Nafamostat mesilate attenuates neuronal damage in a rat model of transient focal cerebral ischemia through thrombin inhibition.
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ABSTRACT:
SUBMITTER: Chen T
PROVIDER: S-EPMC4078306 | biostudies-other | 2014
REPOSITORIES: biostudies-other
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Project description:MicroRNAs (miRNAs) are approximately 22 nucleotides long, noncoding RNAs that control cellular function by either degrading mRNAs or arresting their translation. To understand their functional significance in ischemic pathophysiology, we profiled miRNAs in adult rat brain as a function of reperfusion time after transient middle cerebral artery occlusion. Of the 238 miRNAs evaluated, 8 showed increased and 12 showed decreased expression at least at 4 out of 5 reperfusion time points studied between 3 h and 3 days compared with sham. Of those, 17 showed >5 fold change. Bioinformatics analysis indicated a correlation between miRNAs altered to several mRNAs known to mediate inflammation, transcription, neuroprotection, receptors function, and ionic homeostasis. Antagomir-mediated prevention of mir-145 expression led to an increased protein expression of its downstream target superoxide dismutase-2 in the postischemic brain. In silico analysis showed sequence complementarity of eight miRNAs induced after focal ischemia to 877 promoters indicating the possibility of noncoding RNA-induced activation of gene expression. The mRNA expression of the RNases Drosha and Dicer, cofactor Pasha, and the pre-miRNA transporter exportin-5, which modulate miRNA biogenesis, were not altered after transient middle cerebral artery occlusion. Thus, the present studies indicate a critical role of miRNAs in controlling mRNA transcription and translation in the postischemic brain.
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Project description:Neuroinflammation is one of the important factors aggravating brain injury after ischemic stroke. We aimed to investigate the effects of cerebrolysin (CBL) on neuroinflammation in vivo and in vitro and the underlying mechanisms. The gene expressions of pro-inflammatory factors and anti-inflammatory factors were analyzed by real time PCR in rat transient middle cerebral artery occlusion (tMCAO) model, lipopolysaccharides-induced neuroinflammatory mice model and LPS-treated mouse primary microglia cells. The neuroprotective effects of CBL were evaluated by infarct size, Longa test and Rotarod test for long-term functional recovery in rats subjected to ischemia. The role of CREB/PGC-1α pathway in anti-neuroinflammatory effect of CBL was also determined by real time PCR and Western blotting. In the tMCAO model, administration of CBL at 3 h post-ischemia reduced infarct volume, promoted long-term functional recovery, decreased the gene expression of pro-inflammatory factors and increased the gene expression of anti-inflammatory factors. Correspondingly, in LPS-induced neuroinflammatory mice model, CBL treatment attenuated sickness behavior, decreased the gene expression of pro-inflammatory factors, and increased the gene expression of anti-inflammatory factors. In in vitro and in vivo experiments, CBL increased the protein expression levels of PGC-1α and phosphorylated CREB to play anti-inflammatory effect. Additionally, the application of the specific CREB inhibitor, 666-15 compound could effectively reverse the anti-inflammatory effect of CBL in primary mouse microglia cells and anti-ischemic brain injury of CBL in rats subjected to tMCAO. In conclusion, CBL ameliorated cerebral ischemia injury through reducing neuroinflammation partly via the activation of CREB/PGC-1α pathway and may play a therapeutic role as anti-neuroinflammatory agents in the brain disorders associated with neuroinflammation.
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Project description:Whether the effect of miR-181a is sexually dimorphic in stroke is unknown. Prior work showed protection of male mice with miR-181a inhibition. Estrogen receptor-α (ERα) is an identified target of miR181 in endometrium. Therefore we investigated the separate and joint effects of miR-181a inhibition and 17β-estradiol (E2) replacement after ovariectomy. Adult female mice were ovariectomized and implanted with an E2- or vehicle-containing capsule for 14d prior to 1h middle cerebral artery occlusion (MCAO). Each group received either miR-181a antagomir or mismatch control by intracerebroventricular injection 24h before MCAO. After MCAO neurologic deficit and infarct volume were assessed. Primary male and female astrocyte cultures were subjected to glucose deprivation with miR-181a inhibitor or transfection control, and E2 or vehicle control, with/without ESRα knockdown with small interfering RNA. Cell death was assessed by propidium iodide staining, and lactate dehydrogenase assay. A miR-181a/ERα target site blocker (TSB), with/without miR-181a mimic, was used to confirm targeting of ERα by miR-181a in astrocytes. Individually, miR-181a inhibition or E2 decreased infarct volume and improved neurologic score in female mice, and protected male and female astrocyte cultures. Combined miR-181a inhibition plus E2 afforded greater protection of female mice and female astrocyte cultures, but not in male astrocyte cultures. MiR-181a inhibition only increased ERα levels in vivo and in female cultures, while ERα knockdown with siRNA increased cell death in both sexes. Treatment with ERα TSB was strongly protective in both sexes. In conclusion, the results of the present study suggest miR-181a inhibition enhances E2-mediated stroke protection in females in part by augmenting ERα production, a mechanism detected in female mice and female astrocytes. Sex differences were observed with combined miR-181a inhibition/E2 treatment, and miR-181a targeting of ERα.
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Project description:Background Systemic innate immune priming is a recognized sequela of post-ischemic neuroinflammation and contributor to delayed neurodegeneration. Given mounting evidence linking acute stroke with reactive lung inflammation, we asked whether enhanced expression of the endogenous antioxidant extracellular superoxide dismutase 3 (SOD3) produced by alveolar type II pneumocytes would protect the lung from transient global cerebral ischemia and the brain from the delayed effects of ischemia-reperfusion. Methods and Results Following 15 minutes of global cerebral ischemia or sham conditions, transgenic SOD3 and wild-type mice were followed daily for changes in weight, core temperature, and neurological function. Three days after reperfusion, arterial and venous samples were collected for complete blood counts, flow cytometry, and SOD3 protein blotting, and immunohistochemistry was performed on lung and brain tissue to assess tissue injury, blood-brain barrier permeability, and neutrophil transmigration. Relative to ischemic controls, transgenic SOD3 mice performed better on functional testing and exhibited reduced peripheral neutrophil activation, lung inflammation, and blood-brain barrier leak. Once released from the lung, SOD3 was predominantly not cell associated and depleted in the venous phase of circulation. Conclusions In addition to reducing the local inflammatory response to cerebral ischemia, targeted enrichment of SOD3 within the lung confers distal neuroprotection against ischemia-reperfusion injury. These data suggest that therapies geared toward enhancing adaptive lung-neurovascular coupling may improve outcomes following acute stroke and cardiac arrest.
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