Effect of dexamethasone treatment on the transcriptome of injured mouse hamstring muscle. Effect of dexamethasone treatment on the transcriptome of injured mouse hamstring muscle

Healthy muscle heals and repairs after injury. However, when mice are treated with dexamethasone, an agonist of glucocorticoid receptors, heterotopic ossifications (HO) develop in the injured muscle within fibrotic areas that fail to fully regenerate functional muscle. To understand the mechanisms driving heterotopic ossification in response to glucocorticoid receptor agonists, we performed RNA seqencing analysis on injured muscle. Mice underwent muscle injuries by intramuscular injection of cardiotoxin (CDTX). All mice were treated with either vehicle or dexamethasone (10mg/kg, intraperitoneal injection) daily from day 0-4. polyA+ RNA was isolated from whole muscle 4 days post injury, libraries generated and sequenced. Overall design: Mice underwent muscle injuries by intramuscular injection of cardiotoxin (CDTX). All mice were injected intraperitoneally daily for 4 days with either saline (group C) or 10mg/kg dexamethasone (group D). RNA was isolated from whole CDTX-injured hindlimb muscle on 4 days post injury. Just prior to tissue harvest, heterotopic ossification development quantified in CDTX-injured muscles by micro computed tomography (µCT). Only 1 mouse (mouse C4) out of 5 in saline group developed heterotopic ossification (0.67mm3) and mouse C4 therefore was excluded in further analysis. On the other hand, all mice in dexamethasone-treated mice developed HO (2.40±5.42 mm3). PolyA+ RNA was then purfied before library construction and sequencing. contributor: UQ Sequencing Facility

健康肌肉在损伤后可完成愈合与修复过程。然而，当小鼠接受糖皮质激素受体激动剂地塞米松（dexamethasone）处理时，在未能完全再生功能性肌肉的纤维化区域内的损伤肌肉中，会形成异位骨化（heterotopic ossifications, HO）。为阐明糖皮质激素受体激动剂诱导异位骨化的作用机制，我们对损伤肌肉开展了RNA测序（RNA sequencing）分析。实验中，通过肌肉内注射心脏毒素（cardiotoxin, CDTX）构建小鼠肌肉损伤模型。所有小鼠自第0天至第4天，每日均接受溶剂对照或地塞米松（10mg/kg，腹腔注射）处理。于损伤后4天采集全肌肉组织，分离polyA+ RNA，构建测序文库并进行测序。整体实验设计：小鼠通过肌肉内注射心脏毒素（CDTX）造成肌肉损伤。所有小鼠每日腹腔注射给药，连续4天，分为两组：生理盐水对照组（C组）与10mg/kg地塞米松处理组（D组）。于损伤后4天采集受CDTX损伤的后肢肌肉组织以分离RNA。在组织收获前，通过显微计算机断层扫描（micro computed tomography, µCT）定量检测损伤肌肉中的异位骨化形成情况。结果显示，生理盐水组5只小鼠中仅1只（C4小鼠）形成异位骨化（体积0.67mm³），因此C4小鼠被排除在后续分析之外；而地塞米松处理组的全部小鼠均形成异位骨化，体积为2.40±5.42 mm³。随后纯化polyA+ RNA，进行文库构建与测序。本数据集贡献方：UQ测序平台（UQ Sequencing Facility）