NINJ1 hypomethylation enhances endothelial dysfunction in brain arteriovenous malformations.
Brain 2026 May 26. [Online ahead of print]

Abstract

Brain arteriovenous malformations (bAVMs) are a major cause of intracranial haemorrhage in children and young adults, yet the molecular basis of endothelial dysfunction and vascular fragility remains incompletely understood. Although somatic activation of the KRAS-MAPK pathway has been implicated in bAVM pathogenesis, the contribution of epigenetic dysregulation has not been systematically defined. We performed genome-wide DNA methylation profiling using the Illumina Infinium MethylationEPIC v2 (935K) array on human bAVM nidus tissues, followed by targeted pyrosequencing and immunofluorescence validation. Functional and mechanistic studies were conducted in human brain microvascular endothelial cells, complemented by transcriptomic analyses and in vivo assessment in an endothelial-specific zebrafish model. Genome-wide methylation analysis revealed a distinct promoter-centric epigenetic landscape in bAVM tissue, with enrichment of genes involved in cell adhesion, angiogenesis and inflammatory signalling. Among these, Nerve Injury-Induced Protein 1 (NINJ1) emerged as a prominently hypomethylated gene with increased endothelial expression in bAVM lesions. In human brain microvascular endothelial cells, NINJ1 overexpression enhanced proliferation, migration and tube formation, whereas NINJ1 knockdown produced opposite effects. DNMT1 silencing upregulated NINJ1 expression and induced angiogenic behaviour, which was abolished by concurrent NINJ1 knockdown. Mechanistically, NINJ1 enhanced KRAS-MAPK/ERK signalling, at least in part by increasing KRAS protein stability. NINJ1 overexpression also increased endothelial permeability and reduced VE-cadherin and ZO-1 expression. In vivo, endothelial-specific overexpression of ninj1 in zebrafish impaired vascular integrity and induced intracranial haemorrhage. These findings identify promoter hypomethylation-associated activation of NINJ1 as a previously unrecognised epigenetic contributor to endothelial dysfunction in bAVM and support a role for NINJ1 as an upstream enhancer of KRAS-MAPK signalling. Epigenetic dysregulation of NINJ1 may therefore contribute to vascular instability and haemorrhage risk in bAVM.

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Chen CJDepartment of Neurosurgery, Huashan Hospital, Fudan University, Shanghai 200040, China. National Center for Neurological Disorders, Shanghai, 200040, China. Shanghai Key Laboratory of Brain Function and Restoration and Neural Regeneration, Shanghai, 200040, China. Neurosurgical Institute of Fudan University, Shanghai, 200040, China.
Zhang HDepartment of Neurosurgery, Huashan Hospital, Fudan University, Shanghai 200040, China. National Center for Neurological Disorders, Shanghai, 200040, China. Shanghai Key Laboratory of Brain Function and Restoration and Neural Regeneration, Shanghai, 200040, China. Neurosurgical Institute of Fudan University, Shanghai, 200040, China.
Pan TDepartment of Neurosurgery, Huashan Hospital, Fudan University, Shanghai 200040, China. National Center for Neurological Disorders, Shanghai, 200040, China. Shanghai Key Laboratory of Brain Function and Restoration and Neural Regeneration, Shanghai, 200040, China. Neurosurgical Institute of Fudan University, Shanghai, 200040, China.
Song KDepartment of Neurosurgery, Huashan Hospital, Fudan University, Shanghai 200040, China. National Center for Neurological Disorders, Shanghai, 200040, China. Shanghai Key Laboratory of Brain Function and Restoration and Neural Regeneration, Shanghai, 200040, China. Neurosurgical Institute of Fudan University, Shanghai, 200040, China.
Pan JJSchool of Biomedical Engineering, ShanghaiTech University, Shanghai 201210, China.
Liu PDepartment of Neurosurgery, Huashan Hospital, Fudan University, Shanghai 200040, China. National Center for Neurological Disorders, Shanghai, 200040, China. Shanghai Key Laboratory of Brain Function and Restoration and Neural Regeneration, Shanghai, 200040, China. Neurosurgical Institute of Fudan University, Shanghai, 200040, China.
Shi YDepartment of Neurosurgery, Huashan Hospital, Fudan University, Shanghai 200040, China. National Center for Neurological Disorders, Shanghai, 200040, China. Shanghai Key Laboratory of Brain Function and Restoration and Neural Regeneration, Shanghai, 200040, China. Neurosurgical Institute of Fudan University, Shanghai, 200040, China.
Li ZDepartment of Neurosurgery, Huashan Hospital, Fudan University, Shanghai 200040, China. National Center for Neurological Disorders, Shanghai, 200040, China. Shanghai Key Laboratory of Brain Function and Restoration and Neural Regeneration, Shanghai, 200040, China. Neurosurgical Institute of Fudan University, Shanghai, 200040, China.
Hu LDepartment of Neurosurgery, Huashan Hospital, Fudan University, Shanghai 200040, China. National Center for Neurological Disorders, Shanghai, 200040, China. Shanghai Key Laboratory of Brain Function and Restoration and Neural Regeneration, Shanghai, 200040, China. Neurosurgical Institute of Fudan University, Shanghai, 200040, China.
Qiu TDepartment of Neurosurgery, Huashan Hospital, Fudan University, Shanghai 200040, China. National Center for Neurological Disorders, Shanghai, 200040, China. Shanghai Key Laboratory of Brain Function and Restoration and Neural Regeneration, Shanghai, 200040, China. Neurosurgical Institute of Fudan University, Shanghai, 200040, China.
Quan KDepartment of Neurosurgery, Huashan Hospital, Fudan University, Shanghai 200040, China. National Center for Neurological Disorders, Shanghai, 200040, China. Shanghai Key Laboratory of Brain Function and Restoration and Neural Regeneration, Shanghai, 200040, China. Neurosurgical Institute of Fudan University, Shanghai, 200040, China.
Zhu W0000-0002-6483-2434Department of Neurosurgery, Huashan Hospital, Fudan University, Shanghai 200040, China. National Center for Neurological Disorders, Shanghai, 200040, China. Shanghai Key Laboratory of Brain Function and Restoration and Neural Regeneration, Shanghai, 200040, China. Neurosurgical Institute of Fudan University, Shanghai, 200040, China.

Pub Type(s)

Journal Article

Language

eng

PubMed ID

42189175