n-(n-(3-5-difluorophenacetyl)alanyl)phenylglycine-tert-butyl-ester and Infarction--Middle-Cerebral-Artery

Recent studies revealed that folic acid deficiency (FD) increased the likelihood of stroke and aggravated brain injury after focal cerebral ischaemia. The microglia-mediated inflammatory response plays a crucial role in the complicated pathologies that lead to ischaemic brain injury. However, whether FD is involved in the activation of microglia and the neuroinflammation after experimental stroke and the underlying mechanism is still unclear. The aim of the present study was to assess whether FD modulates the Notch1/nuclear factor kappa B (NF-κB) pathway and enhances microglial immune response in a rat middle cerebral artery occlusion-reperfusion (MCAO) model and oxygen-glucose deprivation (OGD)-treated BV-2 cells. Our results exhibited that FD worsened neuronal cell death and exaggerated microglia activation in the hippocampal CA1, CA3 and Dentate gyrus (DG) subregions after cerebral ischaemia/reperfusion. The hippocampal CA1 region was more sensitive to ischaemic injury and FD treatment. The protein expressions of proinflammatory cytokines such as tumour necrosis factor-α, interleukin-1β and interleukin-6 were also augmented by FD treatment in microglial cells of the post-ischaemic hippocampus and in vitro OGD-stressed microglia model. Moreover, FD not only dramatically enhanced the protein expression levels of Notch1 and NF-κB p65 but also promoted the phosphorylation of pIkBα and the nuclear translocation of NF-κB p65. Blocking of Notch1 with N-[N-(3, 5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester partly attenuated the nuclear translocation of NF-κB p65 and the protein expression of neuroinflammatory cytokines in FD-treated hypoxic BV-2 microglia. These results suggested that Notch1/NF-κB p65 pathway-mediated microglial immune response may be a molecular mechanism underlying cerebral ischaemia-reperfusion injury worsened by FD treatment.

Topics: Animals; Brain Injuries; Brain Ischemia; Cell Line; Cytokines; Dipeptides; Folic Acid Deficiency; Glucose; Hippocampus; Infarction, Middle Cerebral Artery; Inflammation; Male; Mice; Microglia; Neurons; Oxygen; Rats, Sprague-Dawley; Receptor, Notch1; Signal Transduction; Transcription Factor RelA

Perioperative discontinuation of aspirin is often considered due to bleeding concern. We determined whether this discontinuation affected neurological outcome after brain ischemia.. Adult male Sprague-Dawley rats were subjected to a 90-minute right middle cerebral arterial occlusion (MCAO). They received 30 mg/kg/day aspirin via gastric gavage: 1) for 2 days at 5 days before MCAO; 2) for 2 days at 5 days before MCAO and for 3 days after MCAO; 3) for 7 days before MCAO; or 4) for 7 days before MCAO and for 3 days after MCAO. Neurological outcome was evaluated 3 days after the MCAO. Ischemic penumbral cortex was harvested 1 or 3 days after MCAO for determining Notch intracellular domain (NICD), IL-6 and IL-1β levels.. Aspirin given by regimen 2 and 3 but not by regimen 1 improved neurological outcome. Neuroprotection was achieved by N-[N-(3,5-Difluorophenacetyl)-L-alanyl]-S-phenylglycine t-butyl ester (DAPT), a Notch activation inhibitor. DAPT and aspirin given only by regimen 2 and 3 reduced NICD, IL-6 and IL-1β in the ischemic penumbral cortex. NICD was found in microglial nuclei. Microglial activation in the ischemic tissues was inhibited by aspirin.. Aspirin use during the perioperative period provides neuroprotection. Inhibition of Notch activation and neuroinflammation may contribute to the neuroprotection of aspirin.

