diseases; Rats; Rats, Sprague-Dawley; Thrombin; Treatment Outcome

Brain edema formation following intracerebral hemorrhage (ICH) appears to be partly related to erythrocyte lysis and hemoglobin release. Erythrocyte lysis may be mediated by the complement cascade, which then triggers parenchymal injury. In this study the authors examine whether the complement cascade is activated after ICH and whether inhibition of complement attenuates brain edema around the hematoma.. This study was divided into three parts. In the first part, 100 microl of autologous blood was infused into the rats' right basal ganglia, and the animals were killed at 24 and 72 hours after intracerebral infusion. Their brains were tested for complement factors C9, C3d, and clusterin (a naturally occurring complement inhibitor) by using immunohistochemical analysis. In the second part of the study, the rats were killed at 24 or 72 hours after injection of 100 microl of blood. The C9 and clusterin proteins were quantitated using Western blot analysis. In the third part, the rats received either 100 microl of blood or 100 microl of blood plus 10 microg of N-acetylheparin (a complement activation inhibitor). Then they were killed 24 or 72 hours later for measurement of brain water and ion contents. It was demonstrated on Western blot analysis that there had been a sixfold increase in C9 around the hematoma 24 hours after the infusion of 100 microl of autologous blood. Marked perihematomal C9 immunoreactivity was detected at 72 hours. Clusterin also increased after ICH and was expressed in neurons 72 hours later. The addition of N-acetylheparin significantly reduced brain edema formation in the ipsilateral basal ganglia at 24 hours (78.5 +/- 0.5% compared with 81.6 +/- 0.8% in control animals, p < 0.001) and at 72 hours (80.9 +/- 2.2% compared with 83.6 +/- 0.9% in control animals, p < 0.05) after ICH.. It was found that ICH causes complement activation in the brain. Activation of complement and the formation of membrane attack complex contributes to brain edema formation after ICH. Blocking the complement cascade could be an important step in the therapy for ICH.

Topics: Animals; Brain Diseases; Brain Edema; Cerebral Hemorrhage; Clusterin; Complement Activation; Complement C3d; Complement C9; Glycoproteins; Hematoma; Heparin; Male; Molecular Chaperones; Rats; Rats, Sprague-Dawley; Up-Regulation