Extracellular Matrix Vesicles and Intercellular Communication of Osteoblast-lineage Cells

成骨细胞谱系细胞的细胞外基质囊泡和细胞间通讯

• 批准号: 10679255
• 负责人: Anne Michelle Skelton
• 金额: $ 4.43万
• 依托单位: VIRGINIA COMMONWEALTH UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-06-01 至 2027-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10679255
• 关键词: Adult; Alkaline Phosphatase; Autologous; Binding; Biological; Biological Assay; Bone Development; Bone Marrow; Bone Regeneration; Bone Transplantation; Cartilage; Cartilage Matrix; Cell Lineage; Cells; Characteristics; Chemicals; Chondrocytes; Chronic; Coculture Techniques; Collagen Fibril; Collagen Type I; Culture Media; Defect; Deposition; Differentiation Antigens; Embryo; Enzymes; Epiphysial cartilage; Experimental Models; Extracellular Matrix; Femur; Foundations; Fracture; Goals; Human body; Hybrid Cells; Hydrogels; Hydroxyapatites; In Vitro; Injectable; Integrin Binding; Lipids; Literature; Luciferases; Mature Bone; Measures; Mediating; Mediation; Membrane; MicroRNAs; Minerals; Modeling; Morphology; Mus; Nucleic Acids; Operative Surgical Procedures; Organelles; Orthopedics; Osteoblasts; Osteocalcin; Osteogenesis; Pathology; Pathway interactions; Patients; Pattern; Phenotype; Phospholipids; Physiologic Ossification; Physiologic calcification; Play; Population; Population Heterogeneity; Process; Proliferating; Property; Proteins; Regenerative Medicine; Research; Role; Signal Pathway; Signal Transduction; Signaling Molecule; Site; Stromal Cells; System; Testing; Therapeutic; Tissues; Translating; Transmission Electron Microscopy; Trypsin; Untranslated RNA; Vesicle; Western Blotting; body system; bone; bone fracture repair; bone healing; bone repair; calcification; calcium phosphate; exosome; extracellular vesicles; fetal; healing; improved; in vivo; in vivo Model; intercellular communication; long bone; mRNA Expression; microCT; mineralization; nanoparticle; nanosized; novel therapeutics; osteoblast differentiation; osteogenic; protein expression; regenerative therapy; repaired; response; standard care; success; therapeutic evaluation; vesicular release

PROJECT SUMMARY Autologous bone grafting, the current gold standard for treating critical sized bone defects and chronic nonunions, has limited success, leaving a need for adjunct treatments to contribute to overall bone healing. Extracellular vesicles, of which there are many types, are a promising new cell-free, membrane-bound therapeutic in the field of regenerative medicine. Specific to the field of bone regeneration and repair are matrix vesicles; small extracellular vesicles released by mineralization-competent cells that become anchored in the extracellular matrix and are a necessary component of mineralized bone formation via endochondral ossification. They are involved in cellular signaling via small, non-coding microRNA in the growth plate, suggesting that they may play a similar role in bone. Previous research indicates that chondrocyte-derived matrix vesicles are enriched with microRNA and other cell signaling molecules that contribute to their ability to influence proliferation and differentiation of target cells, however the role and mechanism of action of osteoblast-derived matrix vesicles, particularly in bone as opposed to the growth plate, remains to be elucidated. With this information, the biological mediation of bone development and regeneration that matrix vesicles provide may translate into novel therapeutic potential for orthopedic pathologies, including critical size bone defects and fracture nonunions. Therefore, we hypothesize that matrix vesicles produced by osteoblast-lineage cells, as a specific subset of extracellular vesicle, use their microRNA cargo to enhance osteogenic differentiation and proliferation, drive osteogenesis, and aid in bone defect healing. First, we aim to determine the relationship between osteoblast- derived matrix vesicles and another class of extracellular vesicle, the exosome. Next, we aim to determine the pathways targeted by matrix vesicle microRNA cargo. Finally, we aim to determine the extent to which osteoblast-derived matrix vesicles modulate osteogenesis and bone defect healing. To test these aims, we will use osteoblast-lineage cells to derive matrix vesicles and characterize protein expression, size, and morphology. We will also use in vitro experimental models to assess pathway involvement and evaluate co-culture response with osteoblast-like cells. To test the therapeutic potential, we will use a translatable in vivo model of a mouse long bone defect and a biorthogonal injectable hydrogel to deliver matrix vesicles. We expect to find that osteoblast-derived matrix vesicles are a specialized subclass of extracellular vesicle with microRNA cargo that targets the canonical Wnt pathway to activate cellular signaling and leads to increased osteoblastic differentiation of target cells when co-cultured in vitro. Our in vivo model is expected to demonstrate improved healing upon treatment with hydrogel-delivered matrix vesicles. All of which together demonstrates the role matrix vesicles play in the coordinated effort to form new bone and their viability as a therapeutic option to improve bone healing.

项目摘要 自体骨移植，目前治疗严重骨缺损和慢性骨缺损的金标准 骨不连的治疗效果有限，因此需要辅助治疗来促进整体骨愈合。 细胞外囊泡是一种很有前途的新型无细胞、膜结合囊泡， 在再生医学领域的治疗。具体到骨再生和修复领域的是基质 囊泡;小的细胞外囊泡释放的矿化能力的细胞，成为锚定在 细胞外基质，并且是通过软骨内骨化形成矿化骨的必要成分。 它们通过生长板中的小的非编码microRNA参与细胞信号传导，这表明它们 可能在骨骼中起着类似的作用。以前的研究表明，软骨细胞衍生的基质囊泡是 富含microRNA和其他细胞信号分子，有助于它们影响增殖的能力 然而，成骨细胞源性基质的作用和作用机制 囊泡，特别是骨中的囊泡，而不是生长板中的囊泡，仍有待阐明。有了这些信息， 基质囊泡提供的骨发育和再生的生物介导可以转化为新的骨发育和再生。 骨科疾病的治疗潜力，包括临界尺寸骨缺损和骨折不愈合。 因此，我们假设成骨细胞系细胞产生的基质囊泡，作为成骨细胞的一个特定亚群， 细胞外囊泡，使用它们的microRNA货物来增强成骨分化和增殖，驱动 骨生成，并有助于骨缺损愈合。首先，我们的目标是确定成骨细胞- 衍生的基质囊泡和另一类细胞外囊泡，外泌体。接下来，我们的目标是确定 基质囊泡microRNA货物靶向的途径。最后，我们的目标是确定在多大程度上， 成骨细胞衍生的基质囊泡调节骨生成和骨缺损愈合。为了验证这些目标，我们将 使用成骨细胞系细胞衍生基质囊泡并表征蛋白质表达、大小和形态。 我们还将使用体外实验模型来评估通路参与和评估共培养反应 成骨细胞样细胞。为了测试治疗潜力，我们将使用可翻译的小鼠体内模型 长骨缺损和双正交可注射水凝胶以递送基质囊泡。我们希望能发现 成骨细胞来源的基质囊泡是具有微小RNA货物的细胞外囊泡的特化亚类， 靶向经典Wnt通路，激活细胞信号传导，导致成骨细胞分化增加 的靶细胞在体外共培养时。我们的体内模型预计将显示出改善的愈合后， 用水凝胶递送的基质囊泡治疗。所有这些共同证明了基质囊泡 在形成新骨的协调努力中发挥作用，并将其作为改善骨愈合的治疗选择。