u-0126 and Brain-Damage--Chronic

The objective of this study is to explore the neuroprotective effect and mechanism of picroside II on ERK1/2-COX2 signal transduction pathway after cerebral ischemic injury in rats. Focal cerebral ischemic models were established by inserting monofilament threads into the middle cerebral artery in 200 Wistar rats. Twenty four rats were randomly selected into control group, while the other rats were randomly divided into six groups: model group, picroside group, lipopolysaccharide (LPS) with picroside group, U0126 with picroside group, LPS group, and U0126 group with each group containing three subgroups with ischemia at 6, 12, and 24 h. Neurobehavioral function in the rats was evaluated by modified neurological severity score points (mNSS) test; structure of neurons was observed using hematoxylin-eosin (HE) staining; apoptotic cells were counted using terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay; expressions of phosphorylated mitogen/extracellular signal-regulated kinase kinas1/2 (pMEK1/2), phosphorylated extracellular signal-regulated protein kinase1/2 (pERK1/2), and cyclooxygenase (COX2) in the cortex were determined using immunohistochemistry (IHC) and Western blot (WB); and real-time PCR was used to determine the level of COX2 mRNA. The neurological behavioral malfunction appeared in all rats with middle cerebral artery occlusion (MCAO). In the model group, neuron damage was extensive, while the neurobehavioral function score, apoptotic cell index, expression of pMEK1/2, pERK1/2, and COX2 and the level of COX2 mRNA increased significantly when compared to the control group. The peak COX2 mRNA level was in ischemia 12 h, prior to the peak in COX2 protein expression. In the picroside and U0126 groups, the neurological behavioral function was improved, and the number of apoptotic cells and the expression of pMEK1/2, pERK1/2, and COX2 decreased significantly when compared to the model group. In the LPS with picroside group, at ischemia 6 h neuron damage was extensive, and pMEK1/2, pERK1/2, and COX2 expression were much higher than in the model group. But at ischemia 12 and 24 h, the expression of pMEK1/2, pERK1/2, and COX2 decreased slightly, and the neurobehavioral function also improved slightly. In LPS group, neuron damage was extensive, pMEK1/2, pERK1/2, and COX2 expression was still at a high level, and COX2 mRNA peak arrived at ischemic 12 h. Picroside II downregulates COX2 expression after MCAO by inhibiting MEK-ERK1/2

Topics: Animals; Apoptosis; Behavior, Animal; Brain Damage, Chronic; Butadienes; Cerebral Cortex; Cinnamates; Cyclooxygenase 2; Cyclooxygenase 2 Inhibitors; Drug Evaluation, Preclinical; Enzyme Induction; Infarction, Middle Cerebral Artery; Iridoid Glucosides; Lipopolysaccharides; Male; MAP Kinase Kinase Kinases; MAP Kinase Signaling System; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Nerve Tissue Proteins; Neuroprotective Agents; Nitriles; Random Allocation; Rats; Rats, Wistar; RNA, Messenger; Severity of Illness Index; Signal Transduction

Mild hypothermia improves survival and neurological outcome after cardiac arrest, as well as increasing activation of the extracellular-signal-regulated kinase (ERK) in hippocampus. ERK signaling is involved in neuronal growth and survival. We tested the hypothesis that the beneficial effects of hypothermia required ERK activation. ERK activation was measured by immunoblotting with phosphorylation-specific antibodies. Rats (n = 8 per group) underwent 8 min of asphyxial cardiac arrest and were resuscitated with chest compressions, ventilation, epinephrine and bicarbonate. At 30 min after resuscitation, vehicle (50% saline:50% DMSO) or the ERK kinase inhibitor U0126 (100 microg) was infused into the lateral ventricle. Cranial temperature was kept at either 33 degrees C (hypothermia) or 37 degrees C (normothermia) between 1 and 24 h. Neurological function was assessed daily for 14 days. Surviving neurons were counted in the hippocampus. A dose of 100 mug U0126 inhibited ERK bilaterally for 12 to 24 h and decreased phosphorylation of the ERK substrates ATF-2 and CREB. As in previous studies, hypothermia improved survival, neurological and histological outcome after cardiac arrest. However, survival, neurological score and histology did not differ between U0126 and vehicle-treated rats after cardiac arrest. Therefore, a dose of U0126 sufficient to inhibit biochemical markers of ERK signaling in hippocampus does not alter the beneficial effects of hypothermia induced after resuscitation in rats and did not affect recovery of normothermia-treated rats. These results suggest that hypothermia-induced improvement in outcomes does not require ERK activation.

Topics: Animals; Asphyxia; Body Temperature; Brain Damage, Chronic; Butadienes; Cell Survival; Dose-Response Relationship, Drug; Enzyme Activation; Enzyme Inhibitors; Extracellular Signal-Regulated MAP Kinases; Heart Arrest; Hippocampus; Hypothermia; Hypothermia, Induced; Injections, Intraventricular; Male; Nitriles; Rats; Rats, Sprague-Dawley; Resuscitation; Signal Transduction