Spatial and Temporal Role of the Runx3 Transcription Factor in Secondary Fracture Healing

Runx3 转录因子在二次骨折愈合中的时空作用

• 批准号: 10454763
• 负责人: HICHAM M DRISSI
• 金额: --
• 依托单位: VETERANS HEALTH ADMINISTRATION
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-01 至 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10454763
• 关键词: Age; Aging; Biomechanics; Blood Vessels; Blood capillaries; Bone Tissue; Bone callus; Cartilage; Cell Differentiation process; Cell Lineage; Cells; Chondrocytes; Clinical; Data; Diabetes Mellitus; Economic Burden; Encapsulated; Evaluation; Event; Femoral Fractures; Flow Cytometry; Fracture; Frequencies; Gelatin; Healthcare Systems; Histologic; Histology; Hydrogels; Image; Impairment; Incidence; Knockout Mice; LoxP-flanked allele; Mediating; Mesenchymal; MicroRNAs; Modality; Modeling; Molecular; Molecular Analysis; Molecular Target; Monitor; Mus; Names; Osteoblasts; Osteocytes; Osteogenesis; Osteoporosis; Osteoporotic; Pathway interactions; Patients; Periosteal Cell; Periosteum; Pharmaceutical Preparations; Population; Process; RNA; RUNX3 gene; Regulator Genes; Regulatory Pathway; Reporter; Reporting; Repression; Roentgen Rays; Role; Site; Smoking; Sorting - Cell Movement; Tamoxifen; Testing; Time; Tissues; Torsion; Transgenic Mice; Translational Research; Vascular blood supply; Veterans; Work; angiogenesis; bone; bone fracture repair; bone healing; bone imaging; bone mass; bone repair; cartilaginous; clinically relevant; comorbidity; conditional knockout; experimental study; healing; ineffective therapies; laser capture microdissection; long bone; lost work time; microCT; multiphoton microscopy; nanofiber; new therapeutic target; novel; novel therapeutic intervention; novel therapeutics; osteogenic; osteoporosis with pathological fracture; pre-clinical; prevent; progenitor; skeletal stem cell; stem cells; success; transcription factor; two photon microscopy

Clinical Dilemma: The frequency of impaired fracture healing is increased with aging as well as in the presence of other patient-related factors such as smoking, osteoporosis, and diabetes. Treatment of fractures in this setting continues to pose a significant economic burden on the US healthcare system due to increases in time lost from work as well as increases in the expenses associated with fracture-associated complications. While various bone anabolic drugs are successful in increasing homeostatic bone mass in osteoporotic patients and decreasing fracture incidence, they have not demonstrated significant success in enhancing fracture repair. Therefore, identifying novel molecular targets to accelerate secondary fracture healing in this very common setting remains of paramount importance. Relevance to the VA: According to the Office of VA Inspector General report in 2010, osteoporotic patients who suffered a single fracture present a higher incidence of subsequent fractures (20-fold increase) than unaffected populations. Impaired or delayed bony union following fracture of long bones prevents or delays a significant percentage of VA patients from resuming their daily activities and returning to work. Ineffective treatment of these fractures maximizes the economic burden on the VA healthcare system. Identifying novel molecular targets to enhance secondary bone repair remains of paramount importance. The objective of this translational research application is to enhance secondary fracture healing by targeting novel regulatory pathways that enhance periosteal cell-induced osteogenesis and angiogenesis during fracture callus formation. Scientific premise: We provide compelling preliminary evidence of the following: 1. Runx3 is expressed in chondrocytes, osteoblasts and osteocytes of C57BL6j murine long bones. 2. Runx3 expression levels are increased in soft cartilaginous calluses and subsequently decreased in bony calluses of murine femoral fractures. 3. Conditional deletion of Runx3 in periosteal cells (cKO) resulted in enhanced secondary bone healing as evidenced by histological, histomorphometric, and molecular analyses. 4. The cellular mechanisms underlying these positive effects on secondary bone healing implicate increased osteogenesis as well as angiogenesis of fractured periosteal cells from Runx3 cKO compared to control mice. 5. Use of multiphoton microscopy demonstrate the feasibility of tracking Prx1+ skeletal progenitor cells during bone repair and longitudinally monitor the bone healing process for lineage tracing experiments. Here we hypothesize that Runx3 is a molecular switch that controls the transition from cartilaginous to bony callus, and its deletion in the chondrogenic cell lineage will accelerate secondary fracture healing. To verify this hypothesis, we propose to first establish the effects of stage-specific repression of Runx3 on secondary fracture healing (Aim 1). We will then determine the mechanisms via which Runx3 controls mesenchymal cell differentiation into the chondro/osteogenic lineages (Aim 2). Finally, we will assess the efficacy of Runx3 inhibition during fracture repair in control C57BL6 mice through controlled and sustained delivery of miRNA encapsulated hydrogel and examine the rate of bone healing and biomechanical strength of healed bone (Aim 3). Impact: Defining the pathways that governthe transition from soft to bony callus will help identify new therapies to accelerate secondary fracture healing. Here, we will establish Runx3 as a novel therapeutic target.

