Microregional RNA sequencing reveals layer-specific smooth muscle cell heterogeneity in human bicuspid aortic valve-associated aortic dilatation.

Objective: Smooth muscle cells in the human ascending aorta exhibit layer-specific heterogeneity; however, their molecular profiles and contribution to aortic diseases remain poorly understood. In this study, we applied microregional RNA sequencing to characterize layer-specific transcriptomic signatures and investigate the differences between bicuspid aortic valve (BAV) and tricuspid aortic valve (TAV) aortopathy. Methods: Dilated ascending aortic wall samples were obtained from nine patients who underwent ascending aorta replacement concomitant with aortic valve replacement: five with BAV and four with TAV. Microdissected tissues were collected from four distinct layers in media using a micro-punching device. RNA was extracted and sequenced, and the transcriptomic differences across the layers and between BAV- and TAV-dilated aortas were analyzed. Results: Microregional RNA sequencing revealed significant upregulation of several extracellular matrix remodeling genes in the innermost layer, including Conclusions: The present study demonstrated the application of microregional RNA sequencing to human aortic tissue, revealing layer-specific transcriptomic differences in aortic diseases. The differences in gene-set-level findings indicate distinct biological contexts and mechanisms in BAV- and TAV-dilated aortas, particularly in the innermost media layer. This approach is useful for assessing layer-specific profiles in aortic diseases. Clinical relevance: This study identified layer-specific heterogeneity within the human aorta, with molecular alterations in the innermost medial layer potentially contributing to differences between TAV- and BAV-associated aortopathy. The findings suggest that layer-specific transcriptome profiles differ across disease types. Accordingly, diagnostic approaches and therapeutic strategies may be more effective when tailored to specific aortic regions and their underlying molecular characteristics.