Development and Validation of Novel Amelogenesis Models
新型釉质生成模型的开发和验证
基本信息
- 批准号:9796443
- 负责人:
- 金额:$ 31.2万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2019
- 资助国家:美国
- 起止时间:2019-08-01 至 2021-07-31
- 项目状态:已结题
- 来源:
- 关键词:Acetyl Coenzyme AAcidsAmeloblastsAmelogenesisAmelogenesis ImperfectaAntibodiesBicarbonatesBiomimeticsBrainCRISPR/Cas technologyCaringCitrate (si)-SynthaseCitratesCitric Acid CycleDefectDentalDental EnamelDental Enamel HypoplasiaDentinDepositionDevelopmentDimensionsDiseaseDisease ProgressionDissectionDistalEnamel FormationEnergy MetabolismEnsureEpitopesExhibitsExtracellular SpaceGene ExpressionGene ProteinsGenesGraphHardnessHeterozygoteHumanImmunohistochemistryIn Situ HybridizationIn VitroIncisorIndividualInheritedIonsKnock-inKnock-in MouseKnock-outKnockout MiceLeftLiverMembraneMicroscopyMineralsMissense MutationMitochondrial MatrixModelingMusMutationNonsense CodonOralOrganOxaloacetatesPatientsPatternPersonsPhasePhenotypePlayPositioning AttributePredispositionProcessProteinsRadioReactionResearchResearch ProposalsRoleSurfaceSyndromeTerminator CodonThickTooth TissueTooth structureTranslationsValidationWaterWild Type Mouseameloblastinbasebiomineralizationbonecitrate carrierdesignepileptic encephalopathiesextracellularhuman diseaseimprovedinfancyinsightloss of functionmalformationmouse modelnoveloutcome forecastprematureretinal rods
项目摘要
Abstract
This research proposal focuses on the secretory stage of amelogenesis where enamel mineral ribbons initiate
on dentin mineral and elongate until the enamel layer reaches its final dimensions. Many genes cause inherited
enamel malformations (amelogenesis imperfecta, AI). Defects in genes necessary for the secretory stage
typically cause thinner (hypoplastic) enamel with a rough surface. Dental enamel forms in a defined extracellular
space that is established and modified by ameloblasts. We cite 19 human AI genes causing enamel hypoplasia,
but focus on those most directly associated with the extracellular enamel matrix. Five genes encode secreted
matrix proteins: ENAM, AMBN, AMELX, MMP20, and ODAPH. Except for ODAPH (formerly C4orf26), these
genes/proteins are well-characterized. There are 2 key secretory stage ion transporters: SLC4A4 (NCBe1;
transports bicarbonate into enamel matrix) and SLC13A5. Bicarbonate is known to neutralize the acid generated
by mineral deposition, but why absence of the citrate transporter (NaCT) encoded by SLC13A5 causes severe
enamel malformations is unknown.
SLC13A5 is required in soft (liver, brain) and hard (bone, teeth) tissues. Surprisingly ~80% of all citrate in the
body is in bone. Citrate is part of the Citric Acid Cycle where citrate is generated from oxaloacetate, acetyl-
coenzyme A, and water in a reaction catalyzed by citrate synthase within the mitochondrial matrix. Citrate likely
plays 1 of 2 roles: It could be transported into ameloblasts (influx) across its proximal membrane to increase
energy metabolism or be transported out of ameloblasts (efflux) across its distal membrane into the developing
enamel matrix. Based upon findings of citrate in bone and enamel, we hypothesize citrate is secreted and helps
to regulate enamel ribbon deposition. The critical roles of ODAPH and SLC13A5 in enamel ribbon formation
represent two major gaps in our understanding of amelogenesis. We close these gaps by generating and
validating Odaph and Slc13a5 knockout (KO) mice expressing premature stop codons homologous to human
AI-causing mutations. We also generate a Slc13a5 knockin (KI) expressing 3 FLAG epitopes on its C-terminus.
Two Specific Aims are proposed:
SA1: To develop and validate a Slc13a5 KO mouse model homologous to human AI (p.Arg333*) and to
generate a Slc13a5FLAG KI mouse for sensitive and specific immunolocalization (IHC).
UG3: generate a Slc13a5-/- mouse AI model in C57BL/6J background using CRISPR/Cas9.
UG3: generate a Slc13a5FLAG mouse wild-type-tagged model for sensitive and specific localization.
UH3: validate the Slc13a5-/- mouse by characterizing its enamel. Validate Slc13a5FLAG mouse by IHC.
SA2: To develop and validate an Odaph KO mouse model homologous to human AI (p.Cys43*).
UG3: generate an Odaph-/- mouse AI model in C57BL/6J background using CRISPR/Cas9.
UH3: validate the Odaph-/- mouse by characterizing its enamel phenotype.
