Miconazole protects blood vessels from MMP9-dependent rupture and hemorrhage.

Hemorrhagic stroke accounts for 10-15% of all strokes and is strongly associated with mortality and morbidity worldwide, but its prevention and therapeutic interventions remain a major challenge. Here, we report the identification of miconazole as a hemorrhagic suppressor by a small-molecule screen in zebrafish. We found that a hypomorphic mutant fn40a, one of several known ß-pix mutant alleles in zebrafish, had the major symptoms of brain hemorrhage, vessel rupture and inflammation as those in hemorrhagic stroke patients. A small-molecule screen with mutant embryos identified the anti-fungal drug miconazole as a potent hemorrhagic suppressor. Miconazole inhibited both brain hemorrhages in zebrafish and mesenteric hemorrhages in rats by decreasing matrix metalloproteinase 9 (MMP9)-dependent vessel rupture. Mechanistically, miconazole downregulated the levels of pErk and Mmp9 to protect vascular integrity in fn40a mutants. Therefore, our findings demonstrate that miconazole protects blood vessels from hemorrhages by downregulating the pERK-MMP9 axis from zebrafish to mammals and shed light on the potential of phenotype-based screens in zebrafish for the discovery of new drug candidates and chemical probes for hemorrhagic stroke.

Inhibition of matrix metalloproteinases attenuates brain damage in experimental meningococcal meningitis.

BACKGROUND: 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. METHODS: 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. RESULTS: 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). CONCLUSIONS: 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.

Induction of hyperhomocysteinemia models vascular dementia by induction of cerebral microhemorrhages and neuroinflammation.

Vascular dementia (VaD) is the second leading cause of dementia behind Alzheimer's disease (AD) and is a frequent comorbidity with AD, estimated to occur in as many as 40% of AD patients. The causes of VaD are varied and include chronic cerebral hypoperfusion, microhemorrhages, hemorrhagic infarcts, or ischemic infarcts. We have developed a model of VaD by inducing hyperhomocysteinemia (HHcy) in wild-type mice. By placing wild-type mice on a diet deficient in folate, B6, and B12 and supplemented with excess methionine, we induced a moderate HHcy (plasma level homocysteine 82.93 ± 3.561 µmol). After 11 weeks on the diet, the hyperhomocysteinemic mice showed a spatial memory deficit as assessed by the 2-day radial-arm water maze. Also, magnetic resonance imaging and subsequent histology revealed significant microhemorrhage occurrence. We found neuroinflammation induced in the hyperhomocysteinemic mice as determined by elevated interleukin (IL)-1ß, tumor necrosis factor (TNF)α, and IL-6 in brain tissue. Finally, we found increased expression and increased activity of the matrix metalloproteinase 2 (MMP2) and MMP9 systems that are heavily implicated in the pathogenesis of cerebral hemorrhage. Overall, we have developed a dietary model of VaD that will be valuable for studying the pathophysiology of VaD and also for studying the comorbidity of VaD with other dementias and other neurodegenerative disorders.

Extension of the thrombolytic time window with minocycline in experimental stroke.

BACKGROUND AND PURPOSE: Thrombolysis with tPA is the only FDA-approved therapy for acute ischemic stroke. But its widespread application remains limited by narrow treatment time windows and the related risks of cerebral hemorrhage. In this study, we ask whether minocycline can prevent tPA-associated cerebral hemorrhage and extend the reperfusion window in an experimental stroke model in rats. METHODS: Spontaneously hypertensive rats were subjected to embolic focal ischemia using homologous clots and treated with: saline at 1 hour; early tPA at 1 hour, delayed tPA at 6 hours; minocycline at 4 hours; combined minocycline at 4 hours plus tPA at 6 hours. Infarct volumes and hemorrhagic transformation were quantified at 24 hours. Gelatin zymography was used to measure blood levels of circulating matrix metalloproteinase-9 (MMP-9). RESULTS: Early 1-hour thrombolysis restored perfusion and reduced infarction. Late 6-hour tPA did not decrease infarction but instead worsened hemorrhagic conversion. Combining minocycline with delayed 6-hour tPA decreased plasma MMP-9 levels, reduced infarction, and ameliorated brain hemorrhage. Blood levels of MMP-9 were also significantly correlated with volumes of infarction and hemorrhage. CONCLUSIONS: Combination therapy with minocycline may extend tPA treatment time windows in ischemic stroke.

Hyperbaric oxygen preconditioning activates ribosomal protein S6 kinases and reduces brain swelling after intracerebral hemorrhage.

BACKGROUND: New protein synthesis is key to ischemic tolerance induced by preconditioning and ribosomal protein S6 kinases (p70 S6 K) are important enzymes in protein synthesis. Hyperbaric oxygen preconditioning (HBOP) reduces ischemic brain damage. This study investigated if HBOP can activate p70 S6 K and increase new protein synthesis and if HBOP induces brain tolerance against brain swelling after intracerebral hemorrhage (ICH). METHODS: There were two parts of the studies. 1) Rats received five consecutive sessions of HBOP. Twenty-four hours after HBOP, the rats had an ICH and were sacrificed one or three days later for brain edema measurement. 2) Rats received five sessions of HBOP or control pretreatment and were sacrificed for Western blot analysis and immunohistochemistry of activated p70 S6 K and heme oxygenase-1 (HO-1). FINDINGS: Five sessions of HBOP significantly reduced brain edema in the ipsilateral basal ganglia after ICH. Western blot analysis showed that HBOP activated p70 S6 K and increased HO-1 levels in the basal ganglia. Strong activated p70 S6 K immunoreactivity was also found in the basal ganglia. CONCLUSIONS: Our results suggest activation of p70 S6 K may have a role in heat shock protein synthesis after HBOP and may contribute to HBOP-induced brain protection.

