EXTRACELLULAR MATRIX AS A SOLID STATE REGULATOR IN ANGIOGENESIS

这一更新提案的总体目标仍然是了解 细胞外基质（ECM）调节毛细血管发育的生物力学机制。 在最后一次资助期间，我们使用了含有ECM涂层的粘附岛的微加工基底，这些粘附岛在微米尺度上具有确定的尺寸和形状，以证明ECM根据其抵抗细胞牵引力、改变细胞骨架（CSK）张力并从而调节细胞形状的能力来控制毛细血管内皮（CE）细胞是否会生长、分化、经历血管扩张或迁移。CSK张力被发现依赖于收缩性微丝、微管和ECM粘附之间的机械相互作用，并且需要整合素受体和CSK之间通过粘着斑蛋白（如黏着斑蛋白）的结合进行有效的机械偶联。 这种物理连接的维持也被发现是由小G蛋白，外消旋体刺激的片状伪足形成所需的，其中CSK张力产生的药理学抑制迅速抑制片状伪足运动。 因此，本建议的主要目的是扩展我们正在进行的研究，并专注于ECM的生物力学机制：调节方向almigration和血管图案响应血管生成刺激。具体目标包括：1）探索ECM结构和机械变化如何改变CSK张力和局灶性粘附形成，2）确定CE细胞和ECM之间的机械相互作用如何控制釉伪足延伸和定向迁移，3）探索CSK张力的局部变化是否有助于毛细血管形态发生过程中的图案形成。

The general goal of this renewal proposal continues to be to understand the biomechanical mechanism by which extracellular matrix (ECM) regulates capillary development. During the last grant period,we used microfabricated substrates containing ECM-coated adhesive islands of defined size and shape on the micronscale to demonstrate that ECM governs whether capillary endothelial(CE) cells will grow, differentiate, undergo apoptosisor migratein response to angiogenic factors based on its ability to resist cell tractional forces, alter tension in the cytoskeleton (CSK), and thereby modulate cell shape. CSK tensionwas found to be dependent on mechanical interactions between contractile microfilaments, microtubules, and ECM adhesions and to require efficient mechanical coupling between integrin receptors and the CSK through binding of focal adhesion proteins,such as vinculin. Maintenance of this physical linkage also was found to be required for formation of lamellipodia stimulated by the small G protein, rac, where as pharmacological inhibition of CSK tension generation rapidly suppressed lamellipodial movement. Thus, the main objective of the present proposal is to extend our ongoing studies and to focus on the biomechanical mechanism bywhich ECM: regulates direction almigration and vascular patterning in response to angiogenic stimuli. The specificaims include: 1) to explore how changes in ECM structure and mechanicsalter CSK tensionand focal; adhesion formation,2) to determine how mechanical interactions between CE cellsand ECM control amellipodia extension and directional migration, and 3) to explore whether local changes in CSK tension contribute to pattern formation during capillary morphogenesis.