batimastat and Cerebral-Hemorrhage

Approximately 7% of survivors from meningococcal meningitis (MM) suffer from neurological sequelae due to brain damage in the course of meningitis. The present study focuses on the role of matrix metalloproteinases (MMPs) in a novel mouse model of MM-induced brain damage.. The model is based on intracisternal infection of BALB/c mice with a serogroup C Neisseria meningitidis strain. Mice were infected with meningococci and randomised for treatment with the MMP inhibitor batimastat (BB-94) or vehicle. Animal survival, brain injury and host-response biomarkers were assessed 48 h after meningococcal challenge.. Mice that received BB-94 presented significantly diminished MMP-9 levels (p < 0.01), intracerebral bleeding (p < 0.01), and blood-brain barrier (BBB) breakdown (p < 0.05) in comparison with untreated animals. In mice suffering from MM, the amount of MMP-9 measured by zymography significantly correlated with both intracerebral haemorrhage (p < 0.01) and BBB disruption (p < 0.05).. MMPs significantly contribute to brain damage associated with experimental MM. Inhibition of MMPs reduces intracranial complications in mice suffering from MM, representing a potential adjuvant strategy in MM post-infection sequelae.

Topics: Animals; Apoptosis; Blood-Brain Barrier; Cerebellum; Cerebral Hemorrhage; Chemokines; Cytokines; Dentate Gyrus; Disease Models, Animal; Female; Kaplan-Meier Estimate; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Matrix Metalloproteinase Inhibitors; Meningitis, Meningococcal; Mice; Phenylalanine; Thiophenes

Intracerebral hemorrhage (ICH) initiates an inflammatory response with secondary growth of hemorrhage and cell death. Matrix metalloproteinase (MMP) gelatinolytic activity is increased in ICH, and synthetic inhibitors to MMPs reduce edema and hemorrhage size. Recently, we found that tissue inhibitor of metalloproteinase-3 (TIMP-3) is elevated after ischemia and colocalizes with TUNEL (terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate-biotin nick end-labeled)-labeled cells. Tissue inhibitor of metalloproteinase-3 promotes neuronal apoptosis in vitro by blocking the shedding of the tumor necrosis factor (TNF) superfamily of death receptors/ligands by stromelysin-1 (MMP-3). However, the effect of TIMP-3 and synthetic MMP inhibitors on cell death in ICH is unclear. Therefore, we used the collagenase-induced intracerebral hemorrhage (CIH) model in Timp-3 knockout and C57Bl/6 wild-type mice to study MMP expression, hemorrhage volume, and cell death. Real-time PCR showed an increase in Mmp-3 mRNA in CIH, but similar Mmp-2 and -9 mRNA expression levels in CIH and saline-injected mice. Protein levels of pro and cleaved MMP-3 were increased in CIH, and zymographic gelatinolytic activity of MMP-9 was elevated after CIH at 72 h, suggesting an exogenous source. Apoptosis was shown by increased caspase-3 levels at 2 and 72 h, and active caspase-8 by 2 and 24 h. The Timp-3 null mouse and wild types had similar hemorrhage sizes and TUNEL-labeled cells. Unexpectedly, the broad-spectrum MMP inhibitor BB-94 increased hemorrhage size and TUNEL-labeled cells. Our results fail to implicate TIMP-3 in apoptosis in CIH, but show that BB-94 increased apoptosis in CIH, possibly by blocking shedding of TNF death receptors and/or their ligands.

Topics: Animals; Apoptosis; Blotting, Western; Cerebral Hemorrhage; Gene Expression; In Situ Nick-End Labeling; Male; Matrix Metalloproteinases; Mice; Mice, Knockout; Phenylalanine; Protease Inhibitors; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Thiophenes; Tissue Inhibitor of Metalloproteinase-3

