Embryonic Gene Expression During Diabetic Embryopathy

描述(申请人提供)：项目摘要：糖尿病胚胎病变是一种糖尿病并发症，患有糖尿病的母亲的早期胚胎会发展为先天性畸形。拟议的实验采用了糖尿病怀孕的小鼠模型，在该模型中，控制必要发育过程的基因的表达受到干扰。这些实验的重点是Pax3，这是一种在糖尿病小鼠胚胎中表达显著减少的基因，它编码一种转录因子，在糖尿病胚胎病变期间，这些结构通常是畸形的，特别是神经管和心脏。在之前的资助期间，我们发现母亲的高血糖增加了葡萄糖向胚胎的输送，而过度的葡萄糖代谢导致的氧化和低氧应激导致Pax3的表达受损，从而导致神经管缺陷(NTD)。除了使用胚胎进行实验外，我们还使用了小鼠胚胎干细胞，这种细胞可以大量生长，并可以被诱导为表达Pax3的神经上皮细胞类型。在这项建议中要检验的中心假设是，早期胚胎葡萄糖代谢增加所影响的生化途径会引起氧化和低氧应激，从而阻止激活必要发育控制基因表达的转录因子的诱导或结合。在这项建议中，我们将：(1)研究低氧应激、氧化应激和蛋白激酶C活性增加导致Pax3表达受损的相互作用；(2)确定与Pax3调控元件结合的因素，该调控元件是诱导分化所需的，并被氧化应激抑制；以及(3)检验以下假设：导致小鼠ES细胞Pax3表达受损的相同生化干扰也导致人类ES细胞PAX3表达受损。相关性：了解母体糖尿病如何在生化和分子水平上干扰早期胚胎发育，对于制定新的预防策略至关重要。这项拟议的研究将使用糖尿病妊娠的小鼠模型和小鼠胚胎干细胞来阐明重要胚胎基因表达受到干扰的生化机制，并使用人类胚胎干细胞来确定小鼠和小鼠干细胞在多大程度上作为人类糖尿病妊娠期间生化紊乱影响的模型。

DESCRIPTION (provided by applicant): Project Summary: Diabetic embryopathy is a diabetic complication in which the early embryo of a mother with diabetes develops congenital malformations. The proposed experiments employ a mouse model of diabetic pregnancy in which expression of genes which control essential developmental processes is disturbed. These experiments focus on Pax3, a gene whose expression is significantly reduced in embryos of diabetic mice, which encodes a transcription factor required for development of structures that are often malformed during diabetic embryopathy, especially the neural tube and the heart. In the previous funding period, we showed that maternal hyperglycemia increases glucose delivery to the embryo, and that oxidative and hypoxic stress, resulting from excess glucose metabolism, causes impaired Pax3 expression, thereby leading to neural tube defects (NTD). In addition to experiments using embryos, we have used murine embryonic stem (ES) cells, which can be grown in quantity and can be induced to a Pax3-expressing neuroepithelial cell type. The central hypothesis to be tested in this proposal is that biochemical pathways affected by increased glucose metabolism by the early embryo causes oxidative and hypoxic stress, which prevents the induction or binding of transcription factors that activate expression of essential developmental control genes. In this proposal we will: (1) Investigate the interaction of hypoxic stress, oxidative stress, and increased protein kinase C activity leading to impaired Pax3 expression; (2) Identify factors which bind to a Pax3 regulatory element that is needed for differentiation-induced expression and which is inhibited by oxidative stress; and (3) Test the hypothesis that the same biochemical disturbances that lead to impaired Pax3 expression in mouse ES cells also lead to impaired PAX3 expression in human ES cells. Relevance: Understanding how maternal diabetes disturbs early embryonic development on a biochemical and molecular level is essential in order to devise new strategies to prevent them. The proposed research will use a mouse model of diabetic pregnancy and mouse embryonic stem cells to elucidate the biochemical mechanisms by which important embryonic gene expression is disturbed, and human embryonic stem cells to determine the extent to which the mouse and mouse stem cells serve as a model for the effects of biochemical disturbances during human diabetic pregnancy.