Topics: Analysis of Variance; Animals; Anti-Inflammatory Agents, Non-Steroidal; Aspirin; Brain Infarction; Dipeptides; Disease Models, Animal; Enzyme Inhibitors; Enzyme-Linked Immunosorbent Assay; Infarction, Middle Cerebral Artery; Interleukin-1beta; Interleukin-6; Male; Nervous System Diseases; Psychomotor Performance; Rats, Sprague-Dawley; Time Factors

Tight junction protein degradation is a principal characteristic of the blood-brain barrier (BBB) damage that occurs during brain ischemia.. We investigated the mechanisms of occludin degradation that underlie permanent middle cerebral artery occlusion (pMCAO) in rats.. Western blot and Co-immunoprecipitation data indicated ubiquitination and degradation of occludin in brain after pMCAO, which was consistent with ZO-1 degradation in penumbra regions as observed at 24 h after pMCAO. We further investigated candidate protease(s) responsible for the degradation of occludin during pMCAO. The intraventricular administration of γ-secretase blocker DAPT significantly inhibited the pMCAO-induced neurovascular damage, whereas ALLM and Batimastat, which are inhibitors of calpain and metalloproteinase proteases, respectively, were less effective. Notably, we found that DAPT significantly inhibited BBB disruption in comparison with vehicle treatment, as assessed by Evans blue excretion. Interestingly, the confocal immunostaining revealed that activation of the E3 ubiquitin ligase Itch is associated with degradation of occludin in brain microvessels following ischemia. Furthermore, our data demonstrate that the inhibition of γ-secretase signaling and the itch-mediated ubiquitination of occludin likely underlie the vasoprotective effect of DAPT after pMCAO.. The γ-secretase blocker DAPT reduces the permeability of the BBB by decreasing the ubiquitination and degradation of occludin during permanent brain ischemia, suggesting that γ-secretase may represent a novel therapeutic target for preventing neurovascular damage.

Topics: Amyloid Precursor Protein Secretases; Analysis of Variance; Animals; Blood-Brain Barrier; Dipeptides; Disease Models, Animal; Endothelium, Vascular; Enzyme Inhibitors; Gene Expression Regulation; Immunoprecipitation; Infarction, Middle Cerebral Artery; Male; Occludin; Permeability; Platelet Endothelial Cell Adhesion Molecule-1; Rats; Rats, Sprague-Dawley; Tight Junctions; Time Factors; Ubiquitin-Protein Ligases; Ubiquitination; Zonula Occludens-1 Protein

Abnormal β-amyloid (Aβ) deposits in the thalamus have been reported after cerebral cortical infarction. In this study, we investigated the association of Aβ deposits, with the secondary thalamic damage after focal cortical infarction in rats. Thirty-six stroke-prone renovascular hypertensive rats were subjected to distal middle cerebral artery occlusion (MCAO) and then randomly divided into MCAO, vehicle, and N-[N-(3,5-difluorophenacetyl)-L-alanyl]-S-phenylglycine t-butyl ester (DAPT) groups and 12 sham-operated rats as control. The DAPT was administered orally at 72 hours after MCAO. Seven days after MCAO, sensory function, neuron loss, and glial activation and proliferation were evaluated using adhesive removal test, Nissl staining, and immunostaining, respectively. Thalamic Aβ accumulation was evaluated using immunostaining and enzyme-linked immunosorbent assay (ELISA). Compared with vehicle group, the ipsilateral thalamic Aβ, neuronal loss, glial activation and proliferation, and the mean time to remove the stimulus from right forepaw significantly decreased in DAPT group. The mean time to remove the stimulus from the right forepaw and thalamic Aβ burden were both negatively correlated with the number of thalamic neurons. These findings suggest that Aβ deposits are associated with the secondary thalamic damage. Reduction of thalamic Aβ by γ-secretase inhibitor may attenuate the secondary damage and improve sensory function after cerebral cortical infarction.

Topics: Amyloid beta-Peptides; Amyloid Precursor Protein Secretases; Animals; Cell Proliferation; Cerebral Infarction; Coloring Agents; Dipeptides; Enzyme-Linked Immunosorbent Assay; Image Processing, Computer-Assisted; Immunohistochemistry; Infarction, Middle Cerebral Artery; Neuroglia; Neurons; Protease Inhibitors; Rats; Rats, Inbred SHR; Rats, Sprague-Dawley; Sensation; Thalamus