Role of METTL20 in regulating ß-oxidation and heat production in mice under fasting or ketogenic conditions.

METTL20 is a seven-ß-strand methyltransferase that is localised to the mitochondria and tri-methylates the electron transfer flavoprotein (ETF) ß subunit (ETFB) at lysines 200 and 203. It has been shown that METTL20 decreases the ability of ETF to extract electrons from medium-chain acyl-coenzyme A (CoA) dehydrogenase (MCAD) and glutaryl-CoA dehydrogenase in vitro. METTL20-mediated methylation of ETFB influences the oxygen consumption rate in permeabilised mitochondria, suggesting that METTL20-mediated ETFB methylation may also play a regulatory role in mitochondrial metabolism. In this study, we generated Mettl20 knockout (KO) mice to uncover the in vivo functions of METTL20. The KO mice were viable, and a loss of ETFB methylation was confirmed. In vitro enzymatic assays revealed that mitochondrial ETF activity was higher in the KO mice than in wild-type mice, suggesting that the KO mice had higher ß-oxidation capacity. Calorimetric analysis showed that the KO mice fed a ketogenic diet had higher oxygen consumption and heat production. A subsequent cold tolerance test conducted after 24 h of fasting indicated that the KO mice had a better ability to maintain their body temperature in cold environments. Thus, METTL20 regulates ETF activity and heat production through lysine methylation when ß-oxidation is highly activated.

[Relationship of Distal Anterior Cerebral Artery Aneurysm Arising from the Supracallosal Portion and Accessory Anterior Cerebral Artery].

Median artery of the corpus callosum(MACC)refers to the median artery of the triplicate anterior cerebral artery(ACA). When MACC distributes to one or to both hemispheres, it is known as the accessory ACA. We performed detailed angiographic analysis of 32 consecutive patients operated upon for distal ACA(DACA)aneurysms, and noted that all DACA aneurysms occurring in the supracallosal portion were accompanied by an accessory ACA as vascular malformation. Such relationship between accessory ACA and DACA aneurysm in the supracallosal portion has not been previously reported.

[Successful surgical treatment of an extrasinusal dural arteriovenous fistula located in the lateral wall of the cavernous sinus: a case report].

We report the successful surgical treatment of a dural arteriovenous fistula(AVF)located in the lateral wall of the cavernous sinus. A 44-year-old woman presented with facial numbness in the left V3 area. Magnetic resonance imaging showed a flow void sign around the sphenoid ridge on a T2-weighted image. Digital subtraction angiography(DSA)demonstrated a dural AVF that was supplied by feeding arteries from the C4 portion of the left internal carotid artery. Without sinus enhancement, the draining vein flowed directly through the superficial sylvian vein into the vein of Labbé. Following treatment of the dural AVF located in the anterior cranial fossa, we ligated the draining vein at the penetrating point in the intradural region using surgical management. Intraoperative findings showed that the shunt point was located in the left lateral wall of the cavernous sinus. The postoperative course was uneventful and DSA showed no evidence of the AVF. The patient was discharged with no neurological deficits. Few studies have reported the use of direct surgery alone for the treatment of dural AVFs located in the cavernous sinus with a perfusion pattern as in this case.

[Intracranial hemorrhage associated with injury of the critical diploic venous system in clipping for unruptured cerebral aneurysm: a case report].

