Mechanisms of skeletal patterning during limb development and regeneration

肢体发育和再生过程中骨骼模式的机制

• 批准号: 10611365
• 负责人: James R Monaghan
• 金额: $ 29.55万
• 依托单位: NORTHEASTERN UNIVERSITY
• 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 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10611365
• 关键词: Adopted; Affect; Agonist; Ambystoma; Ambystoma mexicanum; Amputation; Animals; Back; Birth; CRISPR/Cas technology; Cell surface; Cells; Child; Congenital Abnormality; Connective Tissue Cells; Deformity; Disease; Distal; Engineering; Epigenetic Process; Event; Fluorescent in Situ Hybridization; Future; Gene Expression; Genes; Genetic; Genetic Transcription; Genomic approach; Goals; Grant; Human; Individual; Knowledge; Laboratories; Ligands; Limb Development; Limb structure; Mediating; Medicine; Memory; Modification; Molecular; Molecular Target; Natural regeneration; Outcome; Pattern; Play; Process; Prosthesis; Regenerative Medicine; Regenerative research; Resolution; Retinoic Acid Receptor; Role; Salamander; Signal Transduction; Signaling Molecule; Site; Structure; System; Testing; Therapeutic; Tissues; Trauma; Tretinoin; United States; blastema; chondrodysplasia; clinically relevant; disability; epithelial wound; in vivo; injured; insight; knockout gene; limb regeneration; morphogens; pharmacologic; predictive modeling; progenitor; programs; receptor; regenerative; regenerative approach; regenerative biology; regenerative therapy; retinoic acid receptor alpha; retinoic acid receptor gamma; single-cell RNA sequencing; skeletal; skeletal disorder; skeletal regeneration; therapeutic target; transcriptomics

PROJECT SUMMARY Limb congenital defects are among the most common causes of disability in the United States affecting 0.8/1000 children born each year. Recent realizations of plausible regenerative medicine therapies in humans has increased efforts towards understanding examples of animal regeneration and the mechanisms that drive the process. In this proposal, we aim to provide insight into the mechanisms that facilitate regeneration by studying a clinically relevant signaling molecule, retinoic acid, during limb regeneration in the salamander. Axolotl salamanders (Ambystoma mexicanum) are particularly useful, because they regenerate entire limbs after amputation by generating a heterogeneous mass of lineage- restricted progenitors called a blastema at the amputation site. The blastema uses positional information retained in connective tissue cells to regenerate the proper limb regions. The goal of this proposal is to the endogenous role in this process and study how positional information is established and maintained in connective tissue cells, which is critical to our understanding of limb regeneration. We will use unbiased single cell RNAseq and spatial transcriptomic approaches to compare gene expression at single-cell resolution throughout the regeneration process. We will also determine the transcriptional targets of retinoic acid specifically in limb connective tissue cells that are undergoing reprogramming from distal to proximal cell identity. Lastly, the positional memory of connective tissue cells will be experimentally programmed in vivo to demonstrate how positional memory is maintained. In the end, the major outcome of this project will be to know how a salamander limb knows what limb structures to grow back in order to stimulate new regenerative approaches in mammalian systems.

项目总结 肢体先天缺陷是美国最常见的残疾原因之一，影响 每年出生的每1000名儿童中就有0.8名。中国再生医学疗法的最新进展 人类已经加大了对动物再生的例子的理解力度 驱动这一过程的机制。在这项提案中，我们旨在提供对机制的洞察 通过研究临床上相关的信号分子维甲酸，促进肢体再生 火蜥蜴的再生。紫杉醇火蜥蜴(Ambystoma Micianum)特别有用， 因为它们在截肢后通过产生不同种类的血统来再生整个肢体- 受限制的祖细胞在截肢部位称为胚泡。胚泡使用体位 保留在结缔组织细胞中的信息，以再生适当的肢体区域。这样做的目的是 建议在这一过程中发挥内生作用，并研究位置信息是如何建立的 并维持在结缔组织细胞中，这对我们理解肢体再生至关重要。 我们将使用无偏见的单细胞RNAseq和空间转录切割方法来比较基因 在整个再生过程中以单细胞分辨率表达。我们还将确定 维甲酸在肢体结缔组织细胞中的特异性转录靶点 从远端细胞到近端细胞的重新编程。最后，结缔组织的位置记忆 细胞将在体内进行实验编程，以演示位置记忆是如何保持的。 最后，这个项目的主要成果将是了解火蜥蜴的肢体是如何知道什么肢体的 为了在哺乳动物系统中刺激新的再生方法，结构需要重新生长。