Cell-dependent contributions of thrombospondin-1 to the rupture of abdominal aortic aneurysm in mice

Rationale: Abdominal aortic aneurysms (AAA) rupture is a life-threatening event with unclear molecular mechanisms. Our previous work demonstrated elevated levels of the matricellular protein thrombospondin-1 (TSP1, encoded by Thbs1) in human and mouse AAA tissues. Single-cell RNA sequencing analysis identified macrophages, endothelial cells, and smooth muscle cells as the major TSP1-expressing cells in aneurysmal tissues. Global Thbs1 deletion reduces aneurysm formation by inhibiting vascular inflammation. Aims: To investigate how TSP1 deficiency in different cell types affects AAA rupture. Methods and Results: AAA and rupture were induced by angiotensin II infusion in hypercholesterolemic mice. In global Thbs1 deficient mice, hypercholesterolemia was achieved by crossing them with Apoe knockout mice. To generate cell type–specific TSP1 deficient mice, Thbs1 flox/flox mice were crossed with VE-cadherin-Cre, SMMHC-iCreERT2, and Lyz2-Cre mice to target endothelial cells, smooth muscle cells, and myeloid cells, respectively. In these conditional knockout models, hypercholesterolemia was induced via AAV-PCSK9. We found that both global and myeloid-specific Thbs1 deletion increased rupture rate over twofold, whereas endothelial- or smooth muscle cell–specific deletion had no significant effect. Endothelial-specific Thbs1 deletion reduced aneurysm size in the CaCl2 model. Single-cell RNA sequencing and histology in myeloid-specific Thbs1 knockout aortas revealed broad suppression of inflammation and extracellular matrix production. Conclusions: Myeloid-derived TSP1 plays a critical role in inhibiting aneurysm rupture in mice, likely by promoting matrix repair phenotypes in vascular smooth muscle cells, enhancing vascular wall integrity. Overall design: Male Thbs1fl/fl/Lyz2-Cre and Thbs1wt/wt/Lyz2-Cre mice (8-12 weeks of age) were subjected to the PCSK9-AAV/Angiotensin II model of AAA. Abdominal aortas were collected after 7 days of Angiotensin II infusion and digested into a single-cell suspension. For each genotype, cells from five mice were pooled into one sample and subjected to scRNA-seq.