A case of intracranial hemorrhage associated with injury of a critical diploic venous system in clipping for an unruptured cerebral aneurysm was reported. A 67-year-old female presented with a sense of floating. Magnetic resonance angiography (MRA) showed a C1-2 portion aneurysm of the left internal carotid artery 13 mm in size projecting supero-laterally. Three-dimensional CT angiography (3DCTA) volume rendering revealed a developed left fronto-anterior temporal diploic venous system draining the frontal cortical venous return. Because of the large and wide-necked aneurysm, we planned clipping surgery for the purpose of a complete cure. The operation was performed with left fronto-temporal craniotomy at the expense of the diploic venous system. Using techniques such as bipolar coagulation and suction decompression, neck clipping was accomplished via the distal trans-sylvian approach. After the operation, the patient was noticed to be delirious, and post-operative CT demonstrated intracranial hemorrhage in the left frontal lobe with severe brain edema. Motor aphasia was remarkable, but it was gradually relieved, and she left our hospital with no motor weakness. 3 months after the operation, her aphasia was faintly perceptible but she could live independently. We concluded that the injury of a diploic venous system could cause intracranial hemorrhage with intractable brain edema by critically interrupting the frontal venous return.

[Cystic meningioma originating from cavernous sinus: a case report].

A 72-year-old man presented with gait disturbance and right homonymous hemianopsia. Magnetic resonance (MR) imaging showed a solid mass around the left cavernous sinus with rostral development of a multi-lobulated cystic component. The tumor was totally removed including the cystic wall via the left transsylvian approach with orbito-zygomatic craniotomy. Most of the tumor was attached to the lateral wall of the left cavernous sinus. Histological examination showed a meningioma WHO grade I with meningothelial meningioma formed lobules with intranuclear inclusions. His neurological symptoms disappeared immediately after the operation. Postoperative MR imaging revealed total removal of the tumor including the large cystic component.

Modulation of proline-rich akt substrate survival signaling pathways by oxidative stress in mouse brains after transient focal cerebral ischemia.

BACKGROUND AND PURPOSE: A proline-rich Akt substrate (PRAS) contributes to the regulation of apoptosis after a variety of cell death stimuli, as well as in an in vivo transient focal cerebral ischemia (tFCI) model. We reported previously that overexpression of copper/zinc-superoxide dismutase (SOD1) reduces apoptotic cell death after tFCI. Our present study was designed to clarify the relationship between the PRAS signaling pathway and oxidative stress in the regulation of apoptosis after tFCI. METHODS: We used a tFCI model with SOD1 transgenic mice and wild-type littermates to examine the expression of phosphorylated PRAS (pPRAS) by Western blotting and immunohistochemistry and the interaction of pPRAS with phosphorylated Akt (pPRAS/pAkt) or the 14-3-3 protein (pPRAS/14-3-3) by coimmunoprecipitation. Direct oxidation of the carbonyl groups, an indication of oxidative injury to total and individual proteins caused by tFCI, was examined using a 2,4-dinitrophenylhydrazone reaction assay. RESULTS: Expression of pPRAS, pPRAS/pAkt, and pPRAS/14-3-3 decreased 2 hours after tFCI. Oxidized hydroethidine did not colocalize with expression of pPRAS. Individual oxidized carbonyls in pPRAS remarkably increased 2 hours after tFCI but were significantly reduced by SOD1 2 hours after tFCI. Expression of pPRAS, pPRAS/pAkt, and pPRAS/14-3-3 was promoted by SOD1 during the same time course. CONCLUSIONS: These results suggest that overexpression of SOD1 may affect the PRAS pathway after tFCI by reducing the direct oxidative reaction to pPRAS after reperfusion injury.

[A case of intracranial neurinoma arising from the maxillary branch of the trigeminal nerve].

The intracranial portion of the trigeminal branches is rarely involved in neurinomas. The clinical characteristics including symptoms, neuroradiological findings and results of surgical excision of the previously reported 11 neurinomas and the present case arising from the intracranial trigeminal branches are reviewed. A 59-year-old female patient complained of paresthesia and hypesthesia in the infraorbital region with no other neurological abnormalities. Neuroradiological examinations revealed a round cyst-like tumor located at the middle fossa with enlargement of the foramen rotundum and pterygopalatine fossa. No vascular abnormalities were found by angiography. At surgery, the solid tumor with no cystic component arising from the intracranial maxillary nerve was removed totally via a frontotemporal extradural and interdural approach with zygomatic osteotomy. Histopathology revealed a typical neurinoma. The paresthesia disappeared following surgery. Clinical features of neurinomas arising from the trigeminal branches are varied but distinct depending upon the site of origin of the tumors. These tumors can be removed totally in most cases through the extradural and interdural approach without critical morbidity. Special care should be taken not to cause injuries to the surrounding neurovascular structures such as cranial nerves and internal carotid artery. A small amount of residue is acceptable in cases of tumors with tenacious adhesion to these critical structures.

