Resident Beijing Tiantan Hospital, Capital Medical University Beijing, Beijing, China (People's Republic)
Disclosure(s):
Qiheng He, MD, PhD: No financial relationships to disclose
Introduction: Moyamoya disease (MMD) is an uncommon cerebrovascular disease. Specific radiologic features, such as Suzuki stage, bilateral, and PCA involvement, have been validated to affect treatment outcomes. Until recently, molecular biomarkers and corresponding molecular mechanisms related to radiologic features and treatment outcomes remain unknown.
Methods: RNA N6-methyladenosine (m6A) Methyltransferase METTL14 was identified as a differentially expressed gene in MMD patients by analyzing the transcriptome sequencing (RNA-seq) data of middle cerebral arteries. Tube formation and wound healing assays were performed to investigate the effect of METTL14 on angiogenesis, and western blot assay was performed to study its effect on endothelial-mesenchymal transition (EndMT). In addition, Methylated RNA Immunoprecipitation Sequencing technology (MeRIP-seq), RIP, and luciferase reporter assays were performed to screen downstream targets of METTL14 in endothelial cells and to fully clarify the specific underlying molecular mechanisms affecting the phenotype of MMD.
Results: The expression level of METTL14 was significantly reduced in the middle cerebral arteries of MMD patients, and a reduced level of METTL14 predicted higher level of Suzuki stage, bilateral, and PCA involvement. Functionally, the knockdown of METTL14 significantly affected angiogenesis and EndMT of the human endothelial cells. Mechanistically, the total level of m6A was reduced in MMD patients. Downregulation of METTL14 reduced the m6A modification on ITGB4 3’UTR and subsequently reduced ITGB4 mRNA degradation, leading to its increased expression. Moreover, the METTL14-mediated inhibition of ITGB4 expression was dependent on the YTHDC1, which acted as an m6A reader to bind to ITGB4 mRNA and stabilize its expression. In addition, we demonstrated that the overexpression of ITGB4 promoted endothelial cell migration, and proliferation as well as stimulating the EndMT process, thereby continuously activating the MAPK signaling pathway. In addition, we demonstrated that blocking the MPAK pathway could restore the phenotype of METTL14-deficient endothelial cells.
Conclusion : Our study reveals that the downregulation of METTL14 promotes ITGB4 expression via m6A modification to affect angiogenesis and EndMT in human endothelial cells, suggesting the METTL14/MAPK/ITGB4 axis as a potential prognostic biomarker and therapeutic target for MMD.
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