Angiogenic and inflammatory responses in human induced microglia-like cells from patients with moyamoya disease: A reverse-translational study

Background: Moyamoya disease (MMD) is a heterogeneous group of cerebrovascular disorders characterized by regional stenosis and occlusion in the circle of Willis. The RNF213 gene mutation was identified recently; however, the pathogenesis of MMD is still unknown. Angiogenic factors associated with MMD are overexpressed in M2 polarized microglia in ischemic stroke, suggesting that microglia may be involved in the pathophysiology of MMD; however, existing approaches are not applicable to explore this hypothesis. We have recently developed a technique to produce induced microglial-like (iMG) cells from human blood monocytes, which are surrogate cells for human brain microglia. We herein investigate gene expression profiles in iMG cells derived from patients with MMD.Methods: We recruited 25 adult patients with MMD and 24 healthy volunteers. Their RNF213 genotypes were confirmed by Sanger sequencing. To stratify by clinical condition, patients with MMD were subdivided into progressive (N = 7) and stable (N = 18) groups based on observation of novel symptoms or radiographic advancement of Suzuki stage within 1 year. We induced 3 types of iMG cells: resting, M1-, and M2-induced cells; performed RNA sequencing followed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis, and qPCR assays.Results: The heterozygous missense variant (p.R4810K) of RNF213 was detected in 19 of 25 (76%) patients with MMD but not in the control group. RNA sequencing of M2-induced iMG cells revealed that 600 genes were significantly upregulated (338) or downregulated (262) in patients with MMD. Inflammation and immune-related factors and angiogenesis-related factors were specifically associated with MMD in GO analysis. qPCR for MMP9, VEGFA, and TGFB1 expression validated these findings.Conclusion: This study is the first to demonstrate that M2 microglia are involved in the angiogenic process of MMD. Our iMG technique provides a promising approach to explore the bioactivity of microglia in cerebrovascular diseases.