[A case of systemic Lupus erythematosus complicated with multiple subcortical hemorrhage].

We described a case of systemic lupus erythematosus (SLE) complicated with multiple intracerebral hemorrhage. A 52-year-old female with no history of hypertension had been treated with steroid hormone therapy for more than 15 years. Four days before admission, she experienced acute onset of headache and nausea. On the day of admission, the patient suffered from headache and nausea again followed by a deteriorated level of consciousness. The computed tomographic scan revealed two subcortical hematomas in the right frontal and left temporo-occipital area with atypical findings compared to those of the usual intracerebral hemorrhage. Cerebral angiography demonstrated that cortical arteries around the hematomas were markedly stretched and displaced. Multiple segmental stenosis and irregular lumen of the internal carotid artery and bilateral cortical arteries were possibly representative of vasculopathy. Transit time of blood flow was severely delayed with no apparent obstruction of the cerebral arteries, veins and dural sinuses. These findings suggested the existence of severely increased intracranial pressure. Emergent surgical evacuation of the hematomas was successfully performed with a favorable outcome for the patient. Intraoperatively, the brain was congested even after the evacuation of the hematomas. Surgical manipulation was able easily to cause bleeding from the cavity of the hematomas, which was hard to control. It is supposed that vasculopathy including fibrinoid degeneration and vasculitis may have affected the susceptibility of small arteries and veins to rupture in the present case. Subsequent increase of intracranial pressure might cause another hematomas by adding of hemodynamic stress to small vessels apart from those at the site of the first hemorrhage.

[Putaminal hemorrhage in a case of Turner's syndrome with hyperaldosteronemia].

A case of Turner's syndrome complicated with intracerebral hemorrhage in a 21-year-old female is presented. The patient experienced acute onset of unconsciousness and the left-sided hemiparesis due to the right-sided putaminal hemorrhage. Severe hypertension was seen at the hospitalization. Cerebral angiography revealed no vascular lesion other than the displacement of lenticulostriate arteries. Emergent surgical evacuation of the hematoma was successfully performed with her favorable outcome. Postoperatively, several hormonal tests indicated hyperaldosteronemia secondary to the increased levels of ACTH and renin as a result of hyper-response to the environmental stress, leading to critical hypertension. Other examinations including CT scan, MRI, MRA, and laboratory data showed no abnormalities suggestive of renal lesions. It is supposed that hypertension, as a frequent presenting feature, should be strictly controlled in Turner's syndrome from the first time of diagnosis.

Damage to the endoplasmic reticulum and activation of apoptotic machinery by oxidative stress in ischemic neurons.

The endoplasmic reticulum (ER), which plays a role in apoptosis, is susceptible to oxidative stress. Because superoxide is produced in the brain after ischemia/reperfusion, oxidative injury to this organelle may be implicated in ischemic neuronal cell death. Activating transcription factor-4 (ATF-4) and C/EBP-homologous protein (CHOP), both of which are involved in apoptosis, are induced by severe ER stress. Using wild-type and human copper/zinc superoxide dismutase transgenic rats, we observed induction of these molecules in the brain after global cerebral ischemia and compared them with neuronal degeneration. In ischemic, wild-type brains, expression of ATF-4 and CHOP was increased in the hippocampal CA1 neurons that would later undergo apoptosis. Transgenic rats had a mild increase in ATF-4 and CHOP and minimal neuronal degeneration, indicating that superoxide was involved in ER stress-induced cell death. We further confirmed attenuation on induction of these molecules in transgenic mouse brains after focal ischemia. When superoxide was visualized with ethidium, signals for ATF-4 and superoxide overlapped in the same cells. Moreover, lipids in the ER were robustly peroxidized by ischemia but were attenuated in transgenic animals. This indicates that superoxide attacked and damaged the ER, and that oxidative ER damage is implicated in ischemic neuronal cell death.

