Subarachnoid hemorrhage causes pulmonary endothelial cell apoptosis and neurogenic pulmonary edema in mice.

OBJECTS: Neurogenic pulmonary edema (NPE) is a well-known complication of subarachnoid hemorrhage (SAH), which potentially causes a poor outcome. The aim of this study was to examine if NPE occurs in the endovascular perforation model of SAH in mice and if apoptosis contributes to NPE development after SAH in mice. METHODS: Sham-operated or SAH mice were treated with an intraperitoneal administration of vehicle or an antiapoptotic drug Z-Val-Ala-Asp-fluoromethylketone (Z-VAD-FMK) 1 h post-SAH. Pulmonary edema measurements and evaluation of apoptosis occurrence were performed on the lung at 24 h post-SAH. RESULTS: SAH caused NPE, which was associated with apoptosis of pulmonary endothelial cells. Z-VAD-FMK significantly prevented apoptosis and NPE. CONCLUSIONS: Pulmonary endothelial cell apoptosis contributes to the pathophysiology of NPE after SAH in mice.

Effects of recombinant osteopontin on blood-brain barrier disruption after subarachnoid hemorrhage in rats.

OBJECTS: We determined effects of recombinant OPN (r-OPN), a pleiotropic extracellular matrix protein, on blood-brain barrier (BBB) disruption and matrix metalloproteinase (MMP)-9 activation after subarachnoid hemorrhage (SAH) in rats. METHODS: The endovascular perforation model of SAH was used. SAH or sham-operated rats were treated with pre-SAH intracerebroventricular administration of two dosages of r-OPN, r-OPN+GRGDSP (an L-arginyl-glycyl-L-aspartate-dependent integrin receptor antagonist), albumin or vehicle. Neurological impairments, brain edema and BBB disruption were evaluated, and Western blot analyses were performed in the brain at 24 h after SAH. RESULTS: r-OPN significantly prevented brain edema and BBB disruption compared with the control rats, associated with the suppression of nuclear factor-κB and mitogen-activated protein kinase pathways, leading to MMP-9 inactivation. These effects were blocked by GRGDSP. CONCLUSIONS: L-arginyl-glycyl-L-aspartate-dependent integrin receptor-mediated multiple signaling pathways may be involved in the protective effects of r-OPN against BBB disruption after SAH.

Apoptotic mechanisms for neuronal cells in early brain injury after subarachnoid hemorrhage.

OBJECTS: The major causes of death and disability in subarachnoid hemorrhage (SAH) may be early brain injury (EBI) and cerebral vasospasm. Although cerebral vasospasm has been studied and treated by a lot of drugs, the outcome is not improved even if vasospasm is reversed. Based on these data, EBI is considered a primary target for future research, and apoptosis may be involved in EBI after experimental SAH. METHODS: We reviewed the published literature about the relationship between SAH induced EBI and apoptosis in PubMed. RESULT: Most available information can be obtained from the endovascular filament perforation animal model. After onset of SAH, intracranial pressure is increased and then cerebral blood flow is reduced. Many factors are involved in the mechanism of apoptotic cell death in EBI after SAH. In the neuronal cells, both intrinsic and extrinsic pathways of apoptosis can occur. Some antiapoptotic drugs were studied and demonstrated a protective effect against EBI after SAH. However, apoptosis in EBI after SAH has been little studied and further studies will provide us more beneficial findings. CONCLUSIONS: The study of apoptosis in EBI after experimental SAH may give us new therapies for SAH.

Immunological response in early brain injury after SAH.

This study summarized the role of inflammation in the early brain injury after subarachnoid hemorrhage. Elevation of cytokines, activation of MMPs and phosphorylation of MAPK contributes to neuronal apoptosis and brain edema. Anti-inflammation may be potential strategy for the prevention and suppression of early brain injury after subarachnoid hemorrhage.

Role of osteopontin in early brain injury after subarachnoid hemorrhage in rats.

BACKGROUND: Subarachnoid hemorrhage (SAH)-induced early brain injury (EBI) contributes to delayed ischemic neurological deficits, one of whose key pathologic manifestation is the blood-brain barrier (BBB) disruption. Although post-SAH BBB breakdown is a self-repairable phenomenon, the molecular pathways are unknown. We determined the role of osteopontin (OPN), a pleiotropic extracellular matrix glycoprotein, in the post-SAH BBB disruption in rats. METHOD: First, we produced the endovascular perforation model of SAH and studied if OPN is induced in the brain after SAH. Secondly, we examined the effects of blockage of endogenous OPN induction on neurological impairments and BBB disruption. Thirdly, we evaluated the effects of exogenous OPN on neurological impairments, brain edema and BBB disruption, and the related protein expression levels. FINDINGS: OPN was significantly induced and peaked at 72 h after SAH, in the recovery phase of EBI. OPN small interfering RNA significantly aggravated neurological impairment and BBB disruption 72 h after SAH. Exogenous OPN significantly prevented neurological impairment, brain edema and BBB disruption associated with the deactivation of nuclear factor-κB activity, the inhibition of matrix metalloproteinase (MMP)-9 induction and tissue inhibitor of MMP-1 reduction, and the consequent preservation of cerebral microvessel basal lamina protein laminin and tight junction protein zona occludens-1. CONCLUSIONS: These findings suggest the protective effects of OPN against BBB disruption after SAH, a finding which should provide a novel therapeutic approach for post-SAH EBI.

