Genetic regulation of the cartilage callus during large-scale bone repair

大规模骨修复过程中软骨愈伤组织的基因调控

• 批准号: 9996526
• 负责人: Francesca V Mariani
• 金额: $ 47.02万
• 依托单位: UNIVERSITY OF SOUTHERN CALIFORNIA
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-15 至 2023-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9996526
• 关键词: Ablation; Adopted; Adult; Agonist; Biological Assay; Body part; Bone Injury; Bone Marrow; Bone Regeneration; Bone callus; Cartilage; Cells; Chondrocytes; Chromatin; Clinical; Connective Tissue; Data; Defect; Development; Engraftment; Enhancers; Epiphysial cartilage; Erinaceidae; Failure; Femur; Fishes; Future; Genes; Genetic; Genomics; Goals; Homeostasis; Human; Injury; Jaw; Knock-in; Label; LacZ Genes; Lesion; Ligands; Maintenance; Mediating; Mesenchymal Stem Cells; Minerals; Modeling; Morphology; Mus; Natural regeneration; Orthopedics; Osteoblasts; Osteocytes; Pathway interactions; Periosteal Cell; Periosteum; Population; Property; Proteins; Regenerative capacity; Regulation; Role; Signal Transduction; Source; Surgeon; Testing; Therapeutic; Thick; Transgenic Organisms; Zebrafish; bone; bone healing; cartilage cell; cartilage repair; cell type; combinatorial; epigenetic memory; healing; improved; mutant; novel; osteoblast differentiation; osteogenic; preclinical trial; progenitor; receptor; repaired; response to injury; rib bone structure; skeletal; stem cell population; stem cells; transcriptome

The repair of extensive bone injuries remains an unmet clinical challenge. By developing two new and complementary models of large-scale bone regeneration in zebrafish and mouse, we aim to understand the role of the cartilage callus in generating large segments of full thickness bone. While the periosteum generates osteoblasts during homeostasis, the specific subpopulation that builds the repair callus remains poorly defined. Using transgenic lineage tracing, we provide compelling preliminary evidence that a rare bi-potent Sox9+/Runx2+ periosteal population generates new cartilage and bone during repair. This newfound ability to label, manipulate, and isolate a specific stem cell population allows us to test whether the remarkable regenerative capacity of the rib is due to the unique properties of its periosteal stem cells. In Aim 1, we team up with an orthopaedic surgeon to test that Sox9+ cells from the rib periosteum can be expanded in culture and used to heal a critical-sized femoral defect. The formation of a cartilage callus is a common feature in bone repair, yet how the periosteum generates cartilage only during repair remains a mystery. In preliminary data, we find that the cartilage callus is severely compromised when either the Ihha ligand is deleted in zebrafish or the Hh receptor Smo is deleted from Sox9+ cells in mice. In Aim 2, we test that this reflects a repair-specific role for Ihh, which is markedly different from its developmental role in osteoblast differentiation and chondrocyte proliferation. Further, our preliminary data suggest that these Ihh-induced repair chondrocytes differ in important ways from those in the growth plate since repair chondrocytes co-express osteoblast genes even at pre-hypertrophic stages. This increase in osteogenic character subsequently correlates with a conversion of chondrocytes into osteocytes. In Aim 3, we test whether repair and developmental chondrocytes represent distinct cell types by comparing global gene expression at different stages of maturation. We also use lineage tracing to quantitate the extent to which cartilage-derived osteocytes preferentially contribute to full thickness bone during repair. Lastly, we use powerful new chromatin accessibility assays to test that as Sox9+ periosteal cells become cartilage, their greater osteogenic character results from the maintenance of poised osteoblast enhancers. Our findings will reveal how a rare population of periosteal stem cells can be induced to make cartilage during injury, and how this specialized repair cartilage can be used to regenerate full thickness bone. A better understanding of these important stem cells will form the basis of future pre- clinical trials aimed at healing large-scale skeletal lesions in other parts of the body.

大面积骨损伤的修复仍然是一个尚未解决的临床挑战。通过开发两个 在斑马鱼和小鼠的大规模骨再生的新的和互补的模型，我们 目的是了解软骨愈伤组织在产生全厚度大段中的作用 骨头虽然骨膜在体内平衡期间产生成骨细胞，但特定的成骨细胞在体内平衡期间产生骨细胞。 构建修复愈伤组织的亚群仍然定义不清。使用转基因谱系 通过追踪，我们提供了令人信服的初步证据，证明罕见的双能Sox 9 +/Runx 2 + 骨膜群在修复期间产生新的软骨和骨。这种新发现的能力 标记、操作和分离特定的干细胞群，使我们能够测试这些干细胞是否是干细胞。 肋骨的显著再生能力是由于其骨膜干的独特性质 细胞在目标1中，我们与整形外科医生合作，测试肋骨中的Sox 9+细胞， 骨膜可以在培养物中扩张并用于愈合临界尺寸的股骨缺损。的 软骨愈伤组织的形成是骨修复中的常见特征，然而骨膜如何 只在修复过程中产生软骨仍然是一个谜。在初步数据中，我们发现， 当斑马鱼中的Ihha配体缺失或 从小鼠的Sox 9+细胞中删除Hh受体Smo。在目标2中，我们测试这反映了一个 Ihh的修复特异性作用，这与其在成骨细胞中的发育作用明显不同 分化和软骨细胞增殖。此外，我们的初步数据表明， IHH诱导的修复软骨细胞与生长板中的软骨细胞在重要方面不同， 修复软骨细胞共表达成骨细胞基因，甚至在肥大前阶段。这 成骨特性的增加随后与软骨细胞转化成 骨细胞在目标3中，我们测试修复和发育软骨细胞是否代表不同的 通过比较不同成熟阶段的整体基因表达来确定细胞类型。我们还使用 谱系追踪以定量软骨来源的骨细胞优先 有助于在修复过程中形成全层骨。最后，我们使用强大的新染色质 可及性试验测试，随着Sox 9+骨膜细胞变成软骨，它们的更大的 成骨特性来自于保持平衡的成骨细胞增强剂。我们的研究结果 将揭示一种罕见的骨膜干细胞群是如何被诱导成软骨的 以及这种专门的修复软骨如何用于再生全厚度 骨头更好地了解这些重要的干细胞将构成未来的基础， 旨在治愈身体其他部位大规模骨骼损伤的临床试验。

项目成果

Histological assessment of microtia cartilage, a potential source of autograft tissue in ear reconstruction.

小耳软骨的组织学评估，耳重建中自体移植组织的潜在来源。

• DOI: 10.1111/joa.14044
• 发表时间: 2024
• 期刊: Journal of anatomy
• 影响因子: 2.4
• 作者: Lam,DavidD;Hegde,NeelV;Patel,DivyaD;Lakeland,DanielL;Guardino,Nicholas;Kochhar,Amit;Mariani,FrancescaV
• 通讯作者: Mariani,FrancescaV

Ectoderm-mesoderm crosstalk in the embryonic limb: The role of fibroblast growth factor signaling.

• DOI: 10.1002/dvdy.24480
• 发表时间: 2017-04
• 期刊: Developmental dynamics : an official publication of the American Association of Anatomists
• 作者: Mariani FV;Fernandez-Teran M;Ros MA
• 通讯作者: Ros MA