Thrombolytic therapy with tissue plasminogen activator (tPA) for acute ischemic stroke remains complicated by risks of hemorrhagic transformation. In this study we used a previously established quantitative rat model of tPA-associated hemorrhage to test the hypothesis that matrix metalloproteinases (MMPs) are involved.. Spontaneously hypertensive rats were subjected to embolic focal ischemia by placing homologous blood clots into the middle cerebral artery. Three groups of rats were studied: (1) untreated controls that received saline at 6 hours after ischemia; (2) rats that received tPA alone (10 mg/kg at 6 hours after ischemia); and (3) rats that received tPA plus the broad-spectrum MMP inhibitor BB-94 (50 mg/kg of BB-94 before ischemia and at 3 and 6 hours after ischemia plus tPA at 6 hours). Gelatin zymography was used to quantify MMP levels. A hemoglobin spectrophotometry method was used to quantify cerebral hemorrhage. Ischemic lesions were measured at 24 hours with tetrazolium staining.. At 6, 12, and 24 hours, pro-MMP-9 and cleaved MMP-9 were upregulated in ischemic brain. At 12 hours, tPA-treated rats showed significantly higher levels of pro-MMP-9 and cleaved MMP-9 than untreated controls. By 24 hours, all rats showed evidence of hemorrhagic transformation in the ischemic territory. Rats treated with BB-94 and tPA showed significantly reduced hemorrhage volumes compared with those that received tPA alone. There was no effect on infarct size.. These results indicate that (1) tPA treatment increases levels of MMP-9 after embolic focal cerebral ischemia, (2) MMPs are involved in the mechanism of tPA-associated hemorrhage, and (3) combination therapies with MMP inhibitors may be useful for decreasing the risk and severity of this dreaded complication of thrombolytic therapy.

Topics: Animals; Blood Flow Velocity; Brain; Brain Ischemia; Cerebral Hemorrhage; Cerebrovascular Circulation; Disease Models, Animal; Enzyme Inhibitors; Intracranial Embolism; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Matrix Metalloproteinase Inhibitors; Matrix Metalloproteinases; Phenylalanine; Rats; Rats, Inbred SHR; Sodium, Dietary; Survival Rate; Thiophenes; Thrombolytic Therapy; Tissue Plasminogen Activator

A potentially dangerous side effect associated with tissue plasminogen activator (tPA) use is cerebral hemorrhage. We have focused on developing drugs that could be administered with tPA to reduce the rate of hemorrhage. Since recent studies suggest that various matrix metalloproteinases (MMPs) are important in tumor necrosis factor-alpha production and membrane and vessel remodeling after ischemia, we investigated whether MMP inhibition affected the rate of hemorrhage and infarct production in the absence or presence of tPA treatment.. We occluded the middle cerebral artery of New Zealand White rabbits with radiolabeled blood clots. Five minutes after embolization, we administered either the MMP inhibitor BB-94 (30 mg/kg SC) or its vehicle. Additional groups received BB-94 or vehicle in combination with tPA, administered 60 minutes after embolization (3.3 mg/kg tPA). After 48 hours, the rabbits were killed and brains were removed, immersion fixed for 1 week in 4% paraformaldehyde, and then cut into 5-mm coronal sections that were examined for the presence of hemorrhage, infarcts, and recanalization.. Hemorrhage after embolic stroke was detected in 24% of the control group. tPA induced macroscopically visible hemorrhage in 77% of the tPA-treated group. The rabbits treated with BB-94 had an 18% incidence of hemorrhage (P:>0.05 compared with control). However, when the combination of BB-94 and tPA was administered to rabbits, there was only a 41% incidence of hemorrhage (compared with 77% in the tPA group; P:<0. 05). Both tPA and BB-94/tPA were similarly effective at lysing clots, at 49% and 35% (P:<0.05), respectively, compared with the 5% rate of lysis in the control group. There was a trend for a reduction in the number of infarcts, but it did not reach statistical significance.. Our data suggest that MMP inhibition attenuates mechanisms involved in tPA-induced hemorrhage. This novel form of combination therapy may show promise as a treatment strategy for acute stroke.

Topics: Animals; Cerebral Hemorrhage; Disease Models, Animal; Fibrinolytic Agents; Humans; Intracranial Embolism; Metalloendopeptidases; Phenylalanine; Protease Inhibitors; Rabbits; Stroke; Thiophenes; Thromboembolism; Tissue Plasminogen Activator