Transcriptome analysis of in-vitro differentiated myotubes from Emery-Dreifuss Muscular Dystrophy patients.

Emery-Dreifuss muscular dystrophy (EDMD) is a genetically and clinically variable disorder. Here we performed transcriptome analysis on 10 EDMD patients covering mutations in 7 EDMD-linked genes, compared to 2 healthy controls. Myoblasts were isolated from muscle biopsies and differentiated in-vitro for 6 days. Differentiated myotubes were isolated, total RNA extracted, and mRNA sequenced. The genes and mutations included were: patient 1 (TMEM214, p.R179H), patient 2 (PLPP7/NET39, p.M92K), patient 3 (SUN1, p.G68D, p.G388S), patient 4 (SYNE1, p.6869*, p.6869*), patient 5 (EMD, p.S58Sfs*1), patient 6 (FHL1, c.688+1G>A), patient 7 (FHL1, p.C224W), patient 8 (FHL1, p.V280M), patient 9 (LMNA, p.T528K), patient 10 (LMNA, p.R571S). The patients were found to segregate into 3 subgroups, defined as: gp1 = patients 1,2,5,7 and 10; gp2 = patients 3,6, and 8; gp3 = patients 4 and 9. Transcriptome analysis was performed comparing in-vitro differentiated EDMD patients against healthy controls. All samples were analysed in triplicate (separate in-vitro differentiation experiments). Four types of comparisons were considered: G1 = all EDMD patients as a single group compared to the controls; G10 = each individual patient, in triplicate, compared to the controls; G3 = each patient subgroup against the controls; G3x = each patient subgroup against all other patients.

Emery-Dreifuss肌营养不良症（Emery-Dreifuss muscular dystrophy, EDMD）是一类遗传与临床表型均具有异质性的疾病。本研究针对10例EDMD患者开展转录组分析（transcriptome analysis），这些患者携带7个EDMD相关基因的突变，同时设置2例健康对照。研究人员从肌肉活检标本（muscle biopsies）中分离成肌细胞（myoblasts），并进行6天的体外分化；收集分化完成的肌管（myotubes），提取总RNA（total RNA）并进行mRNA测序（mRNA sequencing）。纳入研究的基因及突变为：患者1（TMEM214，p.R179H）、患者2（PLPP7/NET39，p.M92K）、患者3（SUN1，p.G68D、p.G388S）、患者4（SYNE1，p.6869*、p.6869*）、患者5（EMD，p.S58Sfs*1）、患者6（FHL1，c.688+1G>A）、患者7（FHL1，p.C224W）、患者8（FHL1，p.V280M）、患者9（LMNA，p.T528K）、患者10（LMNA，p.R571S）。经分析，所有患者可分为3个亚组：组1（gp1）：患者1、2、5、7及10；组2（gp2）：患者3、6及8；组3（gp3）：患者4及9。本研究通过对比体外分化的EDMD患者样本与健康对照样本开展转录组分析。所有样本均进行3次独立重复实验（独立体外分化实验）。研究共设置4类比较方案：G1：将所有EDMD患者作为单一组别与健康对照进行比较；G10：将每一例患者的3次重复样本分别与健康对照进行比较；G3：将每一个患者亚组分别与健康对照进行比较；G3x：将每一个患者亚组分别与其余所有患者亚组进行比较。