力学信号通过整合素偶联激酶对牙周膜细胞代谢谱的影响

Mechanical stress induces periodontal tissue remodeling.As the periodontal ligament stem cell is highly responsive to mechanical stimuli, it plays the important role in the orthodontic tooth movement. However,it is not definitively determined the initial responses which involved in mechanotransdution mechanism in the periodontal ligament stem cells. Integrin-linked kinase(ILK) is the most important knots in ECM-Integrin-Cytoskeleton mechanotransduction signal pathway. According to references and our preliminary work, the metabolites variation in cell and tissue may be the terminal effects of mechanotransduction. This research will set up the mechanical loading model of human periodontal ligament cell. The ILK gene will be overexpressed or silent by RNA interfering technology in the model. Combining the metabonomics method, we will research the effect of ILK mediated mechanotransduction on the periodontal ligament cellular metabolism from the positive and negative aspects, and we will firstly use a variety of methods of NMR and UPLC-MS to acquise the metabolomics fingerprinting data, apply PCA data analysis methods, and compare with the international common metabolite database to screen out the sensitive metabolic landmarks, and then research the specific mechanism using the blocker of different two downstream signal transduction passageway of ILK, VIII for Akt, LY294002 for PI3K.The ultimate purpose of this study is to discuss the effect of ILK-mediated mechanotransduction on periodontal tissue metabolism spectrum in the levels of metabolites when stress as the influencing factors. The result may help us to clarify the complexity of biological network and the relationship between the metabolites and the genes, the proteins in the remolding of periodontal ligament, then explore the mechanism of stress-mediated periodontal tissue remodeling more deeply and systematically.

细胞力学信号转导及牙周膜改建是正畸牙移动的生物学基础，建立力学信号转导分子生物学系统评价参数体系，有助于在生理范围内促进力学信号转导及适应性改建，对优化正畸力系统、加快牙移动及减轻矫治痛苦均具有十分重要的意义。文献及前期的研究表明，代谢谱的变化可能是整合素力学信号转导通路的终端效应，而整合素偶联激酶(ILK)是沟通整合素与细胞骨架的关键激酶。本研究拟通过RNA干扰技术建立ILK 基因过表达及沉默的人牙周膜细胞力学加载模型，结合代谢组学检测方法，从正反两个方面研究应力通过ILK对牙周膜细胞代谢谱的影响，筛选ILK 敏感代谢标志物，并应用不同阻断剂研究下游各信号通路对不同类型代谢产物表达的影响，在代谢谱层面阐明ILK在正畸牙周组织改建中的具体作用机制，明确牙周组织改建中代谢物与蛋白质、基因之间的关系，旨在从代谢谱角度初步建立牙周膜改建的分子生物学评价参数体系，为临床优化矫治力系统提供实验依据。

细胞力学信号转导及其引发的牙周膜改建是正畸牙移动的生物学基础，建立力学信号转导分子生物学系统评价参数体系，有助于在生理范围内促进力学信号转导及适应性改建，对优化正畸力系统、加快牙移动及减轻矫治痛苦均具有十分重要的意义。前期研究表明，代谢谱变化可能是整合素力学信号转导通路的终端效应，而整合素偶联激酶(ILK)是沟通整合素与细胞骨架的关键激酶。.本研究建立ILK 基因过表达及沉默的人牙周膜细胞力学加载模型，研究应力通过ILK对牙周膜细胞代谢谱的影响，筛选ILK敏感代谢标志物，明确牙周组织改建中代谢物与蛋白质、基因之间的关系，同时研究ILK介导的力学信号转导通路对人牙周膜细胞增殖、凋亡和分化的影响，进行小分子代谢谱与上述生物学行为的相关性分析，以明确小分子代谢谱成分变化与生物学行为间的相互关系。旨在从代谢谱角度初步建立牙周膜改建的分子生物学评价参数体系，为临床优化矫治力系统提供实验依据。.目前研究表明，适宜力学刺激可引起细胞内骨架重排及细胞间连接变化，导致人牙周膜细胞形态排列变化。代谢组学检测结果表明机械应力刺激可影响细胞内及细胞外基质小分子代谢产物的组成，氨基酸及有机酸检测发现多数的产物变化呈现曲线性而非直线性的变化趋势，ILK在力学信号传导通路中作为关键因素影响着某些氨基酸和有机酸的代谢，包括细胞内液中的丙氨酸、精氨酸、天门冬氨酸、谷氨酸、甘氨酸、赖氨酸、丝氨酸、苏氨酸、乳酸、柠檬酸和细胞外液中的天门冬氨酸。这些可以作为可能的ILK敏感代谢标志物，进行进一步实验筛选。根据实验组和对照组呈现出显著性差异的时间点的不同，表明了代谢产物变化对于力学信号及ILK效应时间的差异。引起上述变化的具体机制如何尚需要进一步探究。敏感性小分子代谢物各自具有其代谢途径，力学信号和ILK通过何种途径对它们的代谢产生影响，与它们自身特有的代谢途径产生了何种联系，尚需要我们继续对每种敏感代谢标志物代谢路径及其作用深入研究。