Modulation of p53 degradation via MDM2-mediated ubiquitylation and the ubiquitin-proteasome system during reperfusion after stroke: role of oxidative stress.

The tumor suppressor gene p53 plays an important role in the regulation of apoptosis through transcriptional activation of cell cycle control. Degradation of p53 hinders its role in apoptosis regulation. Recent studies have shown that MDM2-mediated ubiquitylation and the ubiquitin-proteasome system are critical regulating systems of p53 ubiquitylation. However, the mechanism regulating p53-mediated neuronal apoptosis after cerebral ischemia remains unknown. We examined the MDM2 pathway and the ubiquitin-proteasome system using a transient focal cerebral ischemia (tFCI) model and analyzed the interaction between p53 regulation and superoxide using copper/zinc superoxide dismutase (SOD1) transgenic mice after tFCI. p53 degradation and ubiquitylation were detected after tFCI. The accumulation of ubiquitylated p53 was inhibited and p53 degradation was facilitated by SOD1. Nuclear translocation and MDM2/Akt interaction were detected after tFCI and were inhibited by phosphatidylinositol 3-kinase inhibition and promoted by SOD1. Cytosolic translocation of the p53/MDM2 complex was detected after tFCI and was promoted by SOD1. Moreover, accumulation of multiubiquitin chains and direct oxidative injury to a proteasome were detected and inhibited by SOD1 after tFCI. These results suggest that SOD1 promotes the MDM2 pathway and the ubiquitin-proteasome system after tFCI and that production of reactive oxygen species after tFCI prevents p53 degradation by inhibiting both systems.

Oxidative stress affects the integrin-linked kinase signaling pathway after transient focal cerebral ischemia.

BACKGROUND AND PURPOSE: The integrin-linked kinase (ILK) signaling pathway contributes to regulation of cellular adhesion, migration, and differentiation, and to apoptotic cell death after a variety of cell death stimuli. We have reported that overexpression of copper/zinc superoxide dismutase (SOD1) reduces apoptotic cell death by promoting the phosphatidylinositol 3-kinase (PI3-K)/Akt survival pathway after transient focal cerebral ischemia (tFCI). However, the role of the ILK pathway after tFCI and the role of oxygen free radicals in regulation of apoptosis remain unclear. METHODS: To clarify these issues, we used an in vivo tFCI model with SOD1 transgenic mice and wild-type mice. We administered the PI3-K inhibitor, LY294002, into mouse brains after tFCI and examined the role of PI3-K in the ILK pathway and expression of the ILK/Akt complex by immunohistochemistry, Western blot analysis, and coimmunoprecipitation. RESULTS: A transient increase in ILK was detected early after tFCI and was prevented by treatment with LY294002, but promoted by SOD1. Coimmunoprecipitation revealed that the direct reaction of ILK/Akt transiently increased concurrent with the increase in ILK after tFCI. Moreover, the ILK/Akt complex was prevented by LY294002, but promoted by SOD1. CONCLUSIONS: These results suggest that the ILK pathway mediated by PI3-K is affected by tFCI and by SOD1.

The c-Jun N-terminal protein kinase signaling pathway mediates Bax activation and subsequent neuronal apoptosis through interaction with Bim after transient focal cerebral ischemia.

