MCAO rat model-guided identification of crucial miRNAs and stroke-related networks via high-throughput sequencing.

Abstract

Ischemic stroke is a complex polygenic disorder, in which microRNAs (miRNAs) have been implicated in various physiological and pathological processes. However, the specific regulatory networks and mechanisms involving brain miRNAs in the development of stroke remain inadequately elucidated. In this study, a middle cerebral artery occlusion (MCAO) model was established in Sprague-Dawley rats to simulate cerebral ischemia. Deep sequencing was performed to profile miRNA expression in the cerebral cortex after stroke, with an emphasis on identifying pivotal miRNAs. Key differentially expressed miRNAs (DEMis) and their potential target mRNAs were validated by quantitative reverse-transcription PCR (qRT-PCR), showing strong concordance with sequencing results. Cluster analysis revealed distinct miRNA expression patterns between MCAO and control cortical tissues. Gene Ontology (GO) enrichment analysis indicated that target genes of DEMis were significantly associated with stroke-relevant pathways, including calcium transmembrane transport, axon guidance, and MAPK and PI3K signaling pathways. Additionally, by combining DEMis target genes with differentially expressed genes (DEGs) obtained from high-throughput sequencing, we identified 376 disease-related target genes and constructed a miRNA-mRNA regulatory network of key DEGs. Through this analysis, we discovered three novel miRNAs (novel-miR-398, novel-miR-544, and novel-miR-1808) and ten miRNAs previously reported in stroke or other diseases. The target genes of these miRNAs are involved in post-stroke processes such as oxidative stress, apoptosis, inflammatory response, and nerve regeneration through endogenous competition mechanisms. Our findings suggested that miRNAs significantly contributed to the regulation of post-stroke pathophysiological processes, offering potential new targets for therapeutic intervention and advancing our understanding of the molecular mechanisms underlying stroke.