Cytosolic phospholipase A2 alpha amplifies early cyclooxygenase-2 expression, oxidative stress and MAP kinase phosphorylation after cerebral ischemia in mice.

The Department of Anesthesiology and Critical Care Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
Journal of Neuroinflammation (Impact Factor: 4.35). 01/2010; 7:42. DOI: 10.1186/1742-2094-7-42
Source: PubMed

ABSTRACT The enzyme cytosolic phospholipase A2 alpha (cPLA2alpha) has been implicated in the progression of cerebral injury following ischemia and reperfusion. Previous studies in rodents suggest that cPLA2alpha enhances delayed injury extension and disruption of the blood brain barrier many hours after reperfusion. In this study we investigated the role of cPLA2alpha in early ischemic cerebral injury.
Middle cerebral artery occlusion (MCAO) was performed on cPLA2alpha+/+ and cPLA2alpha-/- mice for 2 hours followed by 0, 2, or 6 hours of reperfusion. The levels of cPLA2alpha, cyclooxygenase-2, neuronal morphology and reactive oxygen species in the ischemic and contralateral hemispheres were evaluated by light and fluorescent microscopy. PGE2 content was compared between genotypes and hemispheres after MCAO and MCAO and 6 hours reperfusion. Regional cerebral blood flow was measured during MCAO and phosphorylation of relevant MAPKs in brain protein homogenates was measured by Western analysis after 6 hours of reperfusion.
Neuronal cPLA2alpha protein increased by 2-fold immediately after MCAO and returned to pre-MCAO levels after 2 hours reperfusion. Neuronal cyclooxygenase-2 induction and PGE2 concentration were greater in cPLA2alpha+/+ compared to cPLA2alpha-/- ischemic cortex. Neuronal swelling in ischemic regions was significantly greater in the cPLA2alpha+/+ than in cPLA2alpha-/- brains (+/+:2.2+/-0.3 fold vs. -/-:1.7+/-0.4 fold increase; P<0.01). The increase in reactive oxygen species following 2 hours of ischemia was also significantly greater in the cPLA2alpha+/+ ischemic core than in cPLA2alpha-/- (+/+:7.12+/-1.2 fold vs. -/-:3.1+/-1.4 fold; P<0.01). After 6 hours of reperfusion ischemic cortex of cPLA2alpha+/+, but not cPLA2alpha-/-, had disruption of neuron morphology and decreased PGE2 content. Phosphorylation of the MAPKs-p38, ERK 1/2, and MEK 1/2-was significantly greater in cPLA2a+/+ than in cPLA2alpha-/- ischemic cortex 6 hours after reperfusion.
These results indicate that cPLA2alpha modulates the earliest molecular and injury responses after cerebral ischemia and have implications for the potential clinical use of cPLA2alpha inhibitors.

