Biomimetic Apoptotic Particles for Macrophage-driven Oral Bone Regeneration

用于巨噬细胞驱动的口腔骨再生的仿生凋亡颗粒

• 批准号: 10596206
• 负责人: Rahasudha Kannan
• 金额: $ 4.28万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-01 至 2024-12-05
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10596206
• 关键词: Actins; Adult; Affect; Age; Aging; Apoptotic; Architecture; Behavior; Biochemical; Biocompatible Materials; Bioinformatics; Biological; Biomedical Engineering; Biomimetics; Biophysics; Bone Regeneration; Bone Transplantation; CCL2 gene; Cell Death; Cells; Coculture Techniques; Cues; Cytoskeleton; Data; Dental; Dental Implants; Dentition; Eligibility Determination; Enzyme-Linked Immunosorbent Assay; Event; Gene Expression Profile; Genes; Genetic; Goals; Histology; Hydrogels; Immune system; Impaired wound healing; Impairment; Implant; In Vitro; Infection; Infiltration; Knockout Mice; Macrophage; Mediating; Mesenchymal Stem Cells; Microspheres; Morbidity - disease rate; Mus; Natural regeneration; Nature; Nuclear; Oral; Oral cavity; Osteoblasts; Osteogenesis; Outcome; Pain; Pathway interactions; Patients; Phagocytosis; Phenotype; Population; Process; Production; Proteins; Quality of life; Quantitative Reverse Transcriptase PCR; Role; Shapes; Signal Pathway; Signal Transduction; Site; System; Trauma; Treatment Cost; Work; Wound models; aging population; bone healing; bone quality; bone repair; clinical translation; clinically relevant; comorbidity; cost; design; experimental group; improved; insight; mechanotransduction; microCT; mimicry; mouse model; novel; osteogenic; particle; permanent tooth; receptor; recruit; regenerative; regenerative therapy; repaired; response; restoration; single-cell RNA sequencing; stem cells; success; tool; wound; wound healing

ABSTRACT Adequate bone quality, quantity, and wound repair in the oral cavity are crucial for treatment eligibility, as well as short and long-term success of dental implants. Successful restoration of a functional dentition requires an understanding of the endogenous bone repair process. Often overlooked, one of the first steps in osseous wound repair after trauma, such as in a dental extraction, is cell death and subsequent apoptotic cell (AC) clearance (efferocytosis) by macrophages. As a result of efferocytosis, macrophages secrete a variety of factors that facilitate regeneration, and swiftly alter their behavior in response to a multitude of physical and biological microenvironmental cues. CC-motif chemokine ligand 2 (CCL2), which is secreted by macrophages, mediates mesenchymal stem/progenitor cell (MSPC) recruitment to the wound site. Although previous work has largely focused on biochemical signals that drive CCL2 production, preliminary data in the current proposal suggests that the physical nature of AC engulfment drives cytoskeletal events and subsequent mechanotransductive signaling. Engulfment of apoptotic cells induces changes in macrophage shape, actin organization, and nuclear architecture that likely initiates CCL2 production. Understanding efferocytosis-induced intracellular forces and resulting signaling will inform the design of apoptotic cell mimics (ACM) as a regenerative therapy for patients whose age or co-morbidities impair wound healing capacity. The goals of this project are to determine the role of efferocytosis-induced macrophage mechanotransduction in bone repair and to promote reparative macrophage behavior using ACM, hence catalyzing the endogenous osseous wound healing response. The overall hypothesis is that macrophages promote osteogenic repair through efferocytosis-driven biophysical signaling, which can be recapitulated using apoptotic cell mimicry for a regenerative advantage. The two aims proposed are: 1) to connect CCL2 expression and macrophage phenotype with AC and ACM engulfment-induced changes to the cyto- and nucleoskeleton, and 2) to optimize and deliver ACM to promote macrophage-driven osteogenesis and improve bone repair in a clinically relevant oral osseous wound healing model. To accomplish these aims, first an in vitro co-culture system that allows for macrophage engulfment of AC and ACM will be used. This will be a valuable tool to identify and validate genes and proteins associated with mechanotransduction and osteogenic-repair that are similarly altered in both experimental groups. ACM with tunable size, stiffness, and degradability will be optimized to maximize CCL2 secretion given its role in MSPC recruitment. Next, we will use CCL/R2 genetic knockout mouse models to confirm the role of CCL2 in bone regeneration. We anticipate ACM treatment will induce cytoskeletal changes that result in CCL2 production and promote bone repair. The outcomes of this project will identify underexplored effects of post- efferocytosis macrophage mechanotransduction and reparative activity, providing mechanistic insights into bone regeneration, which can be used to establish new bone regeneration therapies for patients.

摘要 充足的骨质、数量和口腔伤口修复对于治疗合格性也至关重要 短期和长期成功的牙科植入物。功能性牙列的成功修复需要 了解内源性骨修复过程。经常被忽视的是，骨创伤的第一步 创伤后的修复，如拔牙，是细胞死亡和随后的凋亡细胞（AC）清除 （巨噬细胞）。由于巨噬细胞增多，巨噬细胞分泌多种因子， 促进再生，并迅速改变他们的行为，以应对众多的物理和生物 微环境线索由巨噬细胞分泌的CC基序趋化因子配体2（CCL 2）介导 间充质干/祖细胞（MSPC）募集到伤口部位。虽然以前的工作主要是 目前提案中的初步数据表明， AC吞噬的物理性质驱动细胞骨架事件和随后的机械转导 发信号。凋亡细胞的吞噬诱导巨噬细胞形状、肌动蛋白组织和细胞核的变化。 可能启动CCL 2生产的结构。理解细胞凋亡诱导的细胞内力， 由此产生的信号将为设计凋亡细胞模拟物（ACM）作为患者的再生疗法提供信息 其年龄或合并症损害伤口愈合能力。本项目的目标是确定 骨修复中巨噬细胞介导的机械转导的作用，并促进修复 使用ACM的巨噬细胞行为，因此催化内源性骨创伤愈合反应。的 总体假设是巨噬细胞通过巨噬细胞驱动的成骨修复促进成骨修复， 生物物理信号传导，其可以使用凋亡细胞模拟来概括，用于再生的细胞周期。 优势提出的两个目标是：1）将CCL 2表达和巨噬细胞表型与AC联系起来 和ACM吞噬诱导的细胞骨架和核骨架的变化，以及2）优化和递送ACM， 促进巨噬细胞驱动的骨生成并改善临床相关的口腔骨创伤中的骨修复 治愈模式为了实现这些目标，首先，体外共培养系统，允许巨噬细胞 将使用AC和ACM的吞没。这将是鉴定和验证基因和蛋白质的有价值的工具 与机械传导和成骨修复相关，在两种实验中， 组具有可调尺寸、刚度和可降解性的ACM将被优化以最大化CCL 2分泌， 它在MSPC招聘中的作用。接下来，我们将使用CCL/R2基因敲除小鼠模型来证实CCL/R2基因的作用。 CCL 2在骨再生中的作用我们预计ACM处理将诱导细胞骨架的变化，导致CCL 2 生产和促进骨修复。该项目的成果将确定未充分探索的后 巨噬细胞巨噬细胞机械转导和修复活性，提供对骨的机械见解 再生，可用于为患者建立新的骨再生疗法。