Identification of coagulation-related hub genes in ischemic stroke based on bioinformatics integration analysis and investigation of their immune regulatory mechanisms.
Ischemic stroke (IS) is a cerebrovascular disease resulting from insufficient blood supply to specific areas of the brain, often due to atherosclerosis and thrombosis. While the association between polymorphisms in coagulation-related genes (CRGs) and thrombosis has been suggested, the precise relationship between CRGs and IS remains unclear and requires further exploration.
This study utilized IS-related datasets (GSE16561, GSE9877, GSE58294, GSE224273) and CRGs to identify differentially expressed genes (IS-DEGs) through differential expression analysis. Differentially expressed coagulation-related genes (DE-CRGs) were determined by overlapping CRGs and IS-DEGs. Machine learning models, including random forest, gradient boosting decision trees, and support vector machines, were applied to screen for potential hub genes. These candidate hub genes were then validated using an independent dataset (GSE9877) and confirmed by real-time quantitative PCR (RT-qPCR) in clinical samples from IS patients. Single-cell RNA-seq of GSE224273 was processed via Seurat to map hub-gene expression across endothelial, smooth-muscle, macrophage, monocyte, T, NK, and B-cell clusters.
A total of 18 DE-CRGs were acquired by overlapping 450 IS-DEGs and 203 CRGs. Furthermore, 517 cluster-differentially expressed genes (DEGs) and 1,021 coagulation-score-RGs were obtained based on 18 DE-CRGs. The 29 candidate genes were obtained by overlapping IS-DEGs, cluster-DEGs and coagulation-score-related genes (RGs), five hub genes (REPS2, LIME1, MYH9, C5AR1, ECHDC2) were successfully screened. Subsequently, in GSE9877 dataset, only REPS2, MYH9, C5AR1 had the same expression tendency with that in GSE16561 dataset, while our RT-qPCR results suggested that the expression trend of LIME1, MYH9, C5AR1, ECHDC2 were consistent with that in GSE16561 dataset besides REPS2. Single-cell analysis localized MYH9 predominantly to smooth-muscle and endothelial cells, whereas C5AR1 was enriched in macrophages and monocytes, with both genes significantly upregulated in IS-derived cells. Two hub genes, C5AR1 and MYH9, were eventually selected from this group. Further analyses showed that these genes are significantly enriched in pathways related to complement and coagulation cascades and platelet activation.
The two hub genes, C5AR1 and MYH9, may contribute to the development of IS by promoting thrombosis. These findings offer new insights into the molecular mechanisms of IS and suggest potential biomarkers and therapeutic targets for IS management.
This study utilized IS-related datasets (GSE16561, GSE9877, GSE58294, GSE224273) and CRGs to identify differentially expressed genes (IS-DEGs) through differential expression analysis. Differentially expressed coagulation-related genes (DE-CRGs) were determined by overlapping CRGs and IS-DEGs. Machine learning models, including random forest, gradient boosting decision trees, and support vector machines, were applied to screen for potential hub genes. These candidate hub genes were then validated using an independent dataset (GSE9877) and confirmed by real-time quantitative PCR (RT-qPCR) in clinical samples from IS patients. Single-cell RNA-seq of GSE224273 was processed via Seurat to map hub-gene expression across endothelial, smooth-muscle, macrophage, monocyte, T, NK, and B-cell clusters.
A total of 18 DE-CRGs were acquired by overlapping 450 IS-DEGs and 203 CRGs. Furthermore, 517 cluster-differentially expressed genes (DEGs) and 1,021 coagulation-score-RGs were obtained based on 18 DE-CRGs. The 29 candidate genes were obtained by overlapping IS-DEGs, cluster-DEGs and coagulation-score-related genes (RGs), five hub genes (REPS2, LIME1, MYH9, C5AR1, ECHDC2) were successfully screened. Subsequently, in GSE9877 dataset, only REPS2, MYH9, C5AR1 had the same expression tendency with that in GSE16561 dataset, while our RT-qPCR results suggested that the expression trend of LIME1, MYH9, C5AR1, ECHDC2 were consistent with that in GSE16561 dataset besides REPS2. Single-cell analysis localized MYH9 predominantly to smooth-muscle and endothelial cells, whereas C5AR1 was enriched in macrophages and monocytes, with both genes significantly upregulated in IS-derived cells. Two hub genes, C5AR1 and MYH9, were eventually selected from this group. Further analyses showed that these genes are significantly enriched in pathways related to complement and coagulation cascades and platelet activation.
The two hub genes, C5AR1 and MYH9, may contribute to the development of IS by promoting thrombosis. These findings offer new insights into the molecular mechanisms of IS and suggest potential biomarkers and therapeutic targets for IS management.