摘要
这项研究计划的重点是分泌阶段的釉质矿物质带开始
在牙本质矿物质上延伸,直到釉质层达到其最终尺寸。许多基因导致遗传
釉质畸形(釉质形成,AI)。分泌期所必需的基因缺陷
典型地导致具有粗糙表面的较薄(发育不良)釉质。牙釉质形成在一个明确的细胞外
由成釉细胞建立和改变的空间。我们引用了19个人类AI基因导致釉质发育不全,
而是集中在与细胞外釉质基质最直接相关的那些。五个基因编码分泌型
基质蛋白:ENAM、AMBN、AMELX、MMP 20和ODAPH。除了ODAPH(以前的C4 orf 26),
基因/蛋白质被很好地表征。分泌期的离子转运蛋白有两个:SLC 4A 4(NCBe 1;
将碳酸氢盐转运到釉质基质中)和SLC 13 A5。众所周知,重碳酸盐可以中和产生的酸
但为什么SLC 13 A5编码的柠檬酸盐转运蛋白(NaCT)的缺失会导致严重的
牙釉质畸形是未知的。
SLC 13 A5在软组织(肝脏,大脑)和硬组织(骨骼,牙齿)中是必需的。令人惊讶的是,
身体在骨头里。柠檬酸盐是柠檬酸循环的一部分,其中柠檬酸盐由草酰乙酸、乙酰-
辅酶A和水在由线粒体基质内的柠檬酸合酶催化的反应中。可能是柠檬酸盐
发挥2种作用之一:它可以通过成釉细胞近端膜转运到成釉细胞(内流),
能量代谢或被运输出成釉细胞(流出)穿过其远端膜进入发育中的
釉质基质基于骨和釉质中柠檬酸盐的发现,我们假设柠檬酸盐是分泌的,
以调节釉带沉积。ODAPH和SLC 13 A5在釉带形成中的关键作用
代表了我们对釉质形成理解的两个主要空白。我们通过生成和
验证表达与人类同源的提前终止密码子的Odaph和Slc 13 a5敲除(KO)小鼠
导致人工智能的突变我们还产生了在其C-末端表达3个FLAG表位的Slc 13 a5敲入(KI)。
提出了两个具体目标:
SA 1:开发并验证与人AI(p.Arg333*)同源的Slc 13 a5 KO小鼠模型,
产生用于敏感和特异性免疫定位(IHC)的Slc 13 a5 FLAG KI小鼠。
UG 3:使用CRISPR/Cas9在C57 BL/6 J背景中生成Slc 13 a5-/-小鼠AI模型。
UG 3:生成Slc 13 a5 FLAG小鼠野生型标记模型,用于敏感和特异性定位。
UH 3:通过表征其牙釉质来验证Slc 13 a5-/-小鼠。免疫组化法检测Slc 13 a5 FLAG小鼠。
SA 2:开发并验证与人AI(p.Cys43*)同源的Odaph KO小鼠模型。
UG 3:使用CRISPR/Cas9在C57 BL/6 J背景中生成Odaph-/-小鼠AI模型。
UH 3:通过表征其釉质表型来验证Odaph-/-小鼠。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
数据更新时间:{{ journalArticles.updateTime }}
{{
item.title }}
{{ item.translation_title }}
- DOI:
{{ item.doi }} - 发表时间:
{{ item.publish_year }} - 期刊:
- 影响因子:{{ item.factor }}
- 作者:
{{ item.authors }} - 通讯作者:
{{ item.author }}
数据更新时间:{{ journalArticles.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ monograph.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ sciAawards.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ conferencePapers.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ patent.updateTime }}
JAN Ching Chun HU其他文献
JAN Ching Chun HU的其他文献
{{
item.title }}
{{ item.translation_title }}
- DOI:
{{ item.doi }} - 发表时间:
{{ item.publish_year }} - 期刊:
- 影响因子:{{ item.factor }}
- 作者:
{{ item.authors }} - 通讯作者:
{{ item.author }}
{{ truncateString('JAN Ching Chun HU', 18)}}的其他基金
Development and Validation of Novel Amelogenesis Models
新型釉质生成模型的开发和验证
- 批准号:
10460291 - 财政年份:2021
- 资助金额:
$ 31.2万 - 项目类别:
Development and Validation of Novel Amelogenesis Models
新型釉质生成模型的开发和验证
- 批准号:
10416109 - 财政年份:2021
- 资助金额:
$ 31.2万 - 项目类别:
Why is Fam83h critical for enamel formation?
为什么 Fam83h 对于牙釉质形成至关重要?
- 批准号:
8441387 - 财政年份:2009
- 资助金额:
$ 31.2万 - 项目类别:
Why is Fam83h critical for enamel formation?
为什么 Fam83h 对于牙釉质形成至关重要?
- 批准号:
7623768 - 财政年份:2009
- 资助金额:
$ 31.2万 - 项目类别:
Why is Fam83h critical for enamel formation?
为什么 Fam83h 对于牙釉质形成至关重要?
- 批准号:
8048006 - 财政年份:2009
- 资助金额:
$ 31.2万 - 项目类别:
Why is Fam83h critical for enamel formation?
为什么 Fam83h 对于牙釉质形成至关重要?
