自噬性GATA4蛋白质沉积调控成骨前体细胞功能及其在骨形成中的作用

Autophagy is suppressed during senescence, which leads to abnormal accumulation of proteins and impaired cell function. GATA4 is a key new "switch" molecule regulating cell senescence. We first discovered that GATA4 protein accumulated in the osteoblast progenitors (POBs) in aged mice. This study aims to investigate whether the reduction of POBs autophagy caused GATA4 protein accumulation leads to impaired osteoblastic bone formation and occurrence of age-related bone loss. Firstly, we will observe the effects of autophagy on the metabolism of GATA4 protein by selective inhibition and activation of POBs autophagy. Then the cell senescence models will be established to confirm whether the suppressed autophagy of POBs leads to accumulation of GATA4 protein. And the POBs specific GATA knockout or overexpressed transgenic mouse model will be generated to observe the changes of POBs self-renewal and differentiation function as well as bone formation in vivo. The effects of GATA4 protein on Notch signaling pathway will be further measured, and the downstream signal pathway of GATA4 will be analyzed. Lastly, we will test the therapeutic effects of GATA4 antibody on bone loss in aged mice. This study will open up new ideas and new ways for the underlying mechanism as well as the prevention and treatment of senile osteoporosis.

衰老时细胞自噬减弱可导致蛋白质异常堆积，损害细胞功能；GATA4为新发现的调控细胞衰老的关键“开关”分子。我们前期研究首次发现衰老时小鼠成骨前体细胞（POBs）中GATA4蛋白质沉积，本研究拟探讨是否存在衰老时POBs自噬减弱导致GATA4蛋白质沉积，损害POBs功能导致骨形成减少，从而发生老年性骨丢失的新机制。首先，拟选择性抑制与激活POBs自噬，观察自噬变化对GATA4蛋白质代谢的影响；然后建立细胞衰老模型，确证POBs衰老使自噬减弱，导致GATA4蛋白堆积；再通过转基因小鼠模型，观察POBs特异性GATA4表达增加或缺失，对POBs自我更新与分化功能、以及骨形成的作用；进一步观察GATA4蛋白质对Notch信号通路的影响，解析GATA4下游信号通路，并研究GATA4抗体对老年小鼠骨丢失的治疗作用。为老年性骨质疏松症发病新机制与防治研究开辟新思路与新途径。

衰老时细胞自噬减弱可导致蛋白质异常堆积，损害细胞功能；GATA4为新发现的调控细胞衰老的关键“开关”分子。我们前期研究首次发现衰老时小鼠成骨前体细胞（POBs）中GATA4蛋白质沉积。本研究通过体内和体外实验探讨了衰老后自噬减弱导致POBs中GATA4蛋白质沉积，损害POBs功能导致骨形成减少，从而导致骨丢失。首先，通过选择性抑制与激活POBs自噬，发现抑制自噬导致GATA4增多，激活自噬能减少GATA4。然后诱导细胞衰老，发现诱导 POBs衰老能导致GATA4蛋白堆积。再通过过表达或敲减GATA4，发现过表达GATA4能促进POBs衰老，抑制POBs的成骨分化能力，而敲减GATA4能抑制POBs衰老，促进POBs的成骨分化能力。再通过构建腺相关病毒，观察POBs特异性GATA4过表达或降低，对骨形成的作用。最后发现 GATA4蛋白质通过Notch信号通路影响POBs的功能。本研究为老年性骨质疏松的防治提供了新思路。