Pathology Of Cerebral Ischemia Research Articles

Phelligridimer A enhances the expression of mitofusin 2 and protects against cerebral ischemia/reperfusion injury

Mitochondrial dysfunction and endoplasmic reticulum (ER) stress play pivotal roles in the pathology of cerebral ischemia. In this study, we investigated whether phelligridimer A (PA), an active compound isolated from the medicinal and edible fungus Phellinus igniarius, ameliorates ischemic cerebral injury by restoring mitochondrial function and restricting ER stress. An in vitro cellular model of ischemic stroke-induced neuronal damage was established by exposing HT-22 neuronal cells to oxygen-glucose deprivation/reoxygenation (OGD/R). An in vivo animal model was established in rats subjected to middle cerebral artery occlusion/reperfusion (MCAO/R). The results showed that PA (1–10 μM) dose-dependently increased HT-22 cell viability, reduced OGD/R-induced lactate dehydrogenase release, and reversed OGD/R-induced apoptosis. PA reduced OGD/R-induced accumulation of reactive oxygen species, restored mitochondrial membrane potential, and increased ATP levels. Additionally, PA reduced the expression of the 78-kDa glucose-regulated protein (GRP78) and the phosphorylation of inositol-requiring enzyme-1α (p-IRE1α) and eukaryotic translation-initiation factor 2α (p-eIF2α). PA also inhibited the activation of the mitogen-activated protein kinase (MAPK) pathway in the OGD/R model. Moreover, treatment with PA restored the expression of mitofusin 2 (Mfn-2), a protein linking mitochondria and ER. The silencing of Mfn-2 abolished the protective effects of PA. The results from the animal study showed that PA (3–10 mg/kg) significantly reduced the volume of cerebral infarction and neurological deficits, which were accompanied by an increased level of Mfn-2, and decreased activation of the ER stress in the penumbra of the ipsilateral side after MCAO/R in rats. Taken together, these results indicate that PA counteracts cerebral ischemia-induced injury by restoring mitochondrial function and reducing ER stress. Therefore, PA might be a novel protective agent to prevent ischemia stroke-induced neuronal injury.

Electroacupuncture Pretreatment Elicits Tolerance to Cerebral Ischemia/Reperfusion through Inhibition of the GluN2B/m-Calpain/p38 MAPK Proapoptotic Pathway.

Background As one of the first steps in the pathology of cerebral ischemia, glutamate-induced excitotoxicity progresses too fast to be the target of postischemic intervention. However, ischemic preconditioning including electroacupuncture (EA) might elicit cerebral ischemic tolerance through ameliorating excitotoxicity. Objective To investigate whether EA pretreatment based on TCM theory could elicit cerebral tolerance against ischemia/reperfusion (I/R) injury, and explore its potential excitotoxicity inhibition mechanism from regulating proapoptotic pathway of the NMDA subtype of glutamate receptor (GluN2B). Methods The experimental procedure included 5 consecutive days of pretreatment stage and the subsequent modeling stage for one day. All rats were evenly randomized into three groups: sham MCAO/R, MCAO/R, and EA+MCAO/R. During pretreatment procedure, only rats in the EA+MCAO/R group received EA intervention on GV20, SP6, and PC6 once a day for 5 days. Model preparation for MCAO/R or sham MCAO/R started 2 hours after the last pretreatment. 24 hours after model preparation, the Garcia neurobehavioral scoring criteria was used for the evaluation of neurological deficits, TTC for the measurement of infarct volume, TUNEL staining for determination of neural cell apoptosis at hippocampal CA1 area, and WB and double immunofluorescence staining for expression and the cellular localization of GluN2B and m-calpain and p38 MAPK. Results This EA pretreatment regime could improve neurofunction, decrease cerebral infarction volume, and reduce neuronal apoptosis 24 hours after cerebral I/R injury. And EA pretreatment might inhibit the excessive activation of GluN2B receptor, the GluN2B downstream proapoptotic mediator m-calpain, and the phosphorylation of its transcription factor p38 MAPK in the hippocampal neurons after cerebral I/R injury. Conclusion The EA regime might induce tolerance against I/R injury partially through the regulation of the proapoptotic GluN2B/m-calpain/p38 MAPK pathway of glutamate.