Ac-YVAD-CMK Decreases Blood-Brain Barrier Degradation by Inhibiting Caspase-1 Activation of Interleukin-1ß in Intracerebral Hemorrhage Mouse Model.

Among many proinflammatory cytokines, interleukin-1ß (IL-1ß) is considered a key mediator of neuronal injury. However, in order to become activated, it must be processed and cleaved by a caspase-1 enzyme. In this study, we tested the neuroprotective effect of Ac-YVAD-CMK, a known selective caspase-1 inhibitor, in a mouse model of intracerebral hemorrhage (ICH). Sixty-six adult male CD-1 mice were subjected to collagenase-induced ICH. Ac-YVAD-CMK or vehicle was administered into the left lateral ventricle 20 min before ICH modeling. Brain edema and neurological functions were assessed at 24 and 72 h after the surgery. Expression of IL-1ß, phosphorylated JNK, tight junction protein zona occludens 1 (ZO-1), and matrix metalloproteinase-9 (MMP-9) were measured by Western blot along with MMP-9 activity measured by zymography at 24 h after ICH. At 24 h after ICH, Ac-YVAD-CMK treatment significantly reduced brain edema and improved neurological functions. The neuroprotection was associated with downregulation of IL-1ß, JNK, MMP-9, and an inhibition of ZO-1 degradation in brain. We conclude that Ac-YVAD-CMK protects the brain against ICH-induced injury, and the neuroprotective effect may result from anti-inflammation-induced blood-brain barrier protection.

The effects of tetrahydrobiopterin on intracerebral hemorrhage-induced brain injury in mice.

Tetrahydrobiopterin (BH4) is an essential cofactor for nitric oxide synthase (NOS) and is presently used clinically to treat forms of phenylketonuria. BH4 has been reported to restrain superoxide generation of NOS and chemically reduce superoxide. However, there has been no report concerning the effects of BH4 in intracerebral hemorrhage (ICH). In the present study, we investigated the neuroprotective effect of BH4 against ICH-induced brain injury in a mouse model.A total of 26 male CD1 mice (31-39 g) were divided into sham, ICH-vehicle, and ICH-treated with BH4 groups (n = 8 in each group). ICH was induced by collagenase injection into the right basal ganglia. BH4 (20 mg/kg) was administrated intraperitoneally at 1 h after ICH. The effect of BH4 was measured by neurological score and brain water content at 24 h after ICH.Our data demonstrates that ICH caused significant neurological deficit that is associated with brain edema. Treatment with BH4 did not reduce brain edema and neurological deficits at 24 h after ICH in mice. Further study is required to investigate the long-term effect of BH4 in ICH-induced brain injury.

Activation of sphingosine 1-phosphate receptor-1 by FTY720 is neuroprotective after ischemic stroke in rats.

BACKGROUND AND PURPOSE: FTY720 is a known sphingosine 1-phosphate receptor agonist. In the present study, we investigated the neuroprotective effect of postischemic administration of FTY720 in rats with 2 hours transient middle cerebral artery occlusion (MCAO). METHODS: One hundred eleven male rats were randomly assigned to sham-operated and MCAO treated with vehicle, 0.25 mg/kg and 1 mg/kg of FTY720, another selective sphingosine 1-phosphate receptor-1 agonist SEW2871 (5 mg/kg), or 0.25 mg/kg of FTY720 plus a sphingosine 1-phosphate antagonist, VPC23019 (0.5 mg/kg). Drugs were injected intraperitoneally immediately after reperfusion. Neurological score and infarct volume were assessed at 24 and 72 hours after MCAO. Western blotting, immunohistochemistry, and terminal deoxynucleotidyl transferase-mediated uridine 5'-triphosphate-biotin nick end-labeling were conducted at 24 hours after MCAO. RESULTS: FTY720 significantly reduced infarct volume and improved neurological score at 24 and 72 hours after MCAO compared with the vehicle group. SEW2871 showed similar neuroprotective effects to FTY720, whereas VPC 20319 abolished the neuroprotective effects of FTY720. FTY720 significantly retained Akt and extracellular signal-regulated kinase phosphorylation and Bcl-2 expression and decreased cleaved caspase-3 expression and terminal deoxynucleotidyl transferase-mediated uridine 5'-triphosphate-biotin nick end-labeling-positive neurons at 24 hours after MCAO. VPC23019 blocked the antiapoptotic effects of FTY720. CONCLUSIONS: These data suggest that activation of sphingosine 1-phosphate-1 by FTY720 reduces neuronal death after transient MCAO.

