INTERVENTION STRATEGIES FOR NON-FOLATE RESPONSIVE NEURAL TUBE DEFECTS

非叶酸反应性神经管缺陷的干预策略

• 批准号: 9636317
• 负责人: DEAN R APPLING
• 金额: $ 32.87万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-02-15 至 2021-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9636317
• 关键词: INTERVENTION; STRATEGIES; NON; FOLATE; RESPONSIVE

 DESCRIPTION (provided by applicant): Neural tube defects (NTDs) are among the most common birth defects in humans. The causes of NTDs are multifactorial, including genetic, environmental, and nutritional factors. Maternal folic acid (FA) status is one of the strongest links to NTD susceptibility. Numerous studies have shown that supplemental FA can reduce NTD prevalence by as much as 70% in some populations. Despite more than 40 years of intensive effort, we still do not understand the mechanisms that underlie these folate-dependent processes. We have begun to address this existing data gap utilizing a new mouse NTD model (Mthfd1l KO) that closely replicates the human NTD phenotype, and does not require additional nutritional intervention to express the NTD phenotype. In this new mouse model, loss of a specific folate-dependent enzyme (mitochondrial MTHFD1L) leads to NTDs. This is the most specific metabolic defect yet associated with NTD susceptibility/etiology, and suggests that FA provides essential one-carbon units for nucleotide and methyl group biosynthesis. These biosynthetic pathways are especially active in the rapidly growing embryo, where they support cell proliferation and death, migration, and differentiation during neural tube closure (NTC). We will test the following specific hypotheses using this mouse model: (1) Maternal supplementation with methionine, purines, thymidylate and S-adenosylmethionine can protect against NTDs in nullizygous Mthfd1l KO (Mthfd1lz/z) embryos, (2) Depakote (Valproic Acid; VPA), the leading cause of pharmaceutical-induced NTDs, inhibits mitochondrial 1C metabolism, thus it is possible that formate can prevent NTDs caused by this teratogen, and (3) cell proliferation and apoptosis, cell migration, and differentiation programs are disrupted in Mthfd1lz/z embryos, leading to neural tube and orofacial defects. We demonstrated that maternal supplementation of MTHFD1L dams with formate, the product of the MTHFD1L enzymatic reaction, decreases the incidence of NTDs and partially rescues the growth deficit in embryos lacking a functional Mthfd1l. In Specific Aim 1 we will determine which supplements downstream of the MTHFD1L reaction can rescue the NTD phenotype. This will be explored in a number of FA responsive and non-responsive NTD mutant strains, and in VPA-sensitive mouse strains. We will identify which cellular processes are dysregulated in Mthfd1lz/z embryos and VPA-sensitive mouse strains, leading to improper NTC. Metabolomic and epigenetic studies will be pursued to fully characterize the mutant mice. Specific Aim 2 will focus on the requirement for formate in neural stem cells and neural crest stem cells using neurosphere growth and differentiation assays, as well as additional epigenetic investigations. Specific Aim 3 will involve DNA resequencing of the human MTHFD1L gene and functional analyses of identified variants in a spina bifida cohort. This research program offers hope for developing the first effective intervention for non-FA responsive NTDs by illuminating the underlying mechanisms by which formate prevents NTDs. Developing interventions that benefit non-folate responsive NTDs is crucial for preventing these preventable birth defects.

 描述（由申请人提供）：神经管缺陷（NTD）是人类最常见的出生缺陷之一。 NTD 的病因是多因素的，包括遗传、环境和营养因素。母亲叶酸 (FA) 状况是与 NTD 易感性最密切的联系之一。大量研究表明，补充 FA 可以将某些人群的 NTD 患病率降低多达 70%。尽管经过 40 多年的努力，我们仍然不了解这些叶酸依赖性过程的机制。我们已经开始利用新的小鼠 NTD 模型 (Mthfd1l KO) 来解决这一现有的数据差距，该模型紧密复制人类 NTD 表型，并且不需要额外的营养干预来表达 NTD 表型。在这个新的小鼠模型中，一种特定的叶酸依赖性酶（线粒体 MTHFD1L）的缺失会导致 NTD。这是迄今为止与 NTD 易感性/病因学相关的最特异的代谢缺陷，表明 FA 为核苷酸和甲基生物合成提供了必需的一碳单位。这些生物合成途径在快速生长的胚胎中尤其活跃，在神经管闭合 (NTC) 过程中支持细胞增殖和死亡、迁移和分化。我们将使用该小鼠模型测试以下具体假设：(1) 母体补充蛋氨酸、嘌呤、胸苷酸和 S-腺苷甲硫氨酸可以预防 Mthfd1l KO (Mthfd1lz/z) 胚胎无效的 NTD，(2) Depakote（丙戊酸；VPA），药物诱导 NTD 的主要原因， 抑制线粒体1C代谢，因此甲酸盐可能可以预防这种致畸剂引起的NTD，并且（3）Mthfd1lz/z胚胎中的细胞增殖和凋亡、细胞迁移和分化程序被破坏，导致神经管和口面部缺陷。我们证明，母体给 MTHFD1L 母鼠补充甲酸盐（MTHFD1L 酶促反应的产物）可以降低 NTD 的发生率，并部分缓解缺乏功能性 Mthfd1l 的胚胎的生长缺陷。在具体目标 1 中，我们将确定 MTHFD1L 反应下游的哪些补充剂可以挽救 NTD 表型。这将在许多 FA 反应性和非反应性 NTD 突变株以及 VPA 敏感小鼠品系中进行探索。我们将确定 Mthfd1lz/z 胚胎和 VPA 敏感小鼠品系中哪些细胞过程失调，从而导致 NTC 不当。将进行代谢组学和表观遗传学研究，以充分表征突变小鼠的特征。具体目标 2 将通过神经球生长和分化测定以及其他表观遗传学研究，重点关注神经干细胞和神经嵴干细胞对甲酸盐的需求。具体目标 3 将涉及人类 MTHFD1L 基因的 DNA 重测序以及对脊柱裂队列中已识别变异的功能分析。该研究项目通过阐明甲酸盐预防 NTD 的潜在机制，为开发针对非 FA 反应性 NTD 的第一个有效干预措施提供了希望。制定有利于非叶酸反应性 NTD 的干预措施对于预防这些可预防的出生缺陷至关重要。