Neuroprotection by μ-calpain and matrix metalloproteinases inhibition by Piroxicam in cerebral ischemia: an in silico study

An intervention to reduce brain injury before, during, or after an ischemic injury irrespective of the cause remains an exciting prospect. Increasing evidences suggest that the excessive activation of calcium-dependent neutral proteases, calpains, could play a key role in the pathology of cerebral ischemia. Targeting calcium-activated proteolysis could, therefore, be an alternative strategy for protecting neurons against post-ischemic injury. In addition, blood brain barrier (BBB) breakdown after stroke is linked to the up-regulation of metalloproteinases (MMPs) and inflammation. MMPs are zinc-dependent endopeptidases which are capable of degrading many types of extracellular matrix proteins and are also involved in the process of tissue remodeling in various pathologic conditions. Previous studies suggest that MMPs, in particular MMP-2 and MMP-9, are deleterious in the brain after stroke. In acute stage after ischemic stroke, the effects of MMP activity is correlated to the degradation of neurovascular matrix and opening of the BBB which promotes vasogenic edema and results in neurological deficits. Hence, calpain and MMPs are considered to be a potential drug target to develop novel therapeutics against cerebral ischemia. To explore such, we have tried to study the interaction of Piroxicam with calpain and MMPs by exploring molecular docking studies. The in silico studies revealed that Piroxicam occupied the active site of μ-calpain, MMP-2, and MMP-9. The binding energy of the complexes was −5.27, −9.75, and −6.47 kcal/mol for μ-calpain, MMP-2, and MMP-9, respectively. On the basis of these in silico studies collectively, we hypothesize for the first time that Piroxicam might have a strong potential to inhibit μ-calpain, MMP-2, and MMP-9 concomitantly which can be explored as a potent therapeutic target for treating cerebral ischemia in future.

Hypothermia protects the brain from transient global ischemia/reperfusion by attenuating endoplasmic reticulum response-induced apoptosis through CHOP.

Endoplasmic reticulum (ER) stress has been implicated in the pathology of cerebral ischemia. Apoptotic cell death occurs during prolonged period of stress or when the adaptive response fails. Hypothermia blocked the TNF or Fas-mediated extrinsic apoptosis pathway and the mitochondria pathway of apoptosis, however, whether hypothermia can block endoplasmic reticulum mediated apoptosis is never known. This study aimed to elucidate whether hypothermia attenuates brain cerebral ischemia/reperfusion (I/R) damage by suppressing ER stress-induced apoptosis. A 15 min global cerebral ischemia rat model was used in this study. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) positive cells in hippocampus CA1 were assessed after reperfusion of the brain. The expressions of C/EBP-homolo gous protein (CHOP) and glucose-regulated protein 78 (GRP78) in ischemic hippocampus CA1 were measured at 6, 12, 24 and 48 h after reperfusion. The results showed that hypothermia significantly attenuated brain I/R injury, as shown by reduction in cell apoptosis, CHOP expression, and increase in GRP78 expression. These results suggest that hypothermia could protect brain from I/R injury by suppressing ER stress-induced apoptosis.

Ischemic postconditioning protects brain from ischemia/reperfusion injury by attenuating endoplasmic reticulum stress-induced apoptosis through PI3K-Akt pathway

Endoplasmic reticulum (ER) stress has been implicated in the pathology of cerebral ischemia. During prolonged period of stress or when the adaptive response fails, apoptotic cell death ensues. Cerebral ischemic postconditioning (Postcond) has been shown to reduce cerebral ischemia/reperfusion (I/R) injury in both focal and global cerebral ischemia model. However, the mechanism remains to be understood. This study aimed to elucidate whether Postcond attenuates brain I/R damage by suppressing ER stress-induced apoptosis and if the phosphatidylinositol-3kinase/Akt (PI3K/Akt) pathway is involved. A focal cerebral ischemia rat model was used in the study. Rat brain infarct size and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) positive cells in ischemic penumbra were assessed after reperfusion of the brain. The expressions of C/EBP-homologous protein (CHOP), caspase-12, glucose-regulated protein 78 (GRP78) and the phosphorylation of Akt (Ser473) in ischemic penumbra were measured after reperfusion. Our results showed that Postcond significantly attenuated brain I/R injury, as shown by reduction in infarct size, cell apoptosis, CHOP expression, caspase-12 activation and increase in GRP78 expression. LY294002, a phosphoinositide 3-kinase inhibitor, increased the number of TUNEL-positive cells suppressed by Postcond in penumbra. In addition, LY294002 diminished the effect of Postcond on the activation of CHOP, caspase-12 and GRP78. These results suggest that Postcond protects brain from I/R injury by suppressing ER stress-induced apoptosis and PI3K/Akt pathway is involved.

The Role of Excitatory Amino Acid Transporters in Cerebral Ischemia

Glutamate is an excitatory neurotransmitter that plays a major role in the pathogenesis of ischemia brain injury. The regulation of glutamate neurotransmission is carried out by excitatory amino acid transporters (EAATs) that act through reuptake of glutamate into cells. EAATs may also release glutamate into the extracellular space in a calcium-independent manner during ischemia and dysfunction of EAATs is specifically implicated in the pathology of cerebral ischemia. Recent studies show that up-regulation of EAAT2 provides neuroprotection during ischemic insult. This review summarizes current knowledge regarding the role of EAATs in cerebral ischemia.