临床困境：骨折愈合受损的频率随着年龄的增长而增加 存在其他与患者相关的因素，如吸烟、骨质疏松症和糖尿病。骨折的治疗 在这种情况下，继续对美国医疗保健系统构成重大经济负担的原因是 工作时间的损失以及与骨折相关并发症相关费用的增加。 而各种骨合成代谢药物在增加骨质疏松症患者体内平衡骨量方面取得了成功 患者和降低骨折发生率方面，他们并没有表现出显著的成功 骨折修复。因此，寻找新的分子靶点以促进本病的继发性骨折愈合 非常常见的环境仍然是至关重要的。 与退伍军人事务部的相关性：根据退伍军人事务部2010年的报告，骨质疏松症患者 患有一次骨折的人继发骨折的发生率(20倍)高于 未受影响的人群。长骨骨折后受损或延迟的骨愈合阻止或延迟了 相当大比例的VA患者无法恢复日常活动并重返工作岗位。效果不佳 这些骨折的治疗最大限度地增加了退伍军人管理局医疗系统的经济负担。辨别小说 促进二次骨修复的分子靶点仍然是至关重要的。这样做的目的是 翻译研究的应用是通过靶向新的调控来促进继发性骨折愈合 在骨折骨痂形成过程中促进骨膜细胞诱导的成骨和血管生成的途径。 科学前提：我们提供了令人信服的以下初步证据： 1.RUNX3在C57BL6j小鼠长骨软骨细胞、成骨细胞和骨细胞中均有表达。 2.RUNX3在软骨性骨痂中的表达水平升高，随后在骨痂中的表达水平降低 小鼠股骨骨折的骨痂。 3.骨膜细胞(CKO)中Runx3的条件性缺失可促进二次骨愈合 经组织学、组织形态计量学和分子分析证实。 4.这些对继发性骨愈合的积极作用背后的细胞机制可能增加 Runx3CKO骨折后骨膜细胞的成骨和血管生成与对照比较 老鼠。 5.使用多光子显微镜证实了追踪PRX1+骨骼祖细胞的可行性 骨修复和纵向监测骨愈合过程的谱系追踪实验。 这里我们假设Runx3是一个分子开关，控制着从软骨到软骨的转变 骨痂及其在软骨细胞谱系中的缺失将加速继发性骨折 治愈。 为了验证这一假设，我们建议首先建立阶段特异性抑制Runx3对 二次骨折愈合(目标1)。然后我们将确定Runx3控制的机制 间充质细胞分化为软骨/成骨细胞(目标2)。最后，我们将评估 对照C57BL6小鼠骨折修复过程中抑制Runx3的作用 将miRNA包裹的水凝胶植入体内，检测其骨愈合率和生物力学强度。 骨愈合(目标3)。 影响：确定控制从软骨痂向骨痂过渡的途径将有助于确定新的治疗方法 以加速二次骨折愈合。在这里，我们将建立Runx3作为一个新的治疗靶点。