Mechanisms of Circadian Rhythmicity in CLOCK-Deficient Mice

时钟缺陷小鼠的昼夜节律机制

• 批准号: 8506154
• 负责人: DAVID Raymond WEAVER
• 金额: $ 36.39万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-06-01 至 2018-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8506154
• 关键词: Affect; Age; Aging; Alleles; Animals; Aortic Valve Stenosis; Behavioral; Brain; Breeding; Bypass; Calcified; Cartilage; Cells; Circadian Rhythms; Complex; Complication; Couples; Defect; Deposition; Development; Disease; Event; Feedback; Fertility; Fertilization in Vitro; Gene Expression; Generations; Genes; Genetic; Genetic Models; Health; Heart; Heart Valves; Heterotopic Ossification; Hormonal; Human; Infertility; Knock-out; Label; Lead; Lesion; Logic; Male Infertility; Mediating; Medical; Methods; Molecular; Molecular Analysis; Motion; Mus; Operative Surgical Procedures; Osteogenesis; Other Genetics; Pain; Pathologic Processes; Pathology; Periodicity; Phenotype; Population; Prevention; Process; Production; Regulation; Reporting; Reproductive Behavior; Reproductive Endocrinology; Role; Serotyping; Sex Behavior; Site; Skeletal system; Somatic Cell; Spermatogenesis; Sterility; Tamoxifen; Tendon structure; Testing; Testis; Therapeutic; Time; Tissues; Transcription Coactivator; Trauma; age related; aortic valve; arthropathies; bone; calcification; cell type; circadian pacemaker; economic impact; improved; in vivo; male; mineralization; mouse model; prevent; promoter; public health relevance; recombinase; repaired; soft tissue; sperm cell; transcription factor

DESCRIPTION (provided by applicant): The proposed studies will use a mouse model to investigate causes for two conditions, infertility and arthropathy. Infertility affects ~ 10% of couples, with about half of this related to male infertility. Abnormal formation of bone-like substances in tissues that are normally not calcified (arthropathy) interferes with tissue function limits motion, and is often painful. Forms of ectopic calcification range from heterotopic ossification (occurs with aging, following trauma, and in genetic and idiopathic forms), to age-related aortic stenosis, and aortic valve calcification (which is a prominent complication limiting surgical repair of the heart and cardiac valves). Understanding the process of ectopic calcification thus has significant potential therapeutic benefit. Mice with disruption of two genes involved in circadian rhythm generation, CLOCK and NPAS2, not only lose their circadian rhythms, but they are also infertile and developed a severe, age-related and progressive ectopic calcification. CLOCK and NPAS2 normally partner with BMAL1 in regulating circadian rhythms; BMAL1- deficient mice are also have reduced fertility and progressive arthropathy. These pathological phenotypes do not occur in animals lacking circadian rhythms due to other genetic lesions, indicating that it is disruption of the CLOCK/NPAS2:BMAL1 transcriptional complexes, likely unrelated to circadian clock function, that allows these pathologies to develop. Studies of animals with gene disruption throughout the body are complicated by the multiple potential sites of action of these genes, ranging from potential effects on reproductive endocrinology and behavior to defects in spermatogenesis (for infertility), and multiple potential cell types of the musculo-skeletal system (for arthropathy). We will thus use advanced genetics approaches to produce tissue-specific genetic lesions, to identify and isolate the cell types involved in fertiliy regulation and in arthropathy prevention, allowing assessment of the molecular events that lead to the pathological conditions. Our overall hypothesis is that local dysregulation of gene expression in animals lacking CLOCK and NPAS2 (or BMAL1) allows these pathological conditions (calcification in specialized but poorly defined cell types at the junction of cartilage and tendon to bone, and infertility). Our overall objective is to define the cell types involved using an unbiased, genetic approach in vivo, and then use these same approaches to label the cell types involved and to isolate them for molecular analysis at times when the genes are critically important for preventing these phenotypes. Identification of the cellular and molecular events occurring in this very reproducible, genetic model in mice should contribute to understanding and ultimately regulating fertility, and for understanding and ultimately preventing pathological ectopic calcification in humans.

描述（由申请方提供）：拟定的研究将使用小鼠模型来研究两种疾病（不孕症和关节病）的原因。不孕症影响约10%的夫妇，其中约一半与男性不育有关。骨样物质在通常不钙化的组织中的异常形成（关节病）干扰组织功能，限制运动，并且通常是疼痛的。异位钙化的形式包括异位骨化（随着年龄的增长、创伤后以及遗传和特发性形式发生）、年龄相关的主动脉瓣狭窄和主动脉瓣钙化（这是一种显著的并发症， 心脏和心脏瓣膜的外科修复）。因此，了解异位钙化的过程具有显著的潜在治疗益处。两个基因缺失的小鼠 参与昼夜节律产生的CLOCK和NPAS 2不仅失去了它们的昼夜节律，而且它们也是不育的，并发展了严重的、与年龄相关的和进行性的异位钙化。CLOCK和NPAS 2通常与BMAL 1一起调节昼夜节律; BMAL 1缺陷小鼠也具有降低的生育力和进行性关节病。这些病理表型不会发生在由于其他遗传病变而缺乏昼夜节律的动物中，这表明CLOCK/NPAS 2：BMAL 1转录复合物的破坏（可能与昼夜节律钟功能无关）使这些病理得以发展。对全身基因破坏的动物的研究由于这些基因的多个潜在作用位点而变得复杂，范围从对生殖内分泌和行为的潜在影响到精子发生的缺陷（不育）以及肌肉骨骼系统的多种潜在细胞类型（关节病）。因此，我们将使用先进的遗传学方法来产生组织特异性遗传病变，以识别和分离参与受精调节和关节病预防的细胞类型，从而评估导致病理条件的分子事件。我们的总体假设是，缺乏CLOCK和NPAS 2（或BMAL 1）的动物中基因表达的局部失调导致了这些病理状况（软骨交界处专门但定义不清的细胞类型中的钙化） 和肌腱到骨，以及不育）。我们的总体目标是在体内使用无偏倚的遗传方法来定义所涉及的细胞类型，然后使用这些相同的方法来标记所涉及的细胞类型，并在基因对于预防这些表型至关重要时将其分离用于分子分析。在小鼠的这种非常可重复的遗传模型中发生的细胞和分子事件的鉴定应有助于理解并最终调节生育能力，并有助于理解并最终预防人类的病理性异位钙化。