Multiple apoptogenic proteins are involved in the nuclear translocation of Apoptosis Inducing Factor during transient focal cerebral ischemia in rat

Apoptosis Inducing Factor is a mitochondrial protein which upon translocation to nucleus causes large scale DNA fragmentation. The stimulus for the cytosolic release and nuclear translocation for this protein still remains to be understood. The role of calpains, cathepsin-b, Poly ADP (ribose) Polymerase and granzyme-b in the nuclear translocation of AIF has been investigated in the pathology of cerebral ischemia. Calpains, cathepsin-b and PARP-1 which were mostly confined to cytosol, lysosomes and nucleus respectively were found to be elevated in the mitochondrial fraction interacting with AIF in the western blot analysis and double immunofluorescence analysis. Western blot and immunohistochemical analysis revealed elevated levels of granzyme-b secreted by cytotoxic T lymphocytes and natural killer cells in the infarct of ischemic mouse brain. Co-immunoprecipitation revealed and western blot analysis the interaction and break down of Heat Shock Protein-70 an endogenous inhibitor of AIF into signature fragments by granzyme-b facilitating the nuclear translocation of AIF. Break down of HSP-70 correlated with the nuclear translocation of AIF observed in western and immunohistochemical analysis. These results indicate that multiple proteases were involved in the nuclear translocation of AIF during the pathology of cerebral ischemia.

Activation of cPLA2 and sPLA2 in astrocytes exposed to simulated ischemia in vitro

Both cytosolic PLA(2) (cPLA(2)) and secretory PLA(2) (sPLA(2)) have been implicated in pathology of cerebral ischemia. However, which of PLA(2) isoforms in astrocytes is responsible for arachidonic acid (AA) release contributing to their ischemic injury remains to be determined. The aim of the present study was to investigate the time-dependent activation of cPLA(2) and sPLA(2) in astrocytes exposed to combined oxygen glucose deprivation (OGD) as well as to evaluate the effectiveness of their pharmacological blockage as a method of preventing ischemic damage of the glial cells. It was shown that exposure of cultured astrocytes to OGD (0.5-24h) causes an increase in cPLA(2) and sPLA(2) expression and activity. The role of AA liberated mainly by cPLA(2) in the process of apoptosis was also demonstrated. To confirm the specific role of cPLA(2) and sPLA(2) in the mechanism of cells injury by OGD exposure, the effect of AACOCF(3) as cPLA(2) inhibitor and 12-epi-scalaradial as sPLA(2) inhibitor on AA release was examined. It was proved that simultaneous pharmacological blockade of enzymatic activity of cPLA(2) and sPLA(2) during OGD by AACOCF(3) and 12-epi-scalaradial substantially improves survival of ischemic injured glial cells.

TIAs and the pathology of cerebral ischemia.

The term “transient ischemic attack,” or TIA, refers to a clinically detectable event that is neurologically transient. Implicit in the definition is the assumption that the underlying cerebrovascular occlusion is also transient, i.e., that there is an arterial occlusion responsible for the symptoms, the obstruction is relieved and, through the restitution of flow, there is no residual injury but rather a complete recovery from neurologic symptoms. This sequence presumes that any neuron injury is short-lived and that there is no detectable evidence of sustained injury. An alternative to this accepted view of TIA is the hypothesis that, in the setting of transient ischemia, there is residual and permanent injury to neurons and their supporting cells (e.g., glia, astrocytes, microvessels) that remains clinically silent. This section of the supplement examines the evidence supporting the view that the pathologic accompaniment of TIA is a permanent type of neuron injury. I focus on two questions drawn from experimental data: (a) whether a vascular lesion, in this particular case, a carotid artery lesion, can continuously produce local arterial occlusion with downstream embolization into the cerebral microvasculature, and (b) whether experimental transient occlusions of the microvasculature affect neuron integrity of the ischemic territory and whether that impact of emboli produces permanent injury (table). View this table: Table Evidence for permanent neuron injury after transient arterial occlusion The usually rapid resolution of neurologic symptoms that typifies the course of TIA has led to the suggestion that the neuron and cell dysfunctions underlying the symptoms are transient. This hypothesis is based on a scenario involving an evanescent flow impediment in the affected territory in which local flow is rapidly restored. The evidence for this sequence is sparse. However, because angiographically detected obstructions of short duration have been documented in ischemic stroke patients and in experimental animal models, such a sequence …

Adenosine and cerebral ischemia: therapeutic future or death of a brave concept?

