Development of a mechanosensitive synthetic cell for mediating intercellular communication.

项目摘要 我们设计能够与活细胞通信的合成细胞系统的能力仍然有限。这个 长期目标是设计出具有日益复杂的仿生功能的类细胞系统，这些功能可以 作为天然细胞的细胞替代或增强功能。这项提议的目标是制定一项 机械敏感的合成细胞，可以对剪切力的增加做出反应，剪切力在 心血管系统，并分泌生物活性分子影响活细胞。我们体内的细胞 不断感知和响应微环境刺激，包括被动和主动的物理刺激， 如细胞外基质硬度、粘附剂配基密度、拉伸、压缩和流体剪切流动。这个 这一提议的基本原理是，完成这项工作将产生一种新的仿生细胞状系统，作为 可与天然活细胞对接的新型剪应力响应型“材料”。大多数人 工程生物材料对生化环境的差异(例如，氧化还原的差异， PH和酶的组成)在正常和病变组织之间。相比之下，一直有 在利用刺激反应行为的力量方面的努力相对较少。合成细胞的想法是受到启发的 通过天然血小板对升高的切应力和分泌颗粒内容物的结合和反应的能力 被绑在一个表面上。所提出的工作包括三个具体目标：1)表征剪切应力 机械感觉囊泡的反应，2)机械感觉与合成细胞胞吐作用的耦合，3) 体外检测切应力激活的合成细胞与内皮细胞的细胞间通讯。我们 将使用一种创新的方法来实现这些目标，即重新调整剪切机械敏感通道的用途 应力传感和使用基于多肽的膜融合。我们的实验室是第一个证明 机械传感合成细胞，我们在自下而上的合成生物学方面拥有重要的专业知识。这个 提出的研究具有重要意义，因为这将是第一个为通信而开发的合成细胞系统 与哺乳动物细胞利用钙触发的分泌。这项工作将制定基本的战略 将机械感觉与合成细胞中的生化反应相结合。这将为其他人开辟新的途径 有兴趣开发更复杂的类细胞系统的研究人员。其结果将具有重要的意义 立即产生积极影响，因为它将支持机械敏感通道可以感觉到 由于剪应力和长期的横向膜张力，因为他们奠定了基础 设计具有其他传感能力的合成细胞。

Project Summary Our abilities to engineer synthetic cell systems that can communicate with living cells remain limited. The long-term goal is to engineer cell-like systems with increasingly complex biomimetic functions that can serve as cell replacement or augment functions of natural cells. The objective of this proposal is to develop a mechanosensitive synthetic cell that can respond to an increase in shear stress, which is most prevalent in the cardiovascular system, and secrete bioactive molecules to effect living cells. Cells in our bodies constantly sense and respond to microenvironmental stimuli, including passive and active physical stimuli, such as extracellular matrix rigidity, adhesive ligand density, tension, compression, and fluid shear flow. The rationale underlying this proposal is that completion will result in a novel biomimetic cell-like system as a novel shear stress-responsive ‘material’ that can interface with natural living cells. The majority of engineered biomaterials respond to differences in the biochemical environment (e.g. differences in redox, pH, and enzyme composition) between normal and diseased tissues. By comparison, there has been relatively less effort in exploiting forces for stimulus-responsive behaviors. The synthetic cell idea is inspired by natural platelets’ ability to bind and respond to elevated shear stress and secrete granule contents when bound to a surface. The proposed work consists of three specific aims: 1) Characterize shear stress response of mechanosensing vesicles, 2) Couple mechanosensing with exocytosis in synthetic cells, 3) Test intercellular communication of shear stress-activated synthetic cells with endothelial cells in vitro. We will pursue these aims using an innovative approach of repurposing mechanosensitive channel for shear stress sensing and using peptide-based membrane fusion. Our lab was the first group to demonstrate mechanosensing synthetic cells and we have significant expertise in bottom-up synthetic biology. The proposed research is significant, because it will be the first synthetic cell system developed to communicate with mammalian cells using calcium-triggered secretion. The work will develop fundamental strategies for coupling mechanosensing to a biochemical response in synthetic cells. This will open new avenue for other researchers interested in developing more complex cell-like systems. The results will have an important positive impact immediately because it will support the idea that mechanosensitive channels can sense lateral membrane tension due to shear stress and long-term because they lay the groundwork of engineering synthetic cells with other sensing abilities.