RNA Sequencing for Gene Expression Profiles in a Rat Model of Middle Cerebral Artery Occlusion

Abstract

Gene expression regulatory mechanisms in models of middle cerebral artery occlusion (MCAO) have been assessed in some studies, but questions remain. The discovery of differentially expressed genes (DEGs) between MCAO and control rats analyzed by next-generation RNA sequencing is of particular interest. These DEGs may help guide the clinical diagnosis of stroke and facilitate selection of the optimal treatment strategy. Twenty rats were equally divided into control and MCAO groups. Three rats from each group were randomly selected for RNA sequencing analysis. Sequence reads were obtained from an Illumina HiSeq2500 platform and mapped onto the rat reference genome RN6 using Hisat2. We identified 1,007 significantly DEGs with p<0.05, including 785 upregulated (fold change [FC]>2) and 222 downregulated (FC<0.5) DEGs, in brain tissue from MCAO rats compared with that from control rats, and numerous immune response genes were identified. Gene ontology (GO) analysis revealed that the majority of the most enriched and meaningful biological process terms were mainly involved in immune response, inflammatory response, cell activation, leukocyte migration, cell adhesion, response to external stimulus, cell migration, and response to wounding. Also enriched were immune-related pathways and neural-related pathways. Similar to GO molecular function terms, the enriched terms were mainly related to cytokine receptor activity. Six DEGs were verified by quantitative real-time polymerase chain reaction (qRT-PCR). Protein-protein interaction analysis of these hits revealed that MCAO affects complement and coagulation cascades and chemokine signaling pathway. Our study identified novel biomarkers that could help further elucidate MCAO mechanisms and processes.

Integrated Bioinformatics Analysis of Potential mRNA and miRNA Regulatory Networks in Mice With Ischemic Stroke Treated by Electroacupuncture

“…Among these genes, Mki67, Top2a, and Cenpf, which are cellular markers for proliferation, were increased in the MCAO model and decreased by EA treatment. This was consistent with previous RNA-seq reports of ischemic stroke (40,41). Previous studies also reported that following ischemic injury, the number of reactive astrocytes increased, and subsequently, microglia were activated since oxidative radical imbalance occurred (41).…”

Identification and functional analysis of microRNAs in rats following focal cerebral ischemia injury

“…Rats were randomly divided into the control (sham surgery) and MCAO groups (n=10 in each group). A focal cerebral ischemia rat model induced by MCAO was established as previously described (19,20). All surgical procedures were performed under anesthesia to minimize animal suffering, according to the National Institutes of Health (NIH) Guide for Animal Care and Welfare (NIH publication no.…”

Decoding the transcriptional response to ischemic stroke in young and aged mouse brain

“…There is a need to understand the age-related mechanisms of ischemic injury to improve our ability to discern which therapeutics can be translated from the bench to the bedside 2 . Previous global gene expression studies of experimental stroke using microarrays [27][28][29][30][31][32][33][34] and more recently RNA-Seq [35][36][37] have provided useful insights into the pathophysiology of ischemic stroke and uncovered many altered molecular pathways 38 . However, few studies included aged animals.…”