Numerous studies have consistently shown that agonist stimulation of adenosine A 1 receptors results in a significant reduction of morbidity and mortality associated with global and focal brain ischemia in animals. Based on these observations, several authors have suggested utilization of adenosine A 1 receptors as targets for the development of clinically viable drugs against ischemic brain disorders. Recent advent of adenosine A 1 receptor agonists characterized by lowered cardiovascular effects added additional strength to this argument. On the other hand, although cardioprotective, adenosine A 3 receptor agonists proved severely cerebrodestructive when administered prior to global ischemia in gerbils. Moreover, stimulation of adenosine A 3 receptors appears to reduce the efficacy of some of the neuroprotective actions mediated by adenosine A 1 receptors. The review discusses the possible role of adenosine receptor subtypes (A 1, A 2, and A 3) in the context of their involvement in the pathology of cerebral ischemia, and analyzes putative strategies for the development of clinically useful strategies based on adenosine and its receptors. It also stresses the need for further experimental studies before definitive conclusions on the usefulness of the adenosine concept in the treatment of brain ischemia can be made.

Adenosine and cerebral ischemia: therapeutic future or death of a brave concept?

Numerous studies have consistently shown that agonist stimulation of adenosine A 1 receptors results in a significant reduction of morbidity and mortality associated with global and focal brain ischemia in animals. Based on these observations, several authors have suggested utilization of adenosine A 1 receptors as targets for the development of clinically viable drugs against ischemic brain disorders. Recent advent of adenosine A 1 receptor agonists characterized by lowered cardiovascular effects added additional strength to this argument. On the other hand, although cardioprotective, adenosine A 3 receptor agonists proved severely cerebrodestructive when administered prior to global ischemia in gerbils. Moreover, stimulation of adenosine A 3 receptors appears to reduce the efficacy of some of the neuroprotective actions mediated by adenosine A 1 receptors. The review discusses the possible role of adenosine receptor subtypes (A 1, A 2, and A 3) in the context of their involvement in the pathology of cerebral ischemia, and analyzes putative strategies for the development of clinically useful strategies based on adenosine and its receptors. It also stresses the need for further experimental studies before definitive conclusions on the usefulness of the adenosine concept in the treatment of brain ischemia can be made.

Alterations of Bcl-2 family proteins precede cytoskeletal proteolysis in the penumbra, but not in infarct centres following focal cerebral ischemia in mice

Apoptosis has drawn attention in ischemic neuronal death recently. However, studies of apoptosis in cerebral ischemia have concentrated largely in DNA fragmentation, a late phase in apoptotic nuclei, at the expense of possible primary ischemic targets at the subcellular level and of upstream apoptotic signalling. To assess those issues, we used an intraluminal middle cerebral artery occlusion model in mice with or without reperfusion, and examined sequential changes of Bcl-2 family proteins modulating apoptotic signalling immunohistochemically and studied nuclear DNA fragmentation, to compare their chronology in relation to the development of infarct as detected by loss of microtubule-associated protein-2, an early marker of cytoplasmic damage. In the centre of the lesion, Bax protein increased and Bcl-2 and Bcl-x proteins decreased after loss of microtubule-associated protein-2 antigenicity occurred, but at the border of the lesion, the former changes preceded loss of microtubule-associated protein-2 antigenicity. Additionally, close morphologic analysis of DNA fragmentation in situ indicated that transient ischemia predominantly induced apoptotic cells but permanent ischemia produced necrosis of cells in the centre of the lesion.The contrasting cell death mechanisms, apoptosis and necrosis, are selectively involved in the pathology of cerebral ischemia, depending on its severity.

The effect of lesions in the locus coeruleus on the physiological responses of the cerebral blood vessels in cats

The effects on cerebral blood flow (CBF) of lesions placed stereotactically in or near the locus coeruleus were studied in 15 lightly anesthetized cats; 5 control cats in which the electrode was placed but no lesion created, and 10 experimental cats in which a lesion was created. The response of CBF to changes in Pa co 2 and in mean arterial blood pressure was determined by 133Xe-washout studies 10 days after the stereotactic procedures. The sites of the lesions were studied histologically, and their effects on catecholamine concentrations in the paraventricular hypothalamic nucleus, anterior ventral nucleus of the thalamus, and parietal cortex were determined by radiochemical assay. Control animals and those with lesions near, but not in, the locus coeruleus had normal Pa co 2-CBF response curves and normal catecholamine concentrations in the areas of biopsy. Bilateral destruction of the locus coeruleus was confirmed in 3 animals on histological examination and in these animals there were decreased levels of catecholamines in the areas of assay, higher resting CBFs at normocapnia, and significantly abnormal CBF-Pa co 2 response curves. The autoregulatory response to changes in perfusion pressure was preserved. Thus, noradrenergic neurons originating in the locus coeruleus may contribute to the control of intraparenchymal cerebral vessels and disturbance of this control may be important in the pathology of cerebral ischemia.