Collaborative Research: A Nanostructured Model of the Apoptotic Cell Surface

协作性研究：细胞表面凋亡的纳米结构模型模拟细胞膜的传感器有助于理解蛋白质-膜相互作用。然而，凋亡细胞的膜在脂类成分和结构上不同，含有氧化的脂类，导致纳米和微米级的突起。膜的形状是蛋白质识别的潜在部位，也是人体识别受损细胞以去除的关键方式。通过模仿凋亡膜的形状，将创造出更准确地反映凋亡细胞的传感器。纳米结构膜将使用支持的双层技术来创建。这些凋亡细胞表面的纳米结构模型将允许对膜曲率和脂质成分进行单独控制。蛋白质与这些传感器的结合将使用表面敏感的荧光显微镜技术进行监测。使用这种方法设计的传感器将有助于理解蛋白质如何与弯曲的膜表面相互作用。这项研究的第一个目标是构建支持的双层膜，允许控制脂组成以及纳米级的表面结构。将制备嵌入荧光团的二氧化硅纳米颗粒，并在其表面包裹一系列氧化和非氧化的脂类。这些纳米粒子将附着在玻璃表面，并在其上形成一层脂质双层。这项研究的第二个目标是展示这些纳米结构膜传感器测量蛋白质与曲面结合的能力。曲率对识别要去除的凋亡细胞所涉及的蛋白质结合的影响将使用全内反射荧光显微镜来测量。C反应蛋白与膜的结合将被可视化和定量测量，以确定凋亡膜的哪些特征对蛋白质识别至关重要。这些信息将指导未来传感器的设计，这些传感器可以检测身体对凋亡细胞的反应。智力上的优点：尚不清楚是脂类、膜曲率还是两者的组合决定了蛋白质与凋亡细胞的结合。这项工作将回答这个问题，并提高对凋亡细胞识别的理解。反过来，这将使传感器的设计能够识别对凋亡细胞的生理反应，这可能是诊断心血管疾病的关键。这里开发的用于工程模拟凋亡细胞膜的方法将成为基于光学、电子或基于质量的信号转导的传感器的基础。广泛的影响：拟议的研究将影响教学、培训和推广。一名博士后科学家和几名学生将接受纳米科学、生物学、化学和光谱学之间的新兴技术培训。此外，该项目的教育影响将延伸到两个校园之外，并将涉及代表性不足的少数群体。来自奥罗拉社区学院、丹佛社区学院和丹佛大都会州立学院的鼓励科学专业人员计划的学生将参与拟议的研究。第二种外展方法是通过落基山-中学数学与科学伙伴关系将中学科学教师包括在内。

AbstractCollaborative Research: A Nanostructured Model of the Apoptotic Cell SurfaceSensors that mimic cellular membranes have contributed to understanding of protein-membrane interactions. However, the membranes of an apoptotic cells differ in lipid composition and structure, containing oxidized lipids that cause nanoscale and microscale protrusions. Membrane shape is a potential site for protein recognition and a critical way in which the body identifies damaged cells for removal. By mimicking the shapes of apoptotic membranes, sensors will be created that more accurately reflect apoptotic cells. Nanostructured membranes will be created using supported bilayer techniques. These nanostructured models of the apoptotic cell surface will allow for separate control of both membrane curvature and lipid composition. Binding of protein to these sensors will be monitored using a surface sensitive, fluorescence microscopy technique. Sensors designed using this approach will be useful for understanding how proteins interact with curved membrane surfaces. The first goal of this research is to construct supported bilayer membranes that allow for control of lipid composition as well as nanoscale surface structure. Silica nanoparticles embedded with fluorophores will be prepared and coated with a series of oxidized and non-oxidized lipids. These nanoparticles will be attached to glass surfaces and a lipid bilayer will be formed over them. The second goal of this research is to demonstrate the ability of these nanostructured membrane sensors to measure protein binding to curved surfaces. The effects of curvature on the binding of proteins involved with recognizing apoptotic cells for removal will be measured using total internal reflection fluorescence microscopy. Binding of C-reactive protein to the membranes will be visualized and quantitatively measured to determine what features of an apoptotic membrane are critical for protein recognition. This information will guide the future design of sensors that detect how the body responds to apoptotic cells.Intellectual Merits: It is unknown whether lipids, membrane curvature, or the combination determines protein binding to apoptotic cells. This work will answer that question and improve understanding of apoptotic cell recognition. In turn, this will allow for the design of sensors that recognize physiological responses to apoptotic cells that could be critical in diagnosing cardiovascular disease. The approaches developed here for engineering mimics of apoptotic cell membranes will become the foundation for sensors based on optical, electronic, or mass based signal transduction.Broader impacts: The proposed research will impact teaching, training, and outreach. A postdoctoral scientist and several students will be trained in emerging techniques at the interface of nanoscience, biology, chemistry, and spectroscopy. Additionally, the educational impact of this project will extend beyond the two campuses and will reach out to involve underrepresented minorities. Students from the Strides Toward Encouraging Professions in Science program at the Community College of Aurora, Community College of Denver and Metropolitan State College of Denver will be involved in the proposed research. A second method of