The c-Jun N-terminal protein kinase (JNK) signaling pathway is implicated in neuronal apoptosis. The mechanism by which activated JNK induces neuronal apoptosis is strongly linked to mitochondrial apoptogenic proteins, although the molecular machinery downstream of JNK has not been precisely elucidated. Our study examined the relevance of proapoptotic Bcl-2 family members in JNK-mediated apoptosis after transient focal cerebral ischemia (tFCI), which, when induced by 60 min of middle cerebral artery (MCA) occlusion, elevated levels of JNK activity and phospho-JNK in the MCA territory. Phospho-JNK was primarily expressed in neurons and colocalized with terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling (TUNEL)-positive cells. Inhibition of JNK activity by anthra[1,9-cd]pyrazol-6(2H)-one (SP600125), a selective JNK inhibitor, protected neurons from ischemia-induced apoptosis detected by TUNEL staining and an apoptotic-related DNA fragmentation assay. SP600125 blocked translocation of the cell death effector Bax from the cytosol to the mitochondria after tFCI. BimL (Bim long) was induced and phosphorylated parallel to JNK activity. Coimmunoprecipitation studies consistently revealed increased interaction of JNK with BimL, as well as BimL with Bax, after tFCI. SP600125 blocked these interactions at a dose that significantly inhibited JNK-induced neuronal apoptosis. These results suggest that the JNK signaling pathway is involved in ischemia-induced neuronal apoptosis by stimulation, at least in part, of Bax translocation to the mitochondria, in which BimL is likely regulated by JNK as a downstream substrate for transmission of apoptotic signals to Bax.

Modulation of the Omi/HtrA2 signaling pathway after transient focal cerebral ischemia in mouse brains that overexpress SOD1.

Omi/HtrA2 is a novel protein that contributes to the regulation of mitochondrial apoptosis after a variety of cell death stimuli in vitro and is thought to negatively control the inhibitor-of-apoptosis protein (IAP) family. However, the Omi/HtrA2 pathway remains unknown in apoptotic neuronal cell death in vivo. To examine the role of the Omi/HtrA2 pathway and its relationship to oxidative stress after reperfusion following cerebral ischemia, we used a transient focal cerebral ischemia (tFCI) model in copper/zinc-superoxide dismutase (SOD1) transgenic mice and wild-type mice. We evaluated the link between the Omi/HtrA2 pathway and the caspase cascade reaction after tFCI by administration of a pan-caspase inhibitor, Z-VAD-FMK. We observed the time-dependent expression of Omi/HtrA2 and its binding to X-chromosome-linked IAP (Omi/XIAP) by immunohistochemistry, Western blotting and coimmunoprecipitation. Translocation of Omi/HtrA2 into the cytosolic space was detected during the early period after tFCI and was not affected by Z-VAD-FMK administration, but it was prevented by SOD1 overexpression. Coimmunoprecipitation revealed that Omi/XIAP transiently increased and that it was prevented by SOD1 overexpression. These results suggest that the Omi/HtrA2 pathway may play an important role in the progress of apoptotic neuronal cell death and that overexpression of SOD1 may attenuate this apoptotic cell death by preventing the Omi/HtrA2 cell signaling pathway.

Oxidative stress is associated with XIAP and Smac/DIABLO signaling pathways in mouse brains after transient focal cerebral ischemia.

