Cellular and Molecular Mechanisms of Murine Digit Regeneration

项目摘要 人类在广泛的肌肉骨骼损伤后再生潜力有限，尤其是 在失去肢体后。因此，生物修复缺失肢体的能力将显著改善 数百万截肢者的生活质量。要了解引导再生的基本机制 复杂的肌肉骨骼组织或器官，小鼠断指模型已被用于研究 肌肉骨骼再生，在去除远端指尖时以时空控制的方式发生。 虽然在阐明导致这一自然现象的细胞类型和生长因子方面取得了很大进展 再生反应，手指再生背后的确切细胞和分子机制尚未 会被揭穿。最近的研究表明，骨膜来源的细胞参与骨再生。 成骨细胞可能是成骨细胞系细胞的来源。然而，目前还不清楚是什么 在一定程度上，这种特定的细胞群在手指再生中起着直接的作用。 我们假设远端断指后不同分子事件的时空激活 介导骨膜细胞(PC)的激活、增殖和/或分化。为了检验这一假设，我们 将使用OSX-CreERT2研究截肢后PC的时空反应；Ai9可诱导 AIM 1中的报告转基因小鼠，这将使该细胞群体的谱系追踪成为可能。此外，我们还将 复制诱导条件性消融检测手指再生过程中PC细胞的增殖需求 Pc使用3.6Col1a1-tk转基因小鼠。最后，我们将确定激活和维持的信号提示 使用RNA原位杂交和组织特异性RNA测序技术在AIM 2中再生数字，并具有特定的 重点研究规范Wnt信号的时空分布。通过鉴定细胞和分子 对于再生至关重要的途径，这项建议将开辟新的治疗途径，有助于恢复 丢失组织的生物成分和结构。

Project Summary Humans have limited regenerative potential after extensive musculoskeletal injuries, particularly following limb loss. As such, the ability to biologically restore the missing limb will significantly improve the quality of life for millions of amputees. To understand the fundamental mechanisms guiding regeneration of complex musculoskeletal tissues or organs, mouse models of digit amputation have been used to study musculoskeletal regrowth, which occurs in a spatiotemporally controlled manner upon distal digit tip removal. While much progress has been made to elucidate the cell types and growth factors responsible for this natural regenerative response, the exact cellular and molecular mechanisms underlying digit regeneration have yet to be revealed. Recent studies suggest that cells derived from the periosteum participate in bone regrowth in adults, and thus represent a possible source of osteoblast lineage cells. However, it remains unknown to what extent this specific cell population plays a direct role in digit regeneration. We hypothesize that spatiotemporal activation of distinct molecular events after distal digit amputation mediates the activation, proliferation, and/or differentiation of periosteal cells (PCs). To test this hypothesis, we will investigate the spatiotemporal response of PCs after digit amputation using Osx-CreERT2;Ai9 inducible reporter transgenic mice in Aim 1, which will permit lineage tracing of this cell population. Furthermore, we will test the requirement of proliferating PCs during digit regeneration by inducible conditional ablation of replicating PCs using 3.6Col1a1-tk transgenic mice. Finally, we will determine the signaling cues that activate and sustain digit regeneration in Aim 2 using RNA in situ hybridization and tissue-specific RNA-sequencing, with a specific focus on the spatiotemporal distribution of canonical Wnt signaling. By identifying the cells and molecular pathways vital to regeneration, this proposal will open new avenues to therapies that will help restore the biological composition and structure of the lost tissues.