Single-cell RNA-seq of vascular endothelial cells in the mouse model of vascular injury

To elucidate the mechanism of vascular repair, we performed single-cell analysis using the mouse vascular injury model (oxygen-induced retinopathy). In this model, mice were exposed to hyperoxic conditions at postnatal day 7 (P7) to induce ischemic retinopathy and the shedding of blood vessels. A return to normal oxygen conditions (normoxia) at P12 results in neovascularization that peaks at P17 and returns to normal by P30. CD31+/CD45neg vascular endothelial cells (ECs) were harvested at P12 (ischemic phase), P17 (retinopathy phase), and P25 (vascular repair phase) from retina only or retina plus optic nerve (6 samples) and analyzed together with corresponding control samples (total of 12 samples). Isolated tissues were digested, stained for CD31/CD45, and isolated by fluorescence-activated cell sorting (FACS). ECs were then loaded into the 10x Chromium Single Cell 3' Kit (v3) to obtain approximately 5,000 cells per well.