BACKGROUND AND PURPOSE: The interaction of X chromosome-linked inhibitor-of-apoptosis protein (XIAP) with second mitochondria-derived activator of caspase (Smac)/direct inhibitor-of-apoptosis protein-binding protein with low pI (DIABLO) contributes to regulation of apoptosis after a variety of cell death stimuli, and in our reported in vivo transient focal cerebral ischemia (tFCI) model. We have also reported that overexpression of copper/zinc superoxide dismutase (SOD1) reduces apoptotic cell death after tFCI. Our present study was designed to clarify the relationship between the XIAP signaling pathway and oxidative stress in the regulation of apoptosis after tFCI. METHODS: We used a tFCI model of SOD1 transgenic mice and wild-type littermates to examine the expression of XIAP and Smac/DIABLO by Western blotting and the interaction of XIAP with Smac/DIABLO (XIAP/Smac) or caspase-9 (XIAP/caspase-9) by coimmunoprecipitation. The direct oxidation of carbonyl groups, an indication of oxidative injury to total and individual proteins caused by tFCI, was examined using a 2,4-dinitrophenylhydrazone reaction assay. RESULTS: Direct oxidative injury to cytosolic and mitochondrial proteins was reduced by SOD1 after tFCI. The individual oxidized carbonyls in XIAP, mitochondrial Smac/DIABLO, and caspase-9 were also reduced by SOD1. Expression of XIAP and XIAP/caspase-9 was promoted, whereas translocation of Smac/DIABLO and XIAP/Smac was reduced, by SOD1 after tFCI. CONCLUSIONS: These results suggest that overexpression of SOD1 may affect the XIAP pathway after tFCI by reducing the direct oxidative reaction to XIAP regulators after reperfusion injury.

Oxidative injury to the endoplasmic reticulum in mouse brains after transient focal ischemia.

Oxidative damage to the endoplasmic reticulum (ER) could be involved in ischemic neuronal cell death because this organelle is susceptible to reactive oxygen species. Using wild-type mice and copper/zinc-superoxide dismutase (SOD1) transgenic mice, we induced focal cerebral ischemia and compared neuronal degeneration and ER stress, that is, phosphorylation of eukaryotic initiation factor 2alpha (eIF2alpha) and RNA-dependent protein kinase-like ER eIF2alpha kinase (PERK). We found that neurons with severe and prolonged phosphorylation of eIF2alpha and PERK underwent later degeneration, and that this was partially prevented by SOD1 overexpression. Signals for superoxide production and phospho-PERK were colocalized, which further indicates a pivotal role for superoxide in ER damage. We investigated the molecular mechanisms of oxidative ER stress and found that detachment of glucose-regulated protein 78 from PERK was the key step. We conclude that ER damage is involved in oxidative neuronal injury in the brain after ischemia/reperfusion.

Neuroprotective role of a proline-rich Akt substrate in apoptotic neuronal cell death after stroke: relationships with nerve growth factor.

The Akt signaling pathway contributes to regulation of apoptosis after a variety of cell death stimuli. A novel proline-rich Akt substrate (PRAS) was recently detected and found to be involved in apoptosis. In our study, Akt activation was modulated by growth factors, and treatment with nerve growth factor (NGF) reduced apoptotic cell death after ischemic injury. However, the role of the PRAS pathway in apoptotic neuronal cell death after ischemia remains unknown. Phosphorylated PRAS (pPRAS) and the binding of pPRAS/phosphorylated Akt (pPRAS/pAkt) to 14-3-3 (pPRAS/14-3-3) were detected, and their expression transiently decreased in mouse brains after transient focal cerebral ischemia (tFCI). Liposome-mediated pPRAS cDNA transfection induced overexpression of pPRAS, promoted pPRAS/14-3-3, and inhibited apoptotic neuronal cell death after tFCI. The expression of pPRAS, pPRAS/pAkt, and pPRAS/14-3-3 increased in NGF-treated mice but decreased with inhibition of phosphatidylinositol-3 kinase and the NGF receptor after tFCI. These results suggest that PRAS phosphorylation and its interaction with pAkt and 14-3-3 might play an important role in neuroprotection mediated by NGF in apoptotic neuronal cell death after tFCI.

Oxidative damage to the endoplasmic reticulum is implicated in ischemic neuronal cell death.