  • [Show abstract] [Hide abstract]
    ABSTRACT: Abstract Significance: Neuronal superoxide production contributes to cell death in both glutamate excitotoxicity and brain ischemia (stroke). NADPH oxidase-2 (NOX2) is the major source of neuronal superoxide production in these settings, and regulation of NOX2 activity can thereby influence outcome in stroke. Recent Advances: Reduced NOX2 activity can rescue cells from oxidative stress and cell death that otherwise occur in excitotoxicity and ischemia. NOX2 activity is regulated by several factors previously shown to affect outcome in stroke, including glucose availability, intracellular pH, protein kinase ζ/δ, casein kinase 2, phosphoinositide-3-kinase, Rac1/2, and phospholipase A2. The newly identified functions of these factors as regulators of NOX2 activity suggest alternative mechanisms for their effects on ischemic brain injury. Critical Issues: Key aspects of these regulatory influences remain unresolved, including the mechanisms by which rac1 and phospholipase activities are coupled to N-methyl-D-aspartate (NMDA) receptors, and whether superoxide production by NOX2 triggers subsequent superoxide production by mitochondria. Future Directions: It will be important to establish whether interventions targeting the signaling pathways linking NMDA receptors to NOX2 in brain ischemia can provide a greater neuroprotective efficacy or a longer time window to treatment than provided by NMDA receptor blockade alone. It will likewise be important to determine whether dissociating superoxide production from the other signaling events initiated by NMDA receptors can mitigate the deleterious effects of NMDA receptor blockade. Antioxid. Redox Signal. 00, 000-000.
    Antioxidants & Redox Signaling 03/2014; · 8.20 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Phospholipases A2 (PLA2s) are important enzymes for the metabolism of fatty acids in membrane phospholipids. Among the three major classes of PLA2s in the mammalian system, the group IV calcium-dependent cytosolic PLA2 alpha (cPLA2α) has received the most attention because it is widely expressed in nearly all mammalian cells and its active participation in cell metabolism. Besides Ca(2+) binding to its C2 domain, this enzyme can undergo a number of cell-specific post-translational modifications, including phosphorylation by protein kinases, S-nitrosylation through interaction with nitric oxide (NO), as well as interaction with other proteins and lipid molecules. Hydrolysis of phospholipids by cPLA2 yields two important lipid mediators, arachidonic acid (AA) and lysophospholipids. While AA is known to serve as a substrate for cyclooxygenases and lipoxygenases, which are enzymes for the synthesis of eicosanoids and leukotrienes, lysophospholipids are known to possess detergent-like properties capable of altering microdomains of cell membranes. An important feature of cPLA2 is its link to cell surface receptors that stimulate signaling pathways associated with activation of protein kinases and production of reactive oxygen species (ROS). In the central nervous system (CNS), cPLA2 activation has been implicated in neuronal excitation, synaptic secretion, apoptosis, cell-cell interaction, cognitive and behavioral function, oxidative-nitrosative stress, and inflammatory responses that underline the pathogenesis of a number of neurodegenerative diseases. However, the types of extracellular agonists that target intracellular signaling pathways leading to cPLA2 activation among different cell types and under different physiological and pathological conditions have not been investigated in detail. In this review, special emphasis is given to metabolic events linking cPLA2 to activation in neurons, astrocytes, microglial cells, and cerebrovascular cells. Understanding the molecular mechanism(s) for regulation of this enzyme is deemed important in the development of new therapeutic targets for the treatment and prevention of neurodegenerative diseases.
    Molecular Neurobiology 02/2014; · 5.47 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The aims of the present study were to investigate the protective effect of tanshinone IIA on the brain and its therapeutic time window in a rat model of cerebral ischemia-reperfusion. The rat model of cerebral ischemia-reperfusion was established by suture occlusion. In an initial experiment, male Sprague-Dawley (SD) rats were randomly divided into control cerebral ischemia-reperfusion rat model, tanshinone IIA1 (TSA1), tanshinone IIA4 (TSA4), tanshinone IIA6 (TSA6) and tanshinone IIA12 (TSA12) groups (n=8 per group). The rats in the control group were given 4 ml phosphate-buffered saline (PBS) intraperitoneally following suture occlusion. The other groups were respectively treated with 25 mg/kg tanshinone IIA intraperitoneally at 1, 4, 6 and 12 h following the initiation of reperfusion and once a day for a total of three days. The grades of neurologic impairment and volume of cerebral infarction of each group were measured 72 h after suture occlusion. In another experiment, 16 male SD rats were randomly divided into a 6 h reperfusion group and a 24 h reperfusion group following drug administration. The rats in each group were further divided into a control subgroup (4 ml PBS) and a tanshinone IIA subgroup (25 mg/kg). The rats were immediately administered their respective treatments following the establishment of the model. The rats were decapitated 6 and 24 h after the initiation of reperfusion. The expression levels of cytoplasmic thioredoxin (Trx-1) and mitochondrial thioredoxin (Trx-2) in the ischemic penumbra were determined by western blot analysis. The nitric oxide (NO) levels, and total NO synthase (tNOS) and inducible NO synthase (iNOS) activities in the rat blood were measured using a reagent kit. The changes in cerebral blood flow were evaluated by Doppler imaging. The grade of neurological impairment of the TSA1 group was statistically lower than that of the other groups (P<0.05). The cerebral infarction volume results showed that the volumes of infarction in the TSA1 and TSA4 groups were lower than those in the other groups (P<0.05). Tanshinone IIA significantly increased cerebral blood flow compared with that of the control group (P<0.05). Moreover, tanshinone IIA significantly increased the expression levels of Trx-1 and Trx-2 compared with those in the control group (P<0.05). Tanshinone IIA significantly decreased the NO levels and iNOS and tNOS activities compared with those of the control group (P<0.05). However, the iNOS activity in the rats in the 6 h reperfusion group was not statistically significantly different from that of the respective control group (P>0.05). Tanshinone IIA has a protective effect on the cranial nerves when administered during the initial stages of cerebral ischemia. This protective effect is associated with an improvement of cerebral blood flow as well as an increase in anti-oxygen radical and anti-inflammatory activities.
    Experimental and therapeutic medicine 11/2014; 8(5):1616-1622. · 0.34 Impact Factor

Full-text (2 Sources)

Available from
May 19, 2014