Comparison of gene expression data between wild-type and DM1-affected Mesodermal Precursors Cells (MPC). Homo sapiens

Analysis of genes that were differentially expressed in mutant VUB03_DM1 as compared to controls VUB01 and SA01 Mesodermal Precursors Cells. Embryonic stem (ES) cell lines provide, theoretically, unlimited access to any needed amount of any specific cell phenotype of an organism, due to their unique capacities at indefinite self-renewal and pluripotency (Smith 2001; Trounson 2006). These properties allow using the progeny of ES cell lines to model human pathologies (Martinat, Shendelman et al. 2004; Lerou and Daley 2005; Ben-Nun and Benvenisty 2006). In particular, human ES cell lines derived from embryos characterized as gene-carriers following pre-implantation genetic diagnosis (PGD) for any one of major monogenic diseases (Pickering, Minger et al. 2005; Mateizel, De Temmerman et al. 2006) may be considered as perfect cellular replicas of those diseases, as they exhibit the exact genotypes associated to them. Here, we confirm this hypothesis by demonstrating that the cell progeny of an ES cell line derived from an embryo with myotonic dystrophy type 1 (DM1) displayed the morphological stigma associated to the expression of the mutant gene –so-called intranuclear foci- as well as abnormal alternate splicing of the insulin receptor, a characteristic feature of DM1. Further differential transcriptomic analysis of the DM1 gene-carrying cells with phenotypically similar populations from native ES cell lines revealed abnormal expression of 89 genes, among which 48 were down-regulated and 39 over-expressed. This study demonstrates that DM1, though a disease with relatively late clinical onset, is associated with expression of genetic defects early on during development. It underlines the value of PGD-derived ES cell lines as a tool to decipher molecular mechanisms of genetic diseases. Ben-Nun, I. F. and N. Benvenisty (2006). Human embryonic stem cells as a cellular model for human disorders. Mol Cell Endocrinol 252(1-2): 154-9. Lerou, P. H. and G. Q. Daley (2005). Therapeutic potential of embryonic stem cells. Blood Rev 19(6): 321-31. Martinat, C., S. Shendelman, et al. (2004). Sensitivity to oxidative stress in DJ-1-deficient dopamine neurons: an ES- derived cell model of primary Parkinsonism. PLoS Biol 2(11): e327. Mateizel, I., N. De Temmerman, et al. (2006). Derivation of human embryonic stem cell lines from embryos obtained after IVF and after PGD for monogenic disorders. Hum Reprod 21(2): 503-11. Pickering, S. J., S. L. Minger, et al. (2005). Generation of a human embryonic stem cell line encoding the cystic fibrosis mutation deltaF508, using preimplantation genetic diagnosis. Reprod Biomed Online 10(3): 390-7. Smith, A. G. (2001). Embryo-derived stem cells: of mice and men. Annu Rev Cell Dev Biol 17: 435-62. Trounson, A. (2006). The production and directed differentiation of human embryonic stem cells. Endocr Rev 27(2): 208-19. Keywords: disease state analysis Overall design: Two controls hES-derived MPC (VUB01 and SA01) and one mutant hES-derived MPC (VUB03_DM1), with three biological replicats for each

本研究针对突变体VUB03_DM1与对照VUB01、SA01来源的中胚层前体细胞之间的差异表达基因展开分析。胚胎干细胞（embryonic stem, ES）系凭借其无限自我更新与多能性的独特特性，理论上可无限获取生物体任意特定细胞表型所需的材料（Smith 2001; Trounson 2006）。上述特性使得ES细胞系的子代细胞可用于构建人类疾病模型（Martinat、Shendelman等，2004; Lerou与Daley 2005; Ben-Nun与Benvenisty 2006）。
具体而言，通过植入前基因诊断（pre-implantation genetic diagnosis, PGD）确认携带单基因遗传病致病基因的胚胎所建立的人类ES细胞系（Pickering、Minger等，2005; Mateizel、De Temmerman等，2006），可作为对应疾病的完美细胞复现模型，因其携带与疾病完全一致的基因型。
本研究针对1型强直性肌营养不良（myotonic dystrophy type 1, DM1）胚胎来源的ES细胞系验证了上述假说：该细胞系的子代细胞不仅呈现出与突变基因表达相关的典型形态特征——即所谓核内灶（intranuclear foci），还表现出胰岛素受体的异常可变剪接，这也是DM1的典型特征之一。
进一步对携带DM1致病基因的细胞与源自天然ES细胞系的表型相似细胞群开展差异转录组分析，结果显示共有89个基因的表达存在异常，其中48个基因表达下调，39个基因表达上调。
本研究证实，尽管1型强直性肌营养不良属于临床发病相对较晚的疾病，但其遗传缺陷的表达在发育早期便已显现。本研究同时凸显了经PGD建立的ES细胞系作为解析遗传疾病分子机制工具的重要价值。
参考文献：
Ben-Nun, I. F. 与 N. Benvenisty (2006). 人类胚胎干细胞作为人类疾病的细胞模型. Mol Cell Endocrinol 252(1-2): 154-9.
Lerou, P. H. 与 G. Q. Daley (2005). 胚胎干细胞的治疗潜力. Blood Rev 19(6): 321-31.
Martinat, C., S. Shendelman 等 (2004). DJ-1缺陷型多巴胺神经元对氧化应激的敏感性：原发性帕金森病的ES细胞模型. PLoS Biol 2(11): e327.
Mateizel, I., N. De Temmerman 等 (2006). 从体外受精（in vitro fertilization, IVF）后胚胎及单基因病PGD后胚胎中建立人类胚胎干细胞系. Hum Reprod 21(2): 503-11.
Pickering, S. J., S. L. Minger 等 (2005). 利用植入前基因诊断建立携带囊性纤维化突变deltaF508的人类胚胎干细胞系. Reprod Biomed Online 10(3): 390-7.
Smith, A. G. (2001). 胚胎来源的干细胞：小鼠与人类. Annu Rev Cell Dev Biol 17: 435-62.
Trounson, A. (2006). 人类胚胎干细胞的制备与定向分化. Endocr Rev 27(2): 208-19.
关键词：疾病状态分析
整体实验设计：两组对照人类ES细胞来源的中胚层前体细胞（VUB01与SA01）以及一组突变体人类ES细胞来源的中胚层前体细胞（VUB03_DM1），每组均设置3次生物学重复。