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: 9225120
• 负责人: DEAN R APPLING
• 金额: $ 17.5万
• 依托单位: UNIVERSITY OF TEXAS AT AUSTIN
• 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 2017-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9225120
• 关键词: Address; Adenine; Anabolism; Apoptosis; Biochemical Reaction; Biological Assay; Carbon; Cell Death; Cell Differentiation process; Cell Proliferation; Cell physiology; Cellular Assay; Cephalic; Cessation of life; Chinese Hamster Ovary Cell; Code; Congenital Abnormality; Craniofacial Abnormalities; DNA Resequencing; Data; Defect; Developing Countries; Diet; Dietary Intervention; Differentiation and Growth; Embryo; Enzymes; Epigenetic Process; Etiology; Failure; Folic Acid; Folic Acid Deficiency; Genes; Genetic; Growth; Histones; Human; Inbred Strains Mice; Incidence; Inositol; Intervention; Investigation; Knockout Mice; Laboratories; Link; Metabolic; Metabolism; Methionine; Methylation; Mitochondria; Modeling; Mouse Strains; Mus; Mutant Strains Mice; Mutation; Neural Crest; Neural Tube Closure; Neural Tube Defects; Neural tube; Nucleotides; Nutritional; Pathway interactions; Pharmacologic Substance; Phenotype; Plasmids; Population; Predisposition; Prevalence; Process; Public Health; Purines; Reaction; Research; Resistance; Risk; Role; S-Adenosylmethionine; Secondary to; Series; Site-Directed Mutagenesis; Spinal Dysraphism; Stem cells; Structural Congenital Anomalies; Supplementation; Supporting Cell; System; Target Populations; Teratogens; Testing; Tetrahydrofolates; Thymidine; Validation; Valproic Acid; Variant; base; cell growth; cell motility; chromatin immunoprecipitation; cohort; design; effective intervention; folic acid supplementation; fortification; in vivo; metabolome; metabolomics; methyl group; migration; mouse model; mutant; mutant mouse model; nerve stem cell; neuroepithelium; non-genetic; orofacial; prevent; programs; promoter; public health relevance; role model; sound; thymidylate; transcriptome sequencing

 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的原因是多因素的，包括遗传、环境和营养因素。母体叶酸（FA）状态是与NTD易感性最密切的联系之一。许多研究表明，在某些人群中，补充FA可使NTD患病率降低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。我们已经开始利用一种新的小鼠NTD模型（Mthfd 11 KO）来解决现有的数据缺口，该模型密切复制了人类NTD表型，并且不需要额外的营养干预来表达NTD表型。在这种新的小鼠模型中，特定叶酸依赖性酶（线粒体MTHFD 1 L）的缺失导致NTD。这是与NTD易感性/病因学相关的最具体的代谢缺陷，并表明FA为核苷酸和甲基生物合成提供了必需的一碳单元。这些生物合成途径在快速生长的胚胎中特别活跃，它们在神经管闭合（NTC）期间支持细胞增殖和死亡，迁移和分化。我们将使用该小鼠模型检验以下特定假设：（1）母体补充蛋氨酸、嘌呤、胸苷酸和S-腺苷蛋氨酸可以保护缺失型Mthfd 11 KO中的NTD（Mthfd 1 lz/z）胚胎，（2）Depakote（丙戊酸; VPA）是药物诱导的NTD的主要原因，抑制线粒体1C代谢，因此，甲酸可能可以防止由这种致畸剂引起的NTD，和（3）在Mthfd 11 z/z胚胎中细胞增殖和凋亡、细胞迁移和分化程序被破坏，导致神经管和口面缺陷。我们证明，母亲补充MTHFD 1 L母鼠甲酸盐，MTHFD 1 L酶反应的产物，降低NTDs的发生率，并部分挽救缺乏功能性MthFD 1 l的胚胎的生长缺陷。在具体目标1中，我们将确定MTHFD 1 L反应下游的哪些补充剂可以拯救NTD表型。这将在许多FA应答和非应答NTD突变株以及VPA敏感小鼠株中进行探索。我们将确定Mthfd 1 lz/z胚胎和VPA敏感小鼠品系中哪些细胞过程失调，导致不适当的NTC。将进行代谢组学和表观遗传学研究，以全面描述突变小鼠的特征。具体目标2将侧重于甲酸在神经干细胞和神经嵴干细胞使用神经球生长和分化测定，以及额外的表观遗传学研究的需求。具体目标3将涉及人类MTHFD 1 L基因的DNA重测序和脊柱裂队列中已鉴定变体的功能分析。这项研究计划通过阐明甲酸盐预防NTDs的潜在机制，为开发第一种有效干预非FA反应性NTDs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。