"Smooth muscle cell-specific deletion of Col15a1 unexpectedly leads to impaired development of advanced atherosclerotic lesions"

Atherosclerotic plaque rupture or erosion with subsequent embolic events is a major cause of sudden death from myocardial infarction or stroke (ref. 1–6). Plaque stability is classically characterized by a fibrous cap rich in ACTA2+ presumed smooth muscle cells (SMC) and collagen with a paucity of CD68+ presumed macrophages (ref. 7–10). The dogma in the field is that SMC produce collagens that in turn provide tensile strength and stability to atherosclerotic plaques. This has largely been supported by evidence that SMC can produce collagens in vitro (ref. 11–13). However, there is no direct evidence in vivo that SMC produce collagens as: 1) collagens are secreted molecules making it challenging to identify their cell source in vivo; and 2) recent lineage tracing studies by our lab (ref.14–16) and others (ref. 17–20) have shown that marker genes used to identify cell type within lesions are non-specific. For example, SMC marker genes (e.g. ACTA2) can be expressed by myeloid-derived cells and endothelial cells and SMC can express marker genes of macrophages (e.g. CD68 and LGALS3) (ref. 14–20). Therefore major unanswered questions remained in the field as to whether SMC are the primary source of collagens in vivo and if SMC produced collagens in turn impact plaque development.
We are interested in type XV collagen alpha 1 (COL15A1) as we identified a single nucleotide polymorphism (SNP) within human COL15A1 associated with age-related atherosclerosis disease risk (ref. 21). COL15A1 is a non-fibrillar collagen upregulated in human and mouse atherosclerosis that has been shown to link large collagen fibers to confer extracellular matrix (ECM) and tissue stability (ref. 21–26). Moreover, siRNA mediated knockdown of Col15a1 in cultured human aortic SMC results in an increase in SMC migration and a decrease in SMC proliferation (ref. 21). Taken together, we hypothesized that SMC produced COL15A1 would be critical in late stage atherosclerotic lesion stability through promoting organization of the extracellular and collagen matrix in the lesion and fibrous cap.
We demonstrate that SMC specific lineage tracing (YFP+), SMC specific Col15a1 knockout mice have impaired lesion development as compared to wild type controls and fail to form advanced lesions despite 18 weeks of Western diet feeding. SMC Col15a1 knockout lesions exhibit a drastic reduction in lesion size, overall cell number, and complexity as exhibited by an increased proportion of macrophages (YFP–LGALS3+/DAPI+) and decreased proportion of SMC per total cells (YFP+/DAPI+) and proliferating SMC (YFP+Ki67+/DAPI+) as compared to wild-type control lesions. Loss of SMC produced COL15A1 also led to a reduction in lesion and medial collagen content and organization which likely contributed to an increase in carotid artery passive tone. In vivo RNA-seq analysis on SMC Col15a1 knockout and wild type control lesions suggest one mechanism of action to explain these effects is likely through SMC-COL15A1 mediated inhibition of pro-atherogenic inflammatory pathways (ex. lipopolysaccharides, TNF, TGFβ, etc.) involved in lesion development. Combined, these results provide the first direct evidence that a SMC-derived collagen, COL15A1, is critical in lesion development.