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Single-cell transcriptome of the murine carotid artery following acute injury

ABSTRACT: Acute vascular injury is an unwelcome consequence of invasive treatments designed to alleviate symptoms of vascular stenosis. Resulting fibrotic scarring and neointima formation may result in loss of lumen diameter and diminished vascular function. Understanding of the key phases of acute inflammation, resolution and remodeling has the potential to minimise unwanted effects on the vasculature and therefore improve patient outcomes. The cellular landscape of blood vessels is highly hetergeneous in nature, and therefore data at single-cell resolution is of high relevance to this problem. Herein, the cellularity of murine carotid artery tissue is described in a cell- and time-resolved manner. Single-cell RNA-sequencing of carotid tissue isolated at time-points ranging from uninjured vessel to 14 days post-injury enabled the recapitulation of all stages of vascular injury. In these data, a sub-population of smooth muscle cells which also arises in atherosclerosis and myocardial infarction was identified. So-called stem cells/endothelial cells/monocytes (SEM) cells are candidates for repopulating injured vessels, and were amongst the most proliferative cell clusters following wire-injury of the carotid artery. Transcriptional signatures reflecting SEM gene expression patterns could also be detected in bulk RNA-sequencing of neointimal tissue isolated by laser capture microdissection. These data indicate that phenotypic plasticity of smooth muscle cells is highly important to the progression of lumen loss following acute vascular insult.

ORGANISM(S): Mus musculus

PROVIDER: GSE217801 | GEO | 2023/03/09

REPOSITORIES: GEO

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Project description:IRF9 is ubiquitously expressed and mediates the effects of IFNs, previous study showed that IRF9 played an important role in immunity and cell fate decision. Our recent study revealed that IRF9 involved in cardiac hypertrophy, hepatic steatosis and insulin resistance. However, the function of IRF9 in VSMC and neointima formation was largely unknown. We found that IRF9 expression was significantly increased in the VSMCs of mouse carotid artery. More importantly, we generated SMC-specific IRF9 overexpression transgenic mice (IRF9 TG) and found that IRF9 TG significantly increased VSMC proliferation, migration and neointima formation compared with NTG mice in response to injury. To evaluate the underlying mechanism by which IRF9 promotes VSMC proliferation and migration after vascular injury, IRF9 TG and NTG mice were subjected to wire-injury and the carotid arteries were collected at 14 days post-injury. We combined 3-5 vessels for one sample, and 3 samples for each phenotype. Subsequently, a total of 400ng RNA was used following Affymetrix instruction and 10 ug of cRNA were hybridized for 16 hr at 45°. GeneChips were scanned using the Scanner 7G and the data was analyzed with Expression Console using Affymetrix default analysis settings and global scaling as normalization method. RMA analysis was employed to evaluate the gene expression. We used microarrays to detect the global gene expression in the carotid arteries of smooth muscle cell specific IRF9 transgenic mice(IRF9 TG) compared with non transgenic control mice (NTG) at 14 days post-injury and identified distinct classes of altered genes. non-transgenic controls mice (NTG) and smooth muscle specific IRF9 transgenic mice (IRF9 TG) were subjected to wire-injury and the carotid ateries were collected at 14 days post-injury. We combine 3-5 vessels in one tube and for a single Affymetrix microarray. Total RNA was extracted and a total of 400ng RNA was used following Affymetrix instruction. 3 biological samples for each genotype.

Project description:Sympathetic neurons of SCG (Superior Cervical Ganglia) send axonal projections either along the external carotid arteries to innervate the salivary glands, or along the internal carotid arteries to the lacrimal and pineal glands, the eye, blood vessels and skin of the head, and the mucosa of the oral and nasal cavities. Previous studies using Wnt1Cre and R26R have defined the neural crest and mesodermal origins of vascular smooth muscle in the heart outflow tract and great vessels, although not specifically of the segments that are relevant for the projections of the SCG neurons. The third pharyngeal arch arteries are lined by neural crest-derived smooth muscle, and consequently, their derivatives, including the entirety of the external carotid arteries and only the base of the internal carotid arteries, also have a neural crest origin. In contrast, the dorsal aortae are lined by smooth muscle that is mesodermal in origin, and as a result, the internal carotid arteries from just above their origination from the common carotid arteries have a mesoderm-derived smooth muscle layer. To address the possibility that guidance cues for SCG neurons are selectively expressed by the external carotid vs. the internal carotid arteries, we isolated these segments of the vasculature from mouse embryos at E13.5 and extracted RNA to screen microarrays for differentially expressed genes. Experiment Overall Design: Vascular segments were isolated from 22 embryos at E13.5, pooled and extracted RNA for microarray screen. Total RNA samples from the internal or the external carotid arteries were subjected for two-round amplification to synthesize cRNA to probe microarray. Neither experimental nor technical replicate was made for this experiment. Experiment Overall Design: Vascular segments were isolated from 22 embryos at E13.5, pooled and extracted RNA for microarray screen. Total RNA samples from the internal or the external carotid arteries were subjected for two-round amplification to synthesize cRNA to probe microarray. Neither experimental nor technical replicate was made for this experiment.

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Single-cell transcriptome of the murine carotid artery following acute injury

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