Single-Cell RNAseq Analysis of Murine Fate-Mapped Tsc1+/+ and Tsc1-/- Aortic Smooth Muscle Cells. Single-Cell RNAseq Analysis of Murine Fate-Mapped Tsc1+/+ and Tsc1-/- Aortic Smooth Muscle Cells

We characterized the transcriptomic effects at the single cell level of chronic mTOR activation in aortic SMCs using a conditional genetic model to delete Tsc1 under control of a smooth muscle-specific Myh11 promoter Overall design: Myh11-CreERT2.mT/mG (Tsc1+/+) and Tsc1fl/fl.Myh11-CreERT2.mT/mG (Tsc1−/−) male mice were treated with tamoxifen at 1.5 wk, the thoracic aortas were procured and enzyme digested at 24 wk, single, viable, GFP+ SMCs were isolated by FACS, and analyzed by single-cell RNAseq using the 10x system.

本研究借助以平滑肌特异性Myh11启动子调控的条件性遗传模型敲除Tsc1，构建主动脉平滑肌细胞（aortic smooth muscle cells, SMCs）慢性mTOR（哺乳动物雷帕霉素靶蛋白，mammalian target of rapamycin, mTOR）激活模型，并对该模型下单细胞水平的转录组学效应开展了系统性表征。
实验整体设计：将Myh11-CreERT2.mT/mG（Tsc1+/+）与Tsc1fl/fl.Myh11-CreERT2.mT/mG（Tsc1−/−）雄性小鼠于出生后1.5周施以他莫昔芬处理；于24周时采集胸主动脉并进行酶解消化，通过荧光激活细胞分选（fluorescence-activated cell sorting, FACS）分离得到单个存活的GFP阳性平滑肌细胞，随后采用10x单细胞测序系统完成单细胞RNA测序分析。