Molecular control of chondrocyte hypertrophy: an evolutionary approach

软骨细胞肥大的分子控制：一种进化方法

• 批准号: 10606678
• 负责人: Michael Palmer
• 金额: $ 6.95万
• 依托单位: MARINE BIOLOGICAL LABORATORY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2026-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10606678
• 关键词: ATAC-seq; Adult; Age; Biological; Biological Models; CRISPR/Cas technology; Cartilage; Cartilage injury; Cell Therapy; Cells; Chondrichthyes; Chondrocytes; Collagen Fiber; Collagen Type I; Collagen Type II; Degenerative polyarthritis; Development; Elderly; Embryonic Development; Engineering; Enhancers; Evolution; Fibrocartilages; Fishes; Frequencies; Friction; Gene Expression; Gene Transfer Techniques; Genes; Genetic Transcription; Genomic approach; Genomics; Growth; Health; Human; Hyaline Cartilage; Hypertrophy; Implant; In Vitro; Individual; Inferior; Injury; Joints; Knee; Laboratories; Left; Lesion; Life; Mammals; Mechanics; Mesenchymal; Mesenchymal Stem Cells; Minor; Molecular; Nature; Organism; Overweight; Pain; Pathology; Patients; Phenotype; Physiologic Ossification; Population; Prevalence; Process; Property; Quality of life; Regulator Genes; Research; Resources; Shark; Site; Skeletal Development; Skeleton; Stable Populations; Structure; Techniques; Testing; Thinness; Time; Tissue Engineering; Tissues; Transplantation; Untranslated RNA; Variant; Vertebrates; Weight-Bearing state; articular cartilage; bone; bone loss; calcification; cartilage cell; cartilage development; cartilaginous; comparative genomics; differential expression; embryo tissue; gene discovery; genome annotation; genome editing; healing; implantation; improved; in vitro Model; induced pluripotent stem cell; marine; novel; novel strategies; osteogenic; repaired; single-cell RNA sequencing; skeletal; skeletogenesis; stem cell based approach; stem cells; treatment strategy

Project Summary In mammals, cartilage is predominantly an embryonic tissue: the vast majority of cartilage is replaced by bone during the processes of hypertrophy and ossification, with cartilage persisting in relatively few places within the adult skeleton (e.g. in joints, as articular cartilage). Articular cartilage is an aneural, avascular tissue with very limited capacity for spontaneous repair, hence the prevalence in humans of cartilage pathologies like osteoarthritis. In contrast, cartilaginous fishes (sharks, skates, and rays) have undergone an evolutionary loss of bone, instead possessing a skeleton that is composed entirely of pre-hypertrophic cartilage, and that remains cartilaginous throughout life. Understanding how cartilaginous fishes arrest skeletal development prior to chondrocyte hypertrophy will shed new light on transcriptional features that could be employed for in vitro engineering of stable chondrocytes for the treatment of human cartilage injuries. Stem cell-based approaches for the treatment of articular cartilage injuries are currently hindered by the relative instability of mammalian cartilage cells (chondrocytes) in vitro and upon implantation into an injury site. In this project, the molecular development of cartilage in the little skate (Leucoraja erinacea) will be studied to discover gene expression correlates of their permanent cartilaginous skeleton. This project will begin with the use of single-cell RNA-sequencing to identify genes that are differentially expressed between developing and differentiated skate and mammalian chondrocytes (Aim 1), and that may underlie arrest prior to hypertrophy in the former. ATAC-seq and comparative genomic approaches will then be used to test for variation in non-coding regions (i.e. putative enhancers) that might account for divergent gene expression during skate and mammalian skeletogeneis (Aim 2). Finally, skate-inspired molecular manipulations (CRISPR/Cas9 genome editing and/or transgenesis) will be incorporated in an in vitro model of mammalian skeletogenesis, in order to achieve a permanent and stable chondrocyte cell state from mammalian mesenchymal progenitors (Aim 3). This project will capitalize on the unique properties of the skate skeleton, and on the biological resources, facilities, and expertise that are available at the Marine Biological Laboratory in Woods Hole. By taking an evolution-inspired tissue engineering approach, this project will contribute to the development of novel in vitro techniques for cartilage engineering and for the treatment of mammalian skeletal pathologies.

项目摘要 在哺乳动物中，软骨主要是胚胎组织：绝大多数软骨被骨骼取代 在肥大和骨化过程中，软骨持续在相对较少的地方 成人骨骼（例如关节，作为关节软骨）。关节软骨是一种胸腔，无血管组织 自发修复的能力有限，因此软骨病理的人类流行率 骨关节炎。相反，软骨鱼（鲨鱼，溜冰和射线）经历了进化损失 骨头，而是拥有完全由腹部前软骨组成的骨骼，并且 一生仍然是软骨。了解软骨鱼如何阻止骨骼发育 到软骨细胞肥大将为体外使用的转录特征提供新的启示 稳定软骨细胞的工程治疗人体软骨损伤。 目前，基于干细胞的基于干细胞的关节软骨损伤方法受到 哺乳动物软骨细胞（软骨细胞）的相对不稳定在体外和植入损伤部位后。 在这个项目中，将研究小滑板（Leucoraja erinacea）中软骨的分子发育 发现其永久性软骨骨骼的基因表达相关。这个项目将以 使用单细胞RNA测序来识别在之间差异表达的基因 开发和区分滑板和哺乳动物软骨细胞（AIM 1），这可能是逮捕的基础 在前者肥大之前。然后，ATAC-SEQ和比较基因组方法将用于 测试可能解释基因不同的非编码区域（即推定增强子）的变化 滑板和哺乳动物骨架期间的表达（AIM 2）。最后，滑板启发的分子 操纵（CRISPR/CAS9基因组编辑和/或转基因）将纳入体外 哺乳动物骨骼生成的模型，以实现永久稳定的软骨细胞状态 来自哺乳动物间充质祖细胞（AIM 3）。 该项目将利用滑板骨骼的独特特性，以及生物资源， 设施和专业知识可在伍兹孔的海洋生物实验室提供。通过 该项目以进化为灵感的组织工程方法，有助于新的体外发展 软骨工程技术和治疗哺乳动物骨骼病理的技术。