Transcription profiling of mouse forebrain with induced middle cerebral artery occlusion (MCAO) to study permanent focal cerebral ischemia
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ABSTRACT: profiling gene transcription in a mouse model of permanent focal cerebral ischemia that was induced by middle cerebral artery occlusion (MCAO)
Project description:We obtained the profiles of neuronal phosphoproteome after cerebral ischemia and reperfusion by isolating mice hippocampus. Hippocampus combined from either nine sham or nine focal cerebral ischemia 1.5 h and reperfusion 24 h (IR) mice were lysed, digested, labeled with different TMT tags, then pooled and analyzed by LC/LC-MS/MS. In total, we quantified 7,865 phosphopeptides,179 phosphorylation sites of 129 proteins were upregulated and 843 phosphorylation sites of 494 proteins were downregulated in hippocampus during cerebral ischemia 2 h compare with sham operation.
Project description:We obtained the profiles of neuronal proteome after cerebral ischemia and reperfusion by isolating mice hippocampus. Hippocampus combined from either nine sham or nine focal cerebral ischemia 1.5 h and reperfusion 24 h (IR) mice were lysed, digested, labeled with different TMT tags, then pooled and analyzed by LC/LC-MS/MS. In total, we quantified 5,059 proteins. We identified 142 differentially expressed proteins (t-test, p-value<0.05) after IR compared to sham groups. The results showed that 92 proteins were upregulated, and 50 proteins were downregulated after IR compared to sham groups. Gene ontology (GO) enrichment analysis of differentially expressed proteins between sham and IR groups. The results showed that the biological process of most of upregulated genes linked with immune inflammatory related responses were increased. And KEGG pathway analysis for upregulated genes showed that multiple immune inflammatory response pathways also increased significantly, such as TNF-signaling, NF-κB signaling and cytokine-cytokine receptor interaction, as well as NOD-like receptor signaling, and toll-like receptor signaling.
Project description:We obtained the profiles of neuronal phosphoproteome after cerebral ischemia onset by isolating mice hippocampus. Hippocampus combined from either ten sham or ten focal cerebral ischemia 2 h mice were lysed, digested, labeled with different TMT tags, then pooled and analyzed by LC/LC-MS/MS. Five percent of the pool was used for whole proteome analysis, and the remaining 95% was subjected to phosphoproteome profiling. In total, we quantified 5,174 proteins and 9,062 phosphopeptides. Interesting, 21 proteins were upregulated and 7 proteins were downregulated in hippocampus lysates of cerebral ischemia 2 h relative to sham base on fold change. S100a9, Alpha-2-HS-glycoprotein (Ahsg), Fibrinogen beta chain (Fga) and Complement Component C3(c3) are the top significantly changed, which were highly consistent with previous reports in cerebral ischemia injury. Using wolfpsort software to analysis the Subcellular Location, 57% of detected proteins were location to extracellular, 15% were cytoplasmic protein, another 11% were transport to nucleus, and the others were location to plasma membranes (10%), mitochondria (4%) and endoplasmic reticulum (3%). Moreover,184 phosphorylation sites of 135 proteins were upregulated and 689 phosphorylation sites of 420 proteins were downregulated in hippocampus during cerebral ischemia 2 h compare with sham operation. Employing wolfpsort software analysis the subcellular location, 50% of phosphorylated proteins were location to nucleus, 26% were cytoplasmic protein, another 16% were transport to plasma membranes, and the others were location to mitochondria (4%), extracellular (3%) and cytoskeleton (1%). Motif analysis showed that 85% were belongs to serine-type phosphorylation, about 14 were threonine-type phosphorylation and 1% were tyrosine-type phosphorylation.
Project description:This data set shows dramatic changes in gene expression in microglia isolated from C57Bl6/J mice subjected to transient middle cerebral artery occlusion, as compared to those subjected to sham surgery. Mice deficient in Mincle (Clec4e-/-) showed significantly improved injury outcomes 3 and 7 days after transient middle cerebral artery occlusion. However, when comparing changes in gene expression in microglia 24 hours after blood reperfusion, there were no differences between wild-type and Clec4e-/- mice, indicating that Mincle does not participate in early microglial activation. Wild type and Mincle knock-out (Clec4e-/-) mice. After 1 h of transient middle cerebral artery occlusion (tMCAO) and 24 h of reperfusion, mice were perfused with PBS, their brains dissected, and 2 ipsilesional hemispheres (with cerebellum and brainstem removed) pooled for microglia isolation. For sham-operated animals, the whole forebrain was used and brains were not pooled. After myelin separation by Percoll gradient centrifugation, around 80,000 CD45intermediate, CD11b+ microglial cells were sorted from each sample. Sham samples n=3, tMCAO samples n=5.
Project description:Analysis of changes in gene expression induced by a transient (90') middle cerebral artery occulsion, at several time points post stroke induction, with the effect of a treatment with the drug FK506 studied at a single time point.
