Cerebrovascular Section Best Basic Scientific Paper Abstract Award: SMC remodeling implicated in IA pathophysiology via familial WES-identified genes

Introduction: The pathophysiology of intracranial aneurysms (IAs) is complex and poorly understood. Systemic vascular studies have identified that key characteristics of smooth muscle cells (SMCs) – particularly plasticity and remodeling in response to physiologic and pathologic stimuli – can result in morphologic and functional changes. While it is thought that these shifts can cause downstream changes in vascular tone and structure, as noted in atherosclerosis and hypertensive processes, their role in IA formation has not been investigated fully.

Methods: To start, we explored whether a gene we had previously implicated in IA formation, PPIL4, acts via SMC remodeling, we proceeded with a stepwise approach. We bulk RNA sequenced (RNA-seq) and immunostained human primary brain vascular smooth muscle cells (HPBVSMC) treated with lentiviral shRNA and open-reading-frame (ORF) particles to individually downregulate and overexpress PPIL4, respectively.

Results: Bulk RNA-seq revealed that there is a significant upregulation of SMC related encoding genes in PPIL4 knockdown. As such, with PPIL4 downregulation we saw a significant upregulation of SMC-related protein-encoding genes. These included CSRP1, CSRP2, ICAM1, S100A4, ACTA2, CNN1, TNNT1, and TNNT2. Importantly, these genes directly involved in SMC phenotype regulation, as well as inflammation. Gene enrichment analysis revealed that the implicated genes are significantly enriched in pathways involved in collagen fibril organization and bone morphogenesis. We were able to reverse the expression of these genes once we overexpressed PPIL4. Finally, we performed SLAM-seq across both PPIL4 over-expression and controls to elucidate whether PPIL4 is involved in RNA modulation and found a global change in synthesis and degradation kinetics.

Conclusion : We confirmed our hypothesis that PPIl4 plays a role in modulating SMC phenotype. We previously demonstrated the association between PPIL4 and IA pathology, and these findings further suggest that there is a mechanistic correlation between IA formation and SMC remodeling via PPIL4. This work provides the foundation for future familial WES-identified gene studies to further elucidate the underlying mechanisms of IA formation.