Ibuprofen and enteric neural crest precursor migration

布洛芬和肠神经嵴前体迁移

• 批准号: 8593450
• 负责人: Ellen Merrick Schill
• 金额: $ 2.85万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-20 至 2016-09-19
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8593450
• 关键词: Abdomen; Actins; Adhesions; Affect; Age; Age-Years; Anti-Inflammatory Agents; Anti-inflammatory; Biochemical; Cause of Death; Cell Survival; Cells; Cessation of life; Child; Colon; Congenital Abnormality; Congenital Megacolon; Cytoskeleton; Data; Defect; Development; Diet; Disease; Distal; Distant; Enteral; Enteric Nervous System; Environmental Exposure; Environmental Risk Factor; Enzymes; Exhibits; Exposure to; Family; Fetal Development; First Pregnancy Trimester; Fishes; Focal Adhesions; Folic Acid; Genes; Genetic Predisposition to Disease; Genetic Risk; Goals; Human; Ibuprofen; Image; Immunohistochemistry; In Vitro; Incidence; Individual; Infant; Intestines; Investigation; Lead; Length; Life; Measures; Mediating; Medicine; Microscopy; Molecular; Morbidity - disease rate; Mus; Mutation; Nervous system structure; Neural Crest; Neural Tube Defects; Neuroglia; Neurons; Obstruction; PTGS1 gene; PTGS2 gene; Pathway interactions; Pharmaceutical Preparations; Physiology; Population; Preclinical Drug Evaluation; Pregnancy; Proteins; Regulation; Research Project Grants; Risk; Severities; Structure; Supplementation; Susceptibility Gene; Testing; Therapeutic; Time; Transgenic Animals; United States; Vertebrates; Vomiting; Woman; Zebrafish; base; cell motility; cellular imaging; critical period; cyclooxygenase 1; disease-causing mutation; disorder risk; effective intervention; gene environment interaction; gene interaction; improved; in utero; in vivo; inhibitor/antagonist; migration; mortality; mouse model; nervous system development; novel; null mutation; premature; prevent; public health relevance; research study; time use

DESCRIPTION (provided by applicant): Birth defects are the most common cause of death in children under one year of age in the United States. The best way to reduce the morbidity and mortality of specific birth defects is to develop strategies to prevent them from occurring, as exemplified by the decrease of neural tube defects due to folic acid supplementation. Hirschsprung Disease (HSCR) is a birth defect of the enteric nervous system that occurs when enteric neural crest-derived cells (ENCCs) do not migrate all the way to the end of the bowel. The resulting distal bowel aganglionosis leads to a functional obstruction, which causes vomiting, abdominal distension and predisposes to premature death. There are many proteins important for normal ENCC migration and mutations in these genes increase the risk of HSCR, but none of these mutations cause disease in all affected children. Therefore, children who have HSCR must have additional factors that further increase their HSCR risk. To develop strategies to decrease HSCR occurrence, it is important to identify alterable factors, such as maternal diet or medications that could in combination with genetic defects cause disease in a child who would otherwise have been born healthy. Ibuprofen, a non-steroidal anti-inflammatory drug (NSAID) commonly used in pregnancy, inhibited ENCC migration in a zebrafish drug screen, causing a HSCR-like distal bowel aganglionosis in affected fish. Experiments in mammalian primary culture confirmed that ibuprofen slows ENCC migration and appears to disrupt actin cytoskeletal and focal adhesion regulation needed for efficient migration. The goals of this proposal are to determine the biochemical mechanism of ibuprofen's inhibition of ENCC migration and to determine if ibuprofen increases the likelihood of HSCR in genetically predisposed individuals. The biochemical mechanisms that ibuprofen disrupts to inhibit ENCC migration will be investigated using cutting edge time-lapse live cell imaging of fluorescently-tagged proteins. These investigations will allow for in depth analysis of ENCC actin protrusions and the dynamics of focal adhesions. In addition, ENCCs from transgenic animals with null mutations in Cox1 and Cox2, enzymes inhibited by ibuprofen, will be analyzed by immunohistochemistry and time-lapse imaging to determine if ibuprofen inhibits ENCC migration via a Cox- dependent mechanism. I will also determine if mice with null mutations in Cox1 and Cox2 have HSCR-like distal bowel aganglionosis. Finally, mice genetically predisposed to HSCR due to mutations in genes important for normal ENCC migration will be exposed to ibuprofen in utero during the critical period of ENS development. These experiments will examine if ibuprofen could be a common and avoidable exposure during pregnancy that increases HSCR risk by reducing the efficiency of ENCC migration.

描述（由申请人提供）：出生缺陷是美国一岁以下儿童死亡的最常见原因。降低特定出生缺陷发病率和死亡率的最佳方法是制定预防策略，例如补充叶酸可减少神经管缺陷。先天性巨结肠症（HSCR）是一种先天性肠神经系统缺陷，发生于肠神经嵴源性细胞（ENCC）不能一直迁移到肠末端的时候。由此产生的远端肠无神经节细胞症导致功能性梗阻，引起呕吐，腹胀，并易于过早死亡。有许多蛋白质对正常的ENCC迁移很重要，这些基因的突变会增加HSCR的风险，但这些突变不会在所有受影响的儿童中引起疾病。因此，患有HSCR的儿童必须有其他因素进一步增加其HSCR风险。为了制定减少HSCR发生的策略，重要的是要确定可变因素，例如母亲的饮食或药物，这些因素可能与遗传缺陷相结合，导致原本健康出生的孩子患病。在斑马鱼药物筛选中，常用于妊娠期的非甾体抗炎药（NSAID）伊曲康可抑制ENCC迁移，导致受影响鱼类出现HSCR样远端肠无神经节细胞症。在哺乳动物原代培养物中的实验证实，布洛芬减缓ENCC迁移，似乎破坏肌动蛋白细胞骨架和局部粘附调节所需的有效迁移。 本提案的目的是确定布洛芬抑制ENCC迁移的生化机制，并确定布洛芬是否增加遗传易感个体发生HSCR的可能性。布洛芬破坏抑制ENCC迁移的生化机制将使用荧光标记蛋白质的尖端时间推移活细胞成像进行研究。这些调查将允许在深入分析的ENCC肌动蛋白突起和动力学的局灶性粘连。此外，将通过免疫组织化学和延时成像分析来自具有Cox1和Cox2无效突变（布洛芬抑制的酶）的转基因动物的ENCC，以确定布洛芬是否通过考克斯依赖性机制抑制ENCC迁移。我还将确定Cox1和Cox2无效突变的小鼠是否具有HSCR样远端肠无神经节细胞症。最后，由于对正常ENCC迁移重要的基因突变，遗传上易患HSCR的小鼠将在ENS发育的关键时期在子宫内暴露于布洛芬。这些实验将检查布洛芬是否可能是怀孕期间常见和可避免的暴露，通过降低ENCC迁移的效率来增加HSCR风险。