自噬在机械应力诱导成骨细胞分化中的作用及机制研究

Appropriate mechanical stress is important for bone homeostasis. It has been reported that mechanical stress could promote autophagy of various types of cells, and that autophagy could induce osteogenesis via Wnt/β-catenin signaling pathway. However, whether the mechanical stress promoted bone reconstruction in osteoporosis through the autophagy pathway is still unknown. In our preliminary experiment, we found that cyclic mechanical stretch could stimulate autophagy in osteoblasts. Thus, we hypothesize that mechanical stress can promote osteoblast differentiation and bone reconstruction through enhancing the activity of autophagy. To prove our hypothesis, we use human bone marrow mesenchymal stem cells (BMSCs), primary rat osteoblasts, human osteoblast line (hFOB1.19), transgenic mice with GFP-LC3, tail suspended mice, autophagy regulator, gene transfection and silencing techniques and mechanical stress flexible stretch system to explore the effect of autophagy on the osteogenic differentiation and bone reconstruction induced by mechanical stress. The involved molecular mechanism is also investigated. We believe these studies will deepen our understanding for the signal transduction of osteoblast differentiation and bone reconstruction under mechanical stress and provide a new therapeutic target for the prevention and treatment of osteoporosis.

适度的机械应力刺激对维持骨骼内环境稳态至关重要。研究表明机械应力能增强多种细胞自噬水平，而自噬通过激活成骨细胞Wnt/β-catenin信号通路，诱导成骨分化。但机械应力能否通过增强成骨细胞自噬水平促进骨质疏松后骨重建，目前尚不清楚。我们通过预实验证实：周期性牵张应力增强成骨细胞自噬水平。故我们提出假说：机械应力通过增强成骨细胞自噬，诱导成骨细胞分化，促进骨重建。为验证这一假说，我们使用人原代骨髓间充质干细胞和大鼠原代成骨细胞、人成骨细胞株hFOB1.19、GFP-LC3转基因小鼠，采用鼠尾悬吊模型和机械应力柔性基底拉伸系统于体内、体外模拟应力刺激，利用自噬调节剂和基因敲入/敲除技术，从分子、细胞、组织以及动物体内多层次探讨自噬在机械应力诱导成骨分化、促进骨重建中的重要作用及可能机制。本项目的研究有助于深入了解机械应力调控成骨分化的信号传导，为骨质疏松的防治提供新思路。

适度的机械应力刺激对维持骨骼内环境稳态至关重要。研究表明机械应力能增强多种细胞自噬水平，而自噬可通过激活成骨细胞Wnt/β-catenin信号通路，诱导BMSCs成骨分化。但机械应力能否通过增强成骨细胞自噬水平参与骨质疏松后骨重建，目前尚不清楚。本课题通过体内外实验发现机械牵张拉力刺激可以激活自噬并进一步促进BMSCs成骨分化。研究发现：机械应力缺失可以降低BMSCs成骨分化和自噬水平。抑制BMSCs自噬可以减弱机械应力诱导的成骨分化能力，增强自噬可以改善机械应力缺失导致的骨量丢失。这些结果证实了机械应力通过激活BMSCs自噬来促进其成骨分化。自噬激活的具体机制是通过ERK-mTOR信号通路实现的。本课题以自噬为切入点，通过体内、外实验，从分子、细胞、组织多层面研究自噬在机械应力调控BMSCs成骨分化中的作用，发现了机械应力通过自噬通路促进BMSCs成骨分化和骨矿化，这一发现具有十分重要的理论意义和应用价值。有助于进一步揭示机械应力促进骨形成的机制，也为防治骨质疏松提供了新思路与新途径。