Bioinformatics analysis and in vivo validation of ferroptosis-related genes in ischemic stroke

Chang Liu; Zhixi Li; Hongjie Xi

doi:10.3389/fphar.2022.940260

Bioinformatics analysis and in vivo validation of ferroptosis-related genes in ischemic stroke

Front Pharmacol. 2022 Nov 24:13:940260. doi: 10.3389/fphar.2022.940260. eCollection 2022.

Authors

Chang Liu^{1

2

3}, Zhixi Li^{1

2

3}, Hongjie Xi^{1

3}

Affiliations

¹ Heilongjiang Province Key Laboratory of Research on Anesthesiology and Critical Care Medicine, Harbin, China.
² The Key Laboratory of Myocardial Ischemia Organization, Chinese Ministry of Education, Harbin, China.
³ Department of Anesthesiology, The Second Affiliated Hospital of Harbin Medical University, Harbin, China.

Abstract

Ischemic stroke (IS) is a neurological condition associated with high mortality and disability rates. Although the molecular mechanisms underlying IS remain unclear, ferroptosis was shown to play an important role in its pathogenesis. Hence, we applied bioinformatics analysis to identify ferroptosis-related therapeutic targets in IS. IS-related microarray data from the GSE61616 dataset were downloaded from the Gene Expression Omnibus (GEO) database and intersected with the FerrDb database. In total, 33 differentially expressed genes (DEGs) were obtained and subjected to functional enrichment and protein-protein interaction (PPI) network analyses. Four candidate genes enriched in the HIF-1 signaling pathway (HMOX1, STAT3, CYBB, and TLR4) were selected based on the hierarchical clustering of the PPI dataset. We also downloaded the IR-related GSE35338 dataset and GSE58294 dataset from the GEO database to verify the expression levels of these four genes. ROC monofactor analysis demonstrated a good performance of HMOX1, STAT3, CYBB, and TLR4 in the diagnosis of ischemic stroke. Transcriptional levels of the above four genes, and translational level of GPX4, the central regulator of ferroptosis, were verified in a mouse model of middle cerebral artery occlusion (MCAO)-induced IS by qRT-PCR and western blotting. Considering the regulation of the HIF-1 signaling pathway, dexmedetomidine was applied to the MCAO mice. We found that expression of these four genes and GPX4 in MCAO mice were significantly reduced, while dexmedetomidine reversed these changes. In addition, dexmedetomidine significantly reduced MCAO-induced cell death, improved neurobehavioral deficits, and reduced the serum and brain levels of inflammatory factors (TNF-α and IL-6) and oxidative stress mediators (MDA and GSSG). Further, we constructed an mRNA-miRNA-lncRNA network based on the four candidate genes and predicted possible transcription factors. In conclusion, we identified four ferroptosis-related candidate genes in IS and proposed, for the first time, a possible mechanism for dexmedetomidine-mediated inhibition of ferroptosis during IS. These findings may help design novel therapeutic strategies for the treatment of IS.

Keywords: HIF-1 signaling pathway; dexmedetomidine; ferroptosis; inflammatory response; ischemic stroke; microarray; oxidative stress.