Protective effects of recombinant osteopontin on early brain injury after subarachnoid hemorrhage in rats.

OBJECTIVE: Accumulated evidence suggests that the primary cause of poor outcome after subarachnoid hemorrhage is not only cerebral arterial narrowing but also early brain injury. Our objective was to determine the effect of recombinant osteopontin, a pleiotropic extracellular matrix glycoprotein, on early brain injury after subarachnoid hemorrhage in rats. DESIGN: Controlled in vivo laboratory study. SETTING: Animal research laboratory. SUBJECTS: One hundred seventy-seven male adult Sprague-Dawley rats weighing 300 to 370 g. INTERVENTIONS: The endovascular perforation model of subarachnoid hemorrhage was produced. Subarachnoid hemorrhage or sham-operated rats were treated with an equal volume (1 microL) of pre-subarachnoid hemorrhage intracerebroventricular administration of two dosages (0.02 and 0.1 microg) of recombinant osteopontin, albumin, or vehicle. Body weight, neurologic scores, brain edema, and blood-brain barrier disruption were evaluated, and Western blot analyses were performed to determine the effect of recombinant osteopontin on matrix metalloproteinase-9, substrates of matrix metalloproteinase-9 (zona occludens-1, laminin), tissue inhibitor of matrix metalloproteinase-1, inflammation (interleukin-1beta), and nuclear factor-kappaB signaling pathways. MEASUREMENTS AND MAIN RESULTS: Treatment with recombinant osteopontin prevented a significant loss in body weight, neurologic impairment, brain edema, and blood-brain barrier disruption after subarachnoid hemorrhage. These effects were associated with the deactivation of nuclear factor-kappaB activity, inhibition of matrix metalloproteinase-9 induction, the maintenance of tissue inhibitor of matrix metalloproteinase-1, the consequent preservation of the cerebral microvessel basal lamina protein laminin, and the tight junction protein zona occludens-1. CONCLUSIONS: These results demonstrate that recombinant osteopontin treatment is effective for early brain injury after subarachnoid hemorrhage.

Caspase-1 inhibitor prevents neurogenic pulmonary edema after subarachnoid hemorrhage in mice.

BACKGROUND AND PURPOSE: We examined the effects of a caspase-1 inhibitor, N-Ac-Tyr-Val-Ala-Asp-chloromethyl ketone (Ac-YVAD-CMK), on neurogenic pulmonary edema in the endovascular perforation model of subarachnoid hemorrhage (SAH) in mice. METHODS: Ninety-seven mice were assigned to sham, SAH+vehicle, SAH+Ac-YVAD-CMK (6 or 10 mg/kg), and SAH+Z-Val-Ala-Asp-fluoromethylketone (Z-VAD-FMK, 6 mg/kg) groups. Drugs were intraperitoneally injected 1 hour post-SAH. Pulmonary edema measurements, Western blot for interleukin-1beta, interleukin-18, myeloperoxidase, matrix metalloproteinase (MMP)-2, MMP-9, cleaved caspase-3 and zona occludens-1, MMP zymography, terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling staining, and immunostaining were performed on the lung at 24 hours post-SAH. RESULTS: Ten- but not 6-mg/kg of Ac-YVAD-CMK significantly inhibited a post-SAH increase in the activation of interleukin-1beta and caspase-3 and the number of terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling-positive pulmonary endothelial cells, preventing neurogenic pulmonary edema. Another antiapoptotic drug, Z-VAD-FMK, also reduced neurogenic pulmonary edema. SAH did not change interleukin-18, myeloperoxidase, MMP-2, MMP-9, zona occludens-1 levels, or MMP activity. CONCLUSIONS: We report for the first time that Ac-YVAD-CMK prevents lung cell apoptosis and neurogenic pulmonary edema after SAH in mice.

Role of interleukin-1beta in early brain injury after subarachnoid hemorrhage in mice.

