Functional Role of Second Heart Field-derived Smooth Muscle Cells in Thoracic Aortopathies in Mice

Background: The ascending aorta is a common location for thoracic aortopathies. Pathology predominates in the aortic media with disease severity being most apparent in outer laminar layers. In the ascending aorta, smooth muscle cells (SMCs) are derived from two embryonic origins: cardiac neural crest and second heart field (SHF). SMCs of these origins have distinct distributions, and the localization of SHF coincides with the regional specificity in some forms of thoracic aortopathies. However, the role of SHF-derived SMCs in maintaining the structural and functional integrity of the ascending aorta remains unclear. Methods: Mass spectrometry assisted proteomic and single cell transcriptomic analyses were performed in mouse aortas to discriminate molecular features of SHF-derived SMCs in maintaining the aortic homeostasis. Genetic deletion of low-density lipoprotein receptor-related protein 1 (Lrp1) or transforming growth factor-β receptor 2 (Tgfbr2) in SHF-derived SMCs was conducted to examine impact of SHF-derived SMCs on the development of thoracic aortopathies. Results: Proteomic analysis did not detect differences in protein profiles between ascending (disease prone) and descending (disease resistant) thoracic aortas in saline-infused mice. However, angiotensin II infusion altered these profiles in a region-specific manner. Angiotensin II evoked differential expression of multiple LRP1 ligands. Histological analysis demonstrated that angiotensin II-induced medial disruptions were detected mainly in outer laminar layers derived from the SHF. Single cell RNA sequencing using normal mouse aortas revealed lower abundance of elastin mRNA in SHF-derived SMCs compared to SMCs from the cardiac neural crest. In addition, Lrp1 and Tgfbr2 mRNA were abundant in SHF-derived SMCs. To examine biological effects of SHF-derived cells, Lrp1 or Tgfbr2 was deleted in SHF-derived cells in mice. SHF-specific Lrp1 deletion augmented angiotensin II-induced aortic aneurysm and rupture in the ascending region. Proteomic analysis discerned regulation of protein abundances related to TGF-β signaling pathways by Lrp1 deletion in SHF-derived cells. Deletion of Tgfbr2, a key regulator of TGF-β signaling, in SHF-derived cells led to embryonic lethality at E12.5 with dilatation of the outflow tract and retroperitoneal hemorrhage in mice. Conclusion: These results demonstrate that SMCs derived from the SHF play a critical role in the integrity of the ascending aortic wall.