Matrix Organization and Dimensionality

在胚胎发育的不同阶段和成人伤口修复期间，细胞与不同组织中生物化学和结构上不同形式的细胞外基质相互作用。该项目的重点是解决有关这些细胞-细胞外基质相互作用的机制的主要问题。例如，与平面（2D）细胞培养基质或甚至1D纤维基质相比，不同类型的哺乳动物细胞使用什么独特的机制来粘附并迁移通过不同的三维（3D）细胞外基质环境？ 我们目前正在探索在动态条件下形成2D与3D纤维状细胞外基质相比的细胞-基质粘附。在常规2D细胞培养中已经相当深入地研究的细胞外基质分子是纤连蛋白。我们和其他人以前描述了两种不同的机制，用于组装纤维状纤连蛋白，一个涉及横向易位的α 5-β 1整联蛋白沿着平坦的组织培养基质，和第二个机制涉及移位的整个灶性粘连沿着基膜基板驱动的肌动球蛋白“收缩绞盘”系统。我们现在已经开始研究纤连蛋白基质在3D纤维胶原基质中组装的机制。我们也在进一步深入探索我们先前的观察，即在3D纤维胶原环境中存在异常高水平的整合素活化，例如，肌动球蛋白收缩性在这种激活状态中起什么作用。 基底膜的组装发生在胚胎中上皮细胞的基底表面，粘附到其相对2D基底膜基底上。在果蝇和成年唾液腺中，基底膜胶原蛋白的合成和分泌机制位于靠近组装位点的细胞基底端。然而，在胚胎小鼠唾液腺中，IV型胶原和层粘连蛋白的mRNA位于上皮细胞的相反顶端，与内质网和高尔基体一起沿着。我们正在研究这些重要的基底膜成分在胚胎哺乳动物器官的基底端适当地运输和分泌的机制。一个关键的机制似乎是通过微管细胞骨架运输。 这些研究为细胞与细胞外基质相互作用中维度的重要性提供了见解。它们的范围可以从细胞与单束胶原相互作用的“1D”相互作用，到基底膜和平坦基底上的2D，再到天然间质细胞外基质中的3D。了解这些相互作用的不同性质可能会提供有用的见解，合理的组织工程。

Cells interact with biochemically and structurally distinct forms of extracellular matrix in different tissues, at different stages of embryonic development, and during adult wound repair. This project focuses on addressing major questions concerning the mechanisms of these cell-extracellular matrix interactions. For example, what unique mechanisms do different types of mammalian cells use to adhere to and to migrate through different three-dimensional (3D) extracellular matrix environments compared to flat (2D) cell culture substrates, or even 1D fibrillar substrates? We are currently exploring the cell-matrix adhesions formed to 2D compared to 3D fibrillar extracellular matrix under dynamic conditions. An extracellular matrix molecule that has been studied in considerable depth in regular 2D cell culture is fibronectin. We and others previously described two distinct mechanisms for assembly of fibrillar fibronectin, one involving lateral translocation of the alpha5-beta1 integrin along flat tissue culture substrates, and a second mechanism involving the translocation of entire focal adhesions along a basement membrane substrate driven by an actomyosin "contractile winch" system. We have now initiated studies of the mechanisms of fibronectin matrix assembly in a 3D fibrillar collagen matrix. We are also exploring in further depth our previous observation that there is an unusually high level of integrin activation in a 3D fibrillar collagen environment, e.g., what role actomyosin contractility plays in this activation state. The assembly of basement membranes occurs at the basal surfaces of epithelial cells in embryos adhering to their relatively 2D basement membrane substrates. In Drosophila and adult salivary glands, the machinery for synthesis and secretion of basement membrane collagen is at the basal end of the cell close to the site of assembly. In embryonic mouse salivary glands, however, mRNA for collagen IV and laminin is at the opposite, apical end of epithelial cells, along with the endoplasmic reticulum and Golgi apparatus. We are investigating the mechanisms by which these important basement membrane components are transported and secreted appropriately at the basal ends of embryonic mammalian organs. A key mechanism appears to be transport via the microtubule cytoskeleton. These studies are providing insights into the importance of dimensionality in the interactions of cells with the extracellular matrix. They can range from a "1D" interaction as a cell interacts with a single bundle of collagen, to 2D on basement membranes and flat substrates, to 3D in native interstitial extracellular matrices. Understanding the differing nature of these interactions may provide insights useful for rational tissue engineering.