BACKGROUND AND PURPOSE: The role of interleukin (IL)-1beta remains unknown in early brain injury (EBI) after subarachnoid hemorrhage (SAH), although IL-1beta has been repeatedly reported to increase in the brain and cerebrospinal fluid. The aim of this study is to examine the effects of IL-1beta inactivation on EBI after SAH in mice. METHODS: The endovascular perforation model of SAH was produced and 112 mice were assigned to sham, SAH+ vehicle, and SAH+ N-Ac-Tyr-Val-Ala-Asp-chloromethyl ketone (Ac-YVAD-CMK, 6 and 10 mg/kg) groups. Ac-YVAD-CMK, a selective inhibitor of IL-1beta converting enzyme, or vehicle was administered intraperitoneally 1 hour post-SAH. EBI was assessed in terms of mortality within 24 hours, neurological scores, brain water content at 24 and 72 hours, Evans blue dye extravasation and Western blot for IL-1beta, c-Jun N-Terminal kinase (JNK), matrix metalloproteinase (MMP)-9, and zonula occludens (ZO)-1 at 24 hours after SAH. RESULTS: High-dose (10 mg/kg) but not low-dose (6 mg/kg) treatment group significantly improved neurological scores, mortality, brain water content, and Evans blue dye extravasation compared with the vehicle group. Although both dosages of Ac-YVAD-CMK attenuated the mature IL-1beta induction, only high-dose treatment group significantly inhibited the phosphorylation of JNK, MMP-9 induction, and ZO-1 degradation. CONCLUSIONS: IL-1beta activation may play an important role in the pathogenesis of EBI after SAH. The neurovascular protection of Ac-YVAD-CMK may be provided by the inhibition of JNK-mediated MMP-9 induction and the consequent preservation of tight junction protein ZO-1.

Effect of gap junction inhibition on intracerebral hemorrhage-induced brain injury in mice.

It has been reported that gap junction contributes to ischemic brain injury and gap junction inhibitors improve neurological outcome in ischemic brain injury models. In the present study, we investigated the effects of gap junction inhibitor, carbenoxolone, on mortality, neurological deficits and brain edema in mice with intracerebral hemorrhage. A total of 80 male CD-1 mice were divided into two parts with two end-points for this study. In part one, animals were divided into four groups: sham, vehicle treatment following intracerebral hemorrhage induction, low-dose carbenoxolone (33 mg/kg) treatment 1 hour after intracerebral hemorrhage induction and high-dose carbenoxolone (100 mg/kg) treatment 1 hour after intracerebral hemorrhage induction groups. Animals were euthanized after 24 hours. In part two, animals were divided into four groups: sham, vehicle treatment 1 hour after intracerebral hemorrhage induction, single high-dose of carbenoxolone treatment at 1 hour after intracerebral hemorrhage induction and three high-doses of carbenoxolone treatment 1, 24 and 48 hours respectively after intracerebral hemorrhage induction. Animals were euthanized after 72 hours. Intracerebral hemorrhage was induced by collagenase injection. Neurological deficits were evaluated using modified Garcia's neurological test, wire hanging and beam balance tests. Brain edema was measured by brain water content. Our results showed that intracerebral hemorrhage produced brain edema and neurological deficits in mice. Carbenoxolone treatment failed to reduce brain edema and neurological deficits. In fact, the high dose of carbenoxolone aggravated neurological deficits and increased mortality 72 hours after the treatment. In conclusion, inhibition of gap junction has no short-term neuroprotective effect on intracerebral hemorrhage-induced brain injury. Further studies are required to assess the long-term effects of gap junction inhibitors in intracerebral hemorrhage models.

The effects of nicotinamide adenine dinucleotide on intracerebral hemorrhage-induced brain injury in mice.

In the present study, we investigated whether the administration of nicotinamide adenine dinucleotide (NAD+) provides brain protection in a mouse model of intracerebral hemorrhage (ICH). Male CD-1 mice were divided into sham, ICH treated with vehicle and ICH treated with NAD+ (10 or 20 mg/kg, intranasal) groups. Intranasal delivery of NAD+ resulted in an increase in NAD+ contents in the brains. ICH was induced by collagenase injection. Neurological function, hemorrhage volume and brain edema were measured 24 hours after injection. ICH caused significant neurological deficit with associated brain edema. NAD+ (10 and 20 mg/kg) failed to reduce brain injury after ICH. These results suggest that NAD+ has no neuroprotective effect at 24 hours after ICH.

[A case of multiple trauma with sinking skull, whose life was saved by consistent team medical treatment].

A 54 year old man was brought to our hospital by ambulance. He had been injured by falling heavy steel. An examination was performed, and he was diag nosed as having sinking skull, acute extradural hematoma, trauma of the righ eye, right eye laceration, injury of the optic canal (right blind), and multipl fractures. Open fractures were observed in the right ring finger and little finger Simple fractures were observed in the zygomatic bone nasal bone and maxillary bone. An emergency operation (external skeletal fixation, taxis of the skull and maxillary bone, extradural hematoma depletion, suture of right eyelid) was performed. His life was saved by consistent team treatment from preoperation t postoperation. He was discharged from our hospital on foot at 45 days after th operation.