细胞长程力在上皮管状结构自组装中的作用及其信号转导机制研究

Tubular structures are one of the most common tissue types, enabling bulk transport of molecules and other substances in human body. How cells assemble into such long and well-organized tubes is an important question in tubulogenesis. In recent years, the role of long-range mechanical force, which crosses the scale beyond single cells to function on cell-population and tissue/organ levels, has attracted enthusiastic attention in tubulogenesis. During in vitro three-dimensional (3D) culture, it was found that mammary epithelial cells can rely on long-range mechanical force to establish distant crosstalk through assembled type-I collagen fiber straps, thus guiding the tubular pattern formation. However, so far the characterization of the long-range mechanical force and its signaling mechanism during tubulogenesis is still limited. This project is to study the role of long-range force and its mechanism in the self-organization of tubular structures, based on the two three-dimensional culture models of mammary and kidney epithelial cells. The major study aims include: 1) to investigate the mechanical mechanism how long-range force induces the assembly of type I collagen fiber straps by incorporating cell rotation mobility-induced dynamic tension force on the 3D gel; 2) by application of fluorescent probes such as FRET biosensors, to reveal the signaling mechanism how long-range force activates cells via collagen straps to induce self-assembly of epithelial tube patterns; 3) to determine the critical role of long-range force in the rapid and collective polarization of epithelial cell group occurred during tubular morphogenesis. This project is to provide important evidence for the mechanism study on long-range force in tubulogenesis, and to recognize the special biological function of long-range force at cell-population level beside single-cell biomechanics, which has important implications in scientific theory and health-relevant applications.

管状结构是体内最常见的一种组织形式，具有关键的物质运输功能，而细胞群体如何组装形成管状结构，是组织发生中的重要科学问题。近来，细胞长程力作用,即力的传递范围超越单细胞而作用于细胞群体，在管状结构发生中的作用引起了重视。申请人在前期研究中发现，细胞通过长程力诱导胞外基质中I型胶原纤维带形成，从而促进上皮管状结构的自组装，但其作用机制尚未明了。本项目拟在乳腺和肾上皮细胞三维培养模型中，研究长程力在上皮管状结构自组装中的作用及其信号转导机制，包括：1）细胞长程力作用中I型胶原纤维带的诱导和形成机制；2）长程力作用通过I型胶原纤维带激活细胞和诱导管状结构自组装的力学信号转导机制；3）长程力在管状结构形成中对细胞群体快速协同极化的诱导作用。本项目的研究将有助认识长程力作用在细胞群体生物学中的独特功能，为体内上皮管状组织结构发生的力学生物学机制提供理论依据，具有重要的科学意义和应用价值。

体内组织器官的发生是一个精确而自主调控的过程，生物力学因素在该宏观尺度上的功能是值得探索的基础科学问题。本项目围绕揭示细胞长程力作用的基本物理与生物学机制展开，理解其在细胞群体和上皮微组织形态发生中的功能，以下是主要研究内容及结果。.1）.揭示细胞长程力作用的基本物理与生物学机制，发现细胞能敏感响应相邻细胞通过基质传递的张应力，从而建立长距离的力学交流，促进细胞定向迁移和群体自组装，其作为细胞间的一种基本信号交流方式，称为“细胞长距离力学通讯”。.2）.发现液相层面的细胞－基质互作调节上皮微组织的形态发生。肾上皮MDCK细胞摄取溶液中的基质成分，生成的片层状基底膜和纤维状胶原分别介导上皮球形小叶、上皮管状结构与分枝形态的发生，而改变基底的力学微环境后，如悬浮培养或在非弹性形变的琼脂糖凝胶上，细胞不能有效组装成管状结构，体现长距离力学因素的调节作用。.3）.阐明细胞长距离力学通讯的分子机制，证实细胞内质网膜上的钙通道、肌丝骨架、膜上整合素蛋白参与细胞之间的力学响应，N钙黏蛋白是力学调控细胞形成稳定网格结构的必要条件。结合FRET活细胞观测技术，阐明基底拉伸力激活细胞内ERK活性的力学信号机制，发现机械拉升力通过激活钙信号、肌丝骨架等激活ERK。.4）.深入理解细胞力学重构胶原基质的力学机制及相关联的细胞信号。分子动力学和有限元模型高度预测了细胞力学促进不同几何形状的胶原纤维图案发生，证实细胞力学在基质中的动态存在和各向异性分布，促进胶原基质的不均匀重构，并发现细胞肌动蛋白骨架、内质网钙通道、膜上力敏感受体起调节作用。 . 依据本项目研究的一个结论具有重要基础科学意义，即长距离力学作用在细胞群体和组织水平上提供了一种基本的通讯交流方式，同时本工作为组织与器官在发育中形成的异质性结构提供了细胞力学角度的可能解读。

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