The endoplasmic reticulum (ER), which plays important roles in apoptosis, is susceptible to oxidative stress. Because reactive oxygen species (ROS) are robustly produced in the ischemic brain, ER damage by ROS may be implicated in ischemic neuronal cell death. We induced global brain ischemia on wild-type and copper/zinc superoxide dismutase (SOD1) transgenic rats and compared ER stress and neuronal damage. Phosphorylated forms of eukaryotic initiation factor 2 alpha (eIF2 alpha) and RNA-dependent protein kinase-like ER eIF2 alpha kinase (PERK), both of which play active roles in apoptosis, were increased in hippocampal CA1 neurons after ischemia but to a lesser degree in the transgenic animals. This finding, together with the finding that the transgenic animals showed decreased neuronal degeneration, indicates that oxidative ER damage is involved in ischemic neuronal cell death. To elucidate the mechanisms of ER damage by ROS, we analyzed glucose-regulated protein 78 (GRP78) binding with PERK and oxidative ER protein modification. The proteins were oxidatively modified and stagnated in the ER lumen, and GRP78 was detached from PERK by ischemia, all of which were attenuated by SOD1 overexpression. We propose that ROS attack and modify ER proteins and elicit ER stress response, which results in neuronal cell death.

Interaction between XIAP and Smac/DIABLO in the mouse brain after transient focal cerebral ischemia.

The X chromosome-linked inhibitor-of-apoptosis protein (XIAP) contributes to apoptosis regulation after a variety of cell death stimuli. XIAP inhibits the caspase reaction via binding to caspases, and is inhibited via binding to the second mitochondria-derived activator of caspase (Smac)/DIABLO to tightly control apoptotic cell death. However, the interaction among XIAP, Smac/DIABLO, and caspases after in vivo cerebral ischemia is not well known. To clarify this issue, the authors examined time-dependent expression and interaction among XIAP, Smac/DIABLO, and activated caspase-9 by immunohistochemistry, Western blot analysis, and immunoprecipitation using an in vivo transient focal cerebral ischemia model. To examine the relationship of the XIAP pathway to the caspase cascade, a pan-caspase inhibitor was administered. XIAP increased concurrently with the release of Smac/DIABLO and the appearance of activated caspase-9 during the early period after reperfusion injury. The bindings of XIAP to Smac/DIABLO and to caspase-9 and the binding of Smac/DIABLO to caspase-9 reached a peak simultaneously after transient focal cerebral ischemia. Neither XIAP nor Smac/DIABLO expression was affected by caspase inhibition. These results suggest that the XIAP pathway was activated upstream of the caspase cascade and that interaction among XIAP, Smac/DIABLO, and caspase-9 plays an important role in the regulation of apoptotic neuronal cell death after transient focal cerebral ischemia.

Induction of GRP78 by ischemic preconditioning reduces endoplasmic reticulum stress and prevents delayed neuronal cell death.

Although the endoplasmic reticulum (ER) is implicated in neuronal degeneration in some situations, its role in delayed neuronal cell death (DND) after ischemia remains uncertain. The authors speculated that ER stress is involved in DND, that it is reduced by ischemic preconditioning, and that ER stress reduction by preconditioning is due to ER molecular chaperone induction. The phosphorylation status of eukaryotic initiation factor 2alpha (eIF2alpha) and RNA-dependent protein kinase-like ER eIF2alpha kinase (PERK) was investigated in the rat hippocampus after ischemia with and without preconditioning. PERK is phosphorylated by ER stress, which phosphorylates eIF2alpha. To investigate the role of ER molecular chaperones in preconditioning, the authors examined GRP78 and GRP94 expression, both of which are ER chaperones that inhibit PERK phosphorylation, and compared their induction and ischemic tolerance time windows. Phosphorylation of eIF2alpha and PERK was confirmed after severe ischemia but was inhibited by preconditioning. After preconditioning, GRP78 was increased in the brain with a peak at 2 days, which corresponded with the ischemic tolerance time window. Immunoprecipitation and double staining demonstrated involvement of GRP78 in prevention of PERK phosphorylation. These results suggest that GRP78 induced by preconditioning may reduce ER stress and eventual DND after ischemia.