- 批准号:
8246309 - 财政年份:2009
- 资助金额:
$ 31.2万 - 项目类别:
Why is Fam83h critical for enamel formation?
为什么 Fam83h 对于牙釉质形成至关重要?
- 批准号:
7780358 - 财政年份:2009
- 资助金额:
$ 31.2万 - 项目类别:
相似国自然基金
具有抗癌活性的天然产物金霉酸(Aureolic acids)全合成与选择性构建2-脱氧糖苷键
- 批准号:22007039
- 批准年份:2020
- 资助金额:24.0 万元
- 项目类别:青年科学基金项目
海洋放线菌来源聚酮类化合物Pteridic acids生物合成机制研究
- 批准号:
- 批准年份:2019
- 资助金额:10.0 万元
- 项目类别:省市级项目
手性Lewis Acids催化的分子内串联1,5-氢迁移/环合反应及其在构建结构多样性手性含氮杂环化合物中的应用
- 批准号:21372217
- 批准年份:2013
- 资助金额:80.0 万元
- 项目类别:面上项目
对空气稳定的新型的有机金属Lewis Acids催化剂制备、表征与应用研究
- 批准号:21172061
- 批准年份:2011
- 资助金额:30.0 万元
- 项目类别:面上项目
钛及含钛Lewis acids促臭氧/过氧化氢体系氧化性能的广普性、高效性及其机制
- 批准号:21176225
- 批准年份:2011
- 资助金额:60.0 万元
- 项目类别:面上项目
基于Zip Nucleic Acids引物对高度降解和低拷贝DNA检材的STR分型研究
- 批准号:81072511
- 批准年份:2010
- 资助金额:31.0 万元
- 项目类别:面上项目
海洋天然产物Makaluvic acids 的全合成及其对南海鱼虱存活的影响
- 批准号:30660215
- 批准年份:2006
- 资助金额:21.0 万元
- 项目类别:地区科学基金项目
相似海外基金
Lipid nanoparticle-mediated Inhalation delivery of anti-viral nucleic acids
脂质纳米颗粒介导的抗病毒核酸的吸入递送
- 批准号:
502577 - 财政年份:2024
- 资助金额:
$ 31.2万 - 项目类别:
CAREER: Highly Rapid and Sensitive Nanomechanoelectrical Detection of Nucleic Acids
职业:高度快速、灵敏的核酸纳米机电检测
- 批准号:
2338857 - 财政年份:2024
- 资助金额:
$ 31.2万 - 项目类别:
Continuing Grant
Double Incorporation of Non-Canonical Amino Acids in an Animal and its Application for Precise and Independent Optical Control of Two Target Genes
动物体内非规范氨基酸的双重掺入及其在两个靶基因精确独立光学控制中的应用
- 批准号:
BB/Y006380/1 - 财政年份:2024
- 资助金额:
$ 31.2万 - 项目类别:
Research Grant
Quantifying L-amino acids in Ryugu to constrain the source of L-amino acids in life on Earth
量化 Ryugu 中的 L-氨基酸以限制地球生命中 L-氨基酸的来源
- 批准号:
24K17112 - 财政年份:2024
- 资助金额:
$ 31.2万 - 项目类别:
Grant-in-Aid for Early-Career Scientists
Synthetic analogues based on metabolites of omega-3 fatty acids protect mitochondria in aging hearts
基于 omega-3 脂肪酸代谢物的合成类似物可保护衰老心脏中的线粒体
- 批准号:
477891 - 财政年份:2023
- 资助金额:
$ 31.2万 - 项目类别:
Operating Grants
Metabolomic profiles of responders and non-responders to an omega-3 fatty acids supplementation.
对 omega-3 脂肪酸补充剂有反应和无反应者的代谢组学特征。
- 批准号:
495594 - 财政年份:2023
- 资助金额:
$ 31.2万 - 项目类别:
Molecular recognition and enantioselective reaction of amino acids
氨基酸的分子识别和对映选择性反应
- 批准号:
23K04668 - 财政年份:2023
- 资助金额:
$ 31.2万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
Integrated understanding and manipulation of hypoxic cellular functions by artificial nucleic acids with hypoxia-accumulating properties
具有缺氧累积特性的人工核酸对缺氧细胞功能的综合理解和操纵
- 批准号:
23H02086 - 财政年份:2023
- 资助金额:
$ 31.2万 - 项目类别:
Grant-in-Aid for Scientific Research (B)
Basic research toward therapeutic strategies for stress-induced chronic pain with non-natural amino acids
非天然氨基酸治疗应激性慢性疼痛策略的基础研究
- 批准号:
23K06918 - 财政年份:2023
- 资助金额:
$ 31.2万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
Molecular mechanisms how arrestins that modulate localization of glucose transporters are phosphorylated in response to amino acids
调节葡萄糖转运蛋白定位的抑制蛋白如何响应氨基酸而被磷酸化的分子机制
- 批准号:
23K05758 - 财政年份:2023
- 资助金额:
$ 31.2万 - 项目类别:
Grant-in-Aid for Scientific Research (C)