Circulating miRNome profiling in Moyamoya disease-discordant monozygotic twins and endothelial microRNA expression analysis using iPS cell line

Abstract

Abstract Background Moyamoya disease (MMD) is characterized by progressive stenosis of intracranial arteries in the circle of Willis with unknown etiology even after the identification of a Moyamoya susceptible gene, RNF213. Recently, differences in epigenetic regulations have been investigated by a case-control study in MMD. Here, we employed a disease discordant monozygotic twin-based study design to unmask potential confounders. Methods Circulating genome-wide microRNA (miRNome) profiling was performed in MMD-discordant monozygotic twins, non-twin-MMD patients, and non-MMD healthy volunteers by microarray followed by qPCRvalidation, using blood samples. Differential plasma-microRNAs were further quantified in endothelial cells differentiated from iPS cell lines (iPSECs) derived from another independent non-twin cohort. Lastly, their target gene expression in the iPSECs was analyzed. Results Microarray detected 309 plasma-microRNAs in MMD-discordant monozygotic twins that were also detected in the non-twin cohort. Principal component analysis of the plasma-microRNA expression level demonstrated distinct 2 groups separated by MMD and healthy control in the twin- and non-twin cohorts. Of these, differential upregulations of hsa-miR-6722-3p/− 328-3p were validated in the plasma of MMD (absolute log2 expression fold change (logFC) > 0.26 for the twin cohort; absolute logFC > 0.26, p < 0.05, and q < 0.15 for the non-twin cohort). In MMD derived iPSECs, hsa-miR-6722-3p/− 328-3p showed a trend of up-regulation with a 3.0- or higher expression fold change. Bioinformatics analysis revealed that 41 target genes of miR-6722-3p/− 328-3p were significantly down-regulated in MMD derived iPSECs and were involved in STAT3, IGF-1-, and PTEN-signaling, suggesting a potential microRNA-gene expression interaction between circulating plasma and endothelial cells. Conclusions Our MMD-discordant monozygotic twin-based study confirmed a novel circulating microRNA signature in MMD as a potential diagnostic biomarker minimally confounded by genetic heterogeneity. The novel circulating microRNA signature can contribute for the future functional microRNA analysis to find new diagnostic and therapeutic target of MMD.

Keywords

• Circulating microRNA
• Moyamoya disease
• Discordant monozygotic twins
• iPS cells
• Endothelial cells