[Protective effect of traditional Chinese medicine nao-yi-an granule in experimental rats with hemorrhagic stroke].

In order to establish a model of intracerebral hemorrhage(ICH), 0.4 U bacterial collagenase was injected stereotaxically into globus pallidus in rats. Neurological deficit score was investigated, and inducible nitric oxide syntahse(iNOS) mRNA was detected in 72 rats. The results showed that: in the group treated by nao-yi-an granule(NYA), the recovery time was shorter than that in other groups, and low concentration nitrite and weak signal of iNOS mRNA were found. It suggests that inhibition of iNOS induction is one of the pharmacological effects of NYA.

[Effects of Nao-Yi-An granule on activity of cytochrome c oxidase in hippocampus of experimental intracerebral hemorrhagic rats].

OBJECTIVE: To explore the neuronal protective effects of a traditional Chinese medicine complex Nao-Yi-An granule(NYA) on experimental intracerebral hemorrhage(ICH) in rats. METHODS: Eighty collagenase-induced ICH rats were used. At 12 h, 24 h, 2 d, 4 d, 7 d, cytochrome c oxidase(CO) activity was measured with histochemistry combined with gray scale scanning. Pyramidal cell(PC) counting was also employed following Nissl's staining. RESULTS: The CO activities in NYA-treated group were less than those in model control group, and the neuronal loss was less in NYA-treated group than that in model control group. CONCLUSIONS: The neuronal protective mechanism of NYA following ICH may be related with its maintaining activity of cytochrome c oxidase, which contributes to improving cellular aerobic metabolism and increasing energy supply for cells.

[Protection of naoyi-an granule in ischemic brain injury secondary to intracerebral hemorrhage in rats].

AIM: To explore the protective effect and its mechanism of the traditional Chinese medicine complex, naoyi-an granule(NYAG), on intracerebral hemorrhage(ICH) in rats. METHODS: Collagenase-induced ICH rats were used. Inducible nitric oxide synthase(iNOS) activity of ischemic cerebral cortex surrounding the intracerebral hematoma was assayed by monitoring the conversion of L-3H-arginine to L-3H-citrulline. Pathologic changes in the same area were observed with transmission electron microscope on the 2nd day, the 4th day and the 7th day after ICH. RESULTS: The iNOS activities of NYA-treated group decreased remarkably(P < 0.01) compared with those of the saline-treated group. Ultrastructural observation also indicated that NYAG ameliorated the brain edema, cell degeneration and necrosis. CONCLUSIONS: The inhibitory effect of NYAG on iNOS activity may be related to its protection against ischemic brain injury secondary to ICH.

Induction of NADPH-diaphorase activity in the forebrain in a model of intracerebral hemorrhage and its inhibition by the traditional Chinese medicine complex Nao Yi An.

Induction of NADPH-diaphorase (NDP) activity in the rat cerebral cortex was studied after autologous blood injection into the internal capsule as experimental model of intracerebral hemorrhage. The potential inhibitory effect on NDP induction by Nao Yi An (NYA), a complex derived from materials of animal and plant origin used in the treatment of intracerebral hemorrhage in traditional Chinese medicine, was also investigated. In animals without therapeutic treatment 2 and 4 days after injection of autologous blood, NDP activity was highly induced in pyramidal neurons in the neocortex, piriform, and entorhinal cortices, in astrocytes and in phagocytes in the hematoma and the area surrounding it, as well as in the subcortical white matter, and in endothelial cells in both the cortex and subcortical white matter bilaterally. Oral administration of NYA failed to inhibit NDP induction in endothelial cells but demonstrated a strong inhibitory effect on NDP activity induced in pyramidal neurons and astrocytes. NDP induction in phagocytes was also inhibited by the administration of NYA. Altogether the present results suggest that intracerebral hemorrhage in the internal capsule may induce nitric oxide synthase activity in different cell populations in the cortex and that administration of NYA can selectively inhibit such induction and, thus, potentially play a neuroprotective role.

[Changes in the cerebellar cortex of rabbits subjected to an electrical current on the posterior hypothalamus]. / Izmeneniia kory polusharii mozzhechka krolika pri deistvii élektricheskogo toka na zadnee pole gipotalamusa.

After 3, 15 and 30 sessions of electrical stimulation of the hypothalamus by means of stereotaxically implanted bipolar electrodes into the area hypothalamica posterior, in the cortex of the cerebellar hemispheres, predominantly in the ganglionic layer, certain reactive, as well as poorly and intensively manifested pathological changes develop in neural and glial cells. Mosaicism of Purkinje's cells lesions within the layer, the state of their cytoplasmic matrix, granular endoplasmic reticulum and mitochondria, essential shifts in activity of oxidation-reduction enzymes at a simultaneous disorder in blood circulation and decreased content of catecholamines in the neural structures of the cerebellar vessels make it possible to estimate the process as centrogenic and neurodistrophic.