Project description:This study aims at a comprehensive understanding of the genomic program activated during early-phase of collateral vessel growth in a rat model for cerebral adaptive arteriogenesis (3-VO). While arteriogenesis constitutes a promising therapeutic concept for cerebrovascular ischemia, genomic profiles essential for therapeutic target identification were analysed solely for collateral arteries of the heart and periphery. Despite challenging anatomical conditions of the brain the 3-VO model allows identification of differentially expressed genes during adaptive cerebral arteriogenesis by selective removal of the posterior cerebral artery (PCA). Experiment Overall Design: Using an established rat model of nonischemic cerebral hypoperfusion (3-VO) (Busch, Buschmann; 2003), RNA was extracted from isolated ipsilateral PCA. Pooled RNAs from groups of intact (0h), sham and 3-VO animals 24h and 3 days after surgery, were hybridised repeatedly for an extensive genome screen of 15866 genes applying standardized Affymetrix technology. For each Array total RNA from 8 animals was processed, pooled and hybridized to a Rat230A GeneChip per group. These groups were classified as follows: intact control (N=3), 24h3VO (N=3); 24h sham (N=3), 3days3VO (N=3); and 3days sham(N=3). Hybridization, washing, antibody amplification, staining, and scanning of probe arrays were performed according to the Affymetrix Technical Manual. Probe arrays were scanned using the GeneChip System (Hewlett-Packard, Santa Cruz, CA)(Affymetrix) and raw data were processed using GCOS and normalized to a global intensity of 500.
Project description:Ischemic stroke triggers severe focal hypoperfusion accompanied with deprivation of oxygen and glucose to the cerebral tissue, together with loss of ATP, depolorization of neurons, elevated extracellular potassium concentration, and subsequently leads to excitotoxicity as well as increased oxidative stress promoting microvascular injury, blood-brain-barrier deregulation, post-ischemic inflammation and eventually the consequential neurological deficit. Although reperfusion of ischemic brain tissue is critical for restoring normal function, it can paradoxically result in secondary damage, called ischemia/reperfusion (I/R) injury. Microarray analysis was performed on the right striatum and cortex (corresponded to infarct area) of post-I/R injured brain tissues of wild-type (WT-MCAO) using Illumina mouse Ref8 V2 genechips. Suture-induced middle cerebral artery occlusion was induced for 2h followed by reperfusion, with tissue extraction taking place 2h, 8h and 24h post-reperfusion (n=4 respectively). Sham controls were included in this study too (n=4 respectively).
Project description:Glutathione peroxidase (Gpx) is a selenium-containing enzyme that catalyses the reduction of a variety of biological peroxides at the expense of reduced glutathione (GSH). Gpx1 is the most abundant isoform and its role has been implicated in neurodegenerative disorders such as ParkinsonM-bM-^@M-^Ys disease (PD), dementia with Lewy bodies tissue (DLB) (Power and Blumbergs, 2009) and traumatic brain injury (Tsuru-Aoyagi et al., 2009). Due to its high abundance, mutation of the Gpx1 allele would lower overall Gpx activity in the brain significantly. Gpx1 knockout (Gpx1-/-) mice do not show overt phenotypic differences, but all indications suggest that these mice are in a chronic M-bM-^@M-^\pro-oxidantM-bM-^@M-^] state (Cheng et al. 1999; de Haan et al. 2004). Indeed, a recent study from our laboratory illustrated that the absence of Gpx1 exacerbated stroke injury via increased ROS production and vascular permeability (Wong et al. 2008). Furthermore, Gpx1-/- mice demonstrated an increase in caspase-3 activation and greater infarct volume (Crack et al. 2001) Microarray analysis was performed on the right striatum and cortex(corresponded to infarct area) of post-I/R injured brain tissues of Gpx1 -/- brains using Illumina mouse Ref8 V2 genechips. Suture-induced middle cerebral artery occlusion was induced for 2h followed by reperfusion, with tissue extraction taking place 2h, 8h and 24h post-reperfusion (n=4 respectively). Sham controls were included in this study too (n=4 respectively).
Project description:This SuperSeries is composed of the following subset Series: GSE23160: Global transcriptomic profiling of ischemic/reperfusion injury in an in vivo wild-type mouse model. GSE23162: Global transcriptomic profiling of ischemic/reperfusion injury in an in vivo Gpx1 -/- transgenic mouse model. Refer to individual Series
Project description:Xuesaitong injection (XST), a standardized patent Chinese medicine of Panax notoginseng roots (Sanqi in Chinese), has long been used for effective prevention and treatment of stroke in China. However, the mechanisms underlying its effects against ischemic stroke are still poorly understood. In this study, we focused on the polypharmacology of XST against ischemic stroke with a XST-regulated stroke network analysis. Male Sprague Dawley rat model of MCAO and reperfusion were administered XST for 7 days while the control group was not treated. Three conditions were compared with three replicates each. These are: (1) sham; (2) model (3) XST treatment. Whole genome microarray analysis was performed using Affymetrix rat Genome 230 2.0 chips.