Assessing lineage decisions of musculoskeletal progenitor cells with aging

评估肌肉骨骼祖细胞随衰老的谱系决定

• 批准号: 7645172
• 负责人: David W. Rowe
• 金额: $ 37.18万
• 依托单位: UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-09-15 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7645172
• 关键词: Adipocytes; Adult; Affect; Age; Aging; Animals; Appearance; Back; Bioinformatics; Bone Marrow; Breeding; Bromodeoxyuridine; Cartilage; Cell Culture Techniques; Cell physiology; Cells; Characteristics; Chromosome Mapping; Clinical; Color; Commit; Deterioration; Development; Environment; Event; Fatty acid glycerol esters; Fluorescence; Foundations; Frail Elderly; Frail Older Adults; Future; Generations; Genes; Genetic; Genetic Models; Genotype; Grant; Health; Immunohistochemistry; In Situ; In Vitro; Inflammatory; Invaded; Label; Mature Bone; Mediator of activation protein; Microarray Analysis; Microscopy; Modeling; Molecular; Mus; Muscle; Muscle Cells; Muscle Fibers; Musculoskeletal; Musculoskeletal System; Myofibroblast; Osteoblasts; Osteogenesis; Osteoporosis; Pathway interactions; Pattern; Phenotype; Population; Primary Cell Cultures; Process; Production; Quantitative Trait Loci; Regulation; Relative (related person); Reporter; Research Personnel; Resistance; Signal Transduction; Skeletal Muscle; Smooth Muscle; Smooth Muscle Actin Staining Method; Speed; Stem cells; Structure; Techniques; Testing; Time; Tissues; Transcription Factor AP-2 Alpha; Transplantation; Validation; age related; aged; base; bone; cell type; congenic; cytokine; design; digital; fiber cell; fluorescence imaging; functional loss; in vivo; indexing; lipid biosynthesis; muscle degeneration; muscle strength; myogenesis; novel strategies; osteogenic; progenitor; programs; research study; sarcopenia; second harmonic; skeletal; time use; tool

DESCRIPTION (provided by applicant): The deterioration of the musculoskeletal system with age that leads to sarcopenia and osteoporosis is a multi-factorial reality that has significant clinical limitations for muscle strength, mobility and loss of functional independence in frail older adults. Murine models may provide a window into the environmental and genetic factors that contribute to the process. This grant will test two independent yet converging hypothesis: one model based on lineage regulation that focuses on a common progenitor cell with the ability to develop into the myogenic, osteogenic or adipogenic lineages, and a second model in which invading adipocytes adversely influence the immunological and contractile status of mature muscle tissue. The first model envisions that age-related changes in gene activity result in a disruption of lineage progression from its primary or default direction (bone and muscle) to a secondary alternative direction (fat). To test this model, GFP-reporter constructs that mark specific levels of cellular differentiation (Col3.6, bone; AP2, fat; myoD, muscle; smooth muscle actin.SMA, myofibroblast) will be introduced in to C57BI/6 (affected) and 129 P3/J (resistant) mice and the animals will be aged to 10 and 24 months. By combining reporter constructs with complementary colors, the ability of the lineage to progress from a progenitor to fully mature bone or muscle cells, or the branch point of the progenitor to select the primary or adipocytic lineage will be assessed in a primary cell culture model. The tempo and extent of differentiation will be assess in real time using repetitive fluorescence imaging of the same culture well, and the proportion of cells at a defined levels of development will be obtained by FAC analysis. In vitro analysis will suggest levels of differentiation that are particularly sensitive to genetic and age-related factors and the potential genetic mediators of the effect will be examined by microarray analysis of the FAC isolated cell populations. In vivo validation of the culture results will be performed by progenitor transplantation experiments into young and aged hosts, and by a BrdU labeling study that assesses lineage choices of the endogenous progenitor cells. To test the second model of in-situ tissue disruption by adipocytes, the two experimental groups will be phenotyped for appearance of GFP positive fat cells in bone and muscle and for a spatial correlation of adipocytes with disorganized microstructure within muscle fiber cells. The tools, techniques and molecular concepts that are developed in this grant will provide a foundation for future QTL studies in mice designed to map genetic factors that contribute to age-related changes in skeletal health.

描述（由申请人提供）：肌肉骨骼系统随着年龄的增长而退化，导致肌肉减少症和骨质疏松症，这是一个多因素的现实，对体弱的老年人的肌肉力量、活动能力和功能独立性的丧失具有显着的临床限制。小鼠模型可能为了解促成这一过程的环境和遗传因素提供了一个窗口。这笔赠款将测试两个独立但趋同的假设：一种基于谱系调节的模型，重点关注具有发育成肌源性、成骨性或脂肪性谱系的能力的共同祖细胞，以及第二种模型，其中入侵的脂肪细胞对免疫和脂肪细胞产生不利影响。成熟肌肉组织的收缩状态。第一个模型设想，与年龄相关的基因活性变化会导致谱系进程从主要或默认方向（骨骼和肌肉）到次要替代方向（脂肪）的破坏。为了测试该模型，标记特定细胞分化水平的 GFP 报告基因构建体（Col3.6，骨；AP2，脂肪；myoD，肌肉；平滑肌肌动蛋白。SMA，肌成纤维细胞）将被引入 C57BI/6（受影响）和129只P3/J（抗性）小鼠，这些动物的年龄将达到10个月和24个月。通过将报告构建体与互补色相结合，将在原代细胞培养模型中评估谱系从祖细胞进展到完全成熟的骨或肌肉细胞的能力，或祖细胞选择原代或脂肪细胞谱系的分支点。将使用同一培养孔的重复荧光成像实时评估分化的速度和程度，并通过 FAC 分析获得处于特定发育水平的细胞比例。体外分析将表明对遗传和年龄相关因素特别敏感的分化水平，并且将通过 FAC 分离细胞群的微阵列分析来检查该效应的潜在遗传介质。培养结果的体内验证将通过年轻和老年宿主的祖细胞移植实验以及评估内源祖细胞谱系选择的 BrdU 标记研究来进行。为了测试脂肪细胞原位组织破坏的第二种模型，将对两个实验组进行表型分析，以确定骨骼和肌肉中 GFP 阳性脂肪细胞的出现，以及脂肪细胞与肌纤维细胞内紊乱的微观结构的空间相关性。这笔资助中开发的工具、技术和分子概念将为未来的小鼠 QTL 研究奠定基础，这些研究旨在绘制导致骨骼健康年龄相关变化的遗传因素。