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Comprehensive Analysis of Blood-Based m6A Methylation in Human Ischemic Stroke

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Abstract

Alterations of N6-methyladenosine (m6A) methylation have been reported in the cerebral cortices of mouse and rat models of ischemic stroke (IS). However, the role of m6A methylation in human IS is still unknown. We assessed m6A levels in peripheral blood from patients with IS and healthy controls. A transient middle cerebral artery occlusion and reperfusion (tMCAO/R) mouse model, and an oxygen–glucose deprivation/reperfusion (OGD/R) model in A172 cells were established to further assess m6A levels. Methylated RNA immunoprecipitation sequencing (MeRIP-seq) and RNA sequencing were performed in the peripheral blood of patients with IS and healthy controls. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses were used to identify underlying biological processes. In this study, we found that global m6A levels were elevated in the peripheral blood of patients with IS, in the cerebral cortex of mice after tMCAO/R treatment and in A172 cells after OGD/R treatment. MeRIP-seq analysis identified 2115 altered m6A peaks in patients with IS, 1052 upregulated and 1063 downregulated. Downregulated methylated mRNAs were enriched in Hippo signaling pathway, cytokine-cytokine receptor interaction, NF-kappa B signaling pathway, etc. Upregulated methylated mRNAs were enriched in calcium signaling pathways, Hedgehog signaling pathway, MAPK signaling pathway, etc. Moreover, a total of 84 differentially expressed mRNAs with altered m6A peaks were identified and enriched in EGFR tyrosine kinase inhibitor, Hematopoietic cell lineage, and cytokine-cytokine receptor interactions. This study is the first to profile the transcriptome-wide m6A methylome of peripheral blood in human IS and uncover increased global m6A levels in the peripheral blood of patients with IS.

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Data Availability

The data in this study are available from the corresponding author upon reasonable request.

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Acknowledgements

The authors thank all of the physicians and nurses in the Department of Internal Neurology, First Affiliated Hospital, Guangxi University of Chinese Medicine.

Funding

This study was supported by the Natural Science Foundation of Guangxi (2018GXNSFAA281224), and the National Natural Science Foundation of China (Nos. 81874395 and 81860822, 82160849). The MEGASTROKE project received funding from sources specified at http://www.megastroke.org/acknowledgments.html. All MEGASTROKE authors were listed at https://www.megastroke.org/authors.html.

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  1. Lulu Zhu, Shengying Liu, and Fangping Liao contributed equally to this work.

Authors and Affiliations

  1. School of Public Health, Guangxi Medical University, 22 Shuangyong Road, Nanning, Guangxi, China

    Lulu Zhu, Shengying Liu, Fangping Liao, Jialei Yang, Tian Liang & Li Su

  2. Department of Internal Neurology, First Affiliated Hospital, Guangxi University of Chinese Medicine, 89-9 Dongge Road, Nanning, Guangxi, China

    Yibing Yang, Xianli Huang & Lian Gu

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  1. Lulu Zhu
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Contributions

All authors contributed to the study’s conception and design. Experiments were performed by Lulu Zhu, Shengying Liu, Fangping Liao, and Jialei Yang; Data collection and analysis were performed by Lulu Zhu, Tian Liang, Yibing Yang, Xianli Huang, Lian Gu, and Li Su. The first draft of the manuscript was written by Lulu Zhu, Lian Gu, and Li Su. All authors read and approved the final manuscript.

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Written informed consent was obtained from all subjects, and the research protocol was approved by Guangxi Medical University Medical Ethics Committee. All animal experiments were approved by the Animal Ethics Committee of Guangxi Medical University.

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Zhu, L., Liu, S., Liao, F. et al. Comprehensive Analysis of Blood-Based m6A Methylation in Human Ischemic Stroke. Mol Neurobiol 60, 431–446 (2023). https://doi.org/10.1007/s12035-022-03064-2

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