Abstract TP274: Spatial RNA Transcriptome Analysis of Moyamoya Disease Intracranial Arteries

Background: Moyamoya disease (MMD) is a rare progressive cerebrovascular disease that can lead to ischemic and hemorrhagic strokes. The pathology of MMD arteries includes thickening of the innermost layer (intima), leading to the narrowing of arteries that supply oxygen to the brain. However, the mechanism underlying this process is largely unknown. We investigated the transcriptome differences of arterial samples from MMD patients and other vascular diseases. Specifically we analyzed the principal cell types - endothelial cells (EC) and vascular smooth muscle cells (VSMC) in MMD superior temporal artery (STA) and middle cerebral artery (MCA). Methods: STA and/or MCA were collected during surgeries in MMD, aneurysm, or cavernous malformation patients. MCA and STA samples were sectioned into 10µm thick slices and RNA transcripts stained through Advance Cell Diagnostics’ multiplex protocol for endothelial cells (PECAM1), vascular smooth muscle cells (ACTA2), proliferation marker (ki67), extracellular matrix (ECM), and DAPI for cell nuclei. Sections were imaged with Zeiss laser scanning microscope (LSM800) and analyzed with ImageJ. For spatial transcriptomic analysis, samples were formalin fixed, paraffin embedded and then processed through the Visium HD pipeline. Results: RNA transcriptome detection through RNAscope noted the presence of ki67, PECAM, ECM, and ACTA2. This aligns with previous immunohistochemistry staining for endothelial cells (CD31) and vascular smooth muscle cells (α-SMA). In addition, the morphology of VSMCs in MMD STA differs significantly; control STAs display a contractile phenotype with elongated, spindle-shaped cells, whereas MMD STAs exhibit synthetic phenotype with shorter, cobblestone appearance. Quantification of the intima layer thickness also show increased intima thickness in MMD STA. Spatial transcriptome analysis of these vessels have been processed to identify gene expression differences in these vessel layers in MMD. Conclusions: Our findings found RNA evidence of proliferation in the media layer, predominantly VSMCs, indicating a shift towards synthetic phenotype associated with increased cellular proliferation and reduced contractility, which can contribute to the MMD pathology. Ongoing studies include analysis of Visium HD transcriptome changes across different layers in MMD, aneurysm, and cavernous malformation to identify key molecular changes driving these pathological MMD features.