Fracture Repair in Aging: Identifying networks by miRNA and mRNA co-sequencing

衰老中的骨折修复：通过 miRNA 和 mRNA 共测序识别网络

• 批准号: 10475273
• 负责人: Anne M Delany
• 金额: $ 20.5万
• 依托单位: UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-01 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10475273
• 关键词: Aging; Biology; Bone Regeneration; Cell Lineage; Cells; Chondrocytes; Clinical; Complex; Defect; Development; Economic Burden; Elderly; Epidermal Growth Factor Receptor; Excision; Fracture; Gene Expression; Gene Expression Regulation; Growth Factor; Healthcare Systems; Hematoma; Heterogeneity; Impairment; Individual; Inflammation; Injury; Investigation; Knowledge; Lead; Mediating; Mesenchymal Stem Cells; Messenger RNA; MicroRNAs; Molecular; Mus; Osteoblasts; Osteoporosis; Patients; Periosteal Cell; Periosteum; Platelet-Derived Growth Factor; Play; Population; Post-Transcriptional Regulation; Process; Protocols documentation; Publishing; RNA; Regulatory Pathway; Rest; Risk; Role; Signal Pathway; Skeletal system; Techniques; Therapeutic; Therapeutic Intervention; Time; Tissue-Specific Gene Expression; Tissues; aged; base; bone; bone fracture repair; bone healing; bone repair; cell type; comorbidity; fracture risk; healing; high reward; high risk; interest; medical complication; new technology; new therapeutic target; novel strategies; novel therapeutics; osteoporotic bone; progenitor; programs; regenerative therapy; repaired; response to injury; single-cell RNA sequencing; skeletal; skeletal regeneration; skeletal stem cell; stem; stem cell differentiation; stem cells; transcriptome; transcriptome sequencing; wound

There is an urgent clinical need to develop new bone regenerative therapeutics to treat delayed and non-union fractures, and critical sized bone defects. Moreover, aging results in severely impaired skeletal repair. A critical aspect of fracture healing begins with expansion of periosteal progenitors, which are activated immediately after injury and then differentiate into bone forming osteoblasts and chondrocytes. However, the molecular mechanisms controlling their activation, expansion, and differentiation in response to injury remains limited. microRNA (miRNA) mediated post-transcriptional regulation of gene expression plays a key role in the commitment and differentiation of stem cells toward the osteoblastic or chondrogenic lineages. Since a single miRNA can potentially target hundreds of distinct mRNAs, identifying the gene expression networks controlled by miRNAs continues to represent a significant gap in our understanding of skeletal biology and repair. Differences in miRNA expression can be observed between young and old bone, and likely mediate some of the impact of aging on fracture repair. Non-biased whole transcriptome analyses have revealed important information about gene expression programs active in the resting periosteum and in healing fractures. Studies utilizing RNA extracted from whole bone have documented changes in either mRNA or miRNAs in these tissues. However, single cell RNA sequencing studies readily demonstrate the cellular heterogeneity present in periosteum and in regenerating bone, making less clear the interpretation of bulk RNAseq studies in these complex tissues. The ability to observe coordinated mRNA-miRNA networks in individual cells during fracture repair will provide paradigm- shifting information that could be used to enhance repair. Further, local delivery of miRNA-based therapeutics represents an exciting novel strategy for facilitating bone healing in the elderly. A thorough understanding of the mRNA-miRNA networks active in each cell type in the healing bone will provide critical rationale for miRNAs to target in a therapeutic setting. We propose to perform single cell mRNA-miRNA co-sequencing on non-hematopoietic lineage cells isolated from resting periosteum and periosteum activated by fracture, comparing young and aged mice. This will allow us to identify expression-based cell subpopulations and mRNA-miRNA interaction networks within those populations, and how these are modulated during fracture repair and in aging. Protocols for single cell miRNA- mRNA co-sequencing were only recently developed. The complex micro-environment of fracture repair is an ideal application for this new technology, which will fill a significant void in our understanding of skeletal biology and provide a rationale for further investigation of miRNA-based therapeutics for bone repair.

临床上迫切需要开发新的骨再生疗法来治疗延迟愈合和骨不连。 骨折和严重的骨缺损。此外，衰老会导致骨骼修复严重受损。一位批评者 骨折愈合始于骨膜祖细胞的扩张，骨膜祖细胞立即被激活。 损伤后分化为成骨细胞和软骨细胞。然而，分子 控制它们的激活、扩张和分化以应对损伤的机制仍然有限。 MicroRNA(MiRNA)介导的基因转录后调控在基因表达调控中起着关键作用。 干细胞向成骨细胞或软骨细胞的定向和分化。因为一首单曲 MiRNA可能针对数百个不同的mRNAs，识别受控制的基因表达网络 通过miRNAs仍然代表着我们对骨骼生物学和修复的理解方面的一个重大差距。 在新骨和老年骨之间可以观察到miRNA表达的差异，并可能介导一些 衰老对骨折修复的影响。 无偏倚的全转录组分析揭示了有关基因表达的重要信息 活跃在静息的骨膜和骨折愈合中的程序。利用从黄瓜中提取的RNA的研究 骨骼已经证明这些组织中的mRNA或miRNAs发生了变化。然而，单细胞RNA 测序研究很容易证明存在于骨膜和再生中的细胞异质性。 在这些复杂的组织中，对大量RNAseq研究的解释不那么清楚。有能力 观察骨折修复过程中单个细胞中协调的mRNA-miRNA网络将提供范例- 转移可以用来加强修复的信息。此外，基于miRNA的治疗药物的本地交付 代表了一种促进老年人骨骼愈合的令人兴奋的新策略。透彻地了解 修复骨中每种细胞类型中活跃的mrna-miRNA网络将为 在治疗环境中靶向miRNAs。 我们建议对分离的非造血系细胞进行单细胞mRNAmiRNA共测序。 从静息骨膜和骨折激活的骨膜，比较幼龄和老龄小鼠。这将允许 美国将确定基于表达的细胞亚群和其中的mRNA-miRNA相互作用网络 以及在骨折修复和衰老过程中这些因素是如何被调节的。单细胞miRNA的协议- 信使核糖核酸共测序是最近才发展起来的。骨折修复的复杂微环境是一种 这项新技术的理想应用，将填补我们对骨骼生物学理解的一个重大空白 为进一步研究基于miRNA的骨修复疗法提供了理论基础。