Organoid Modeling and Single-Cell Profiling Reveal Smooth Muscle Cell Migration in Moyamoya Disease.

Moyamoya disease (MMD) is a cerebrovascular disorder characterized by progressive intracranial arterial stenosis, which can lead to ischemic or hemorrhagic stroke. However, its pathogenesis remains poorly understood, limiting advances in the diagnosis and treatment of MMD. We perform data-independent acquisition (DIA) proteomics on serum samples from 40 MMD patients and 20 healthy controls, followed by ELISA validation of an additional 45 cohort. Vascular organoids are generated from induced pluripotent stem cells (iPSCs) derived from the peripheral blood, and analyzed using histological staining, transcriptomics, and single-cell RNA sequencing (scRNA-seq). Histological examination of the temporal superficial artery (STA) in MMD patients reveals abnormal accumulation of vascular smooth muscle cells (VSMCs). DIA proteomics identify significant upregulation of TUBA4A and TUBB4B in MMD serum. In vitro assays demonstrate that these tubulin proteins promoted VSMC proliferation, migration, and contractile-to-synthetic phenotypic switching through the GJA1/PI3K/AKT/KLF4 signaling pathway. Single-cell RNA sequencing of MMD vascular organoids shows an increased proportion of synthetic VSMCs with upregulated TUBA4A and TUBB4B expression. Our findings suggest that overexpression of TUBA4A and TUBB4B contributes to pathological vascular remodeling in MMD through the GJA1/PI3K/AKT/KLF4 pathway, as well as therapeutic targets for intervention in MMD vascular remodeling.