Single-cell analysis reveals the progress in skin repair after radiation in rats.. Single-cell analysis reveals the progress in skin repair after radiation in rats.

This study aims to compare the cell heterogeneity and mRNA expression between irradiated and non-irradiated rat skin tissues at the single cell level. In this study, the skin was isolated from rats with radiation or non-irradiated rats. Male Sprague Dawley (SD) rats (6 weeks of age) were used. For irradiation, the hair on the gluteal region of the rats was shaved using a razor. The animals were immobilized using adhesive tape on a plastic plate to minimize motion during radiation exposure. A 3-cm-thick piece of lead was used to shield the animals and localize the radiation field (3-4 cm). Irradiation was performed on the treatment area at a dose rate of 1000 cGy/min using a 6-MeV electron beam accelerator. Irradiated skin was taken on day 7, 14, 28 and 60 for RNA-seq, respectively. The control group of skin were collected from four rats without radiation and mixed for RNA-seq. Overall design: Single cell RNA-seq analysis of skin tissue from irradiated and non-irradiated parts of SD rats at different time periods. ScRNA sequencing libraries were further generated by the 10x Chromium Single Cell 3′ Library & Single Cell 3’ v3 Gel Beads (10x Genomics). Briefly, gel beads with cell barcodes and skin cell suspension were wrapped in liquid droplets by a microfluidic platform (10x Genomics). A droplet containing single cells was collected and then lysed to generate Gel Bead-In Emulsions (GEMs).

本研究旨在在单细胞水平上比较受辐照与未受辐照大鼠皮肤组织的细胞异质性及mRNA表达水平。本研究从经辐照及未辐照大鼠体内分离皮肤组织，实验选用6周龄雄性Sprague Dawley（SD）大鼠。辐照前，使用剃刀剃除大鼠臀区毛发；将动物以胶带固定于塑料板上，以最大限度降低辐照过程中的位移。采用3cm厚铅块对动物进行屏蔽，并将辐照野范围限定为3-4cm。使用6-MeV电子束加速器，以1000 cGy/min的剂量率对处理区域实施辐照。分别于辐照后第7、14、28及60天采集辐照皮肤组织，用于RNA测序。对照组皮肤组织取自4只未辐照大鼠，混合后进行RNA测序。整体实验设计：对不同时间点SD大鼠辐照与未辐照皮肤组织开展单细胞RNA测序分析。单细胞RNA测序文库采用10x Chromium单细胞3′文库及单细胞3′ v3凝胶磁珠（10x Genomics）构建。简要流程如下：通过微流控平台（10x Genomics）将带有细胞条码的凝胶磁珠与皮肤细胞悬液包裹为液滴；收集包含单个细胞的液滴并裂解，生成凝胶磁珠包裹的乳液（Gel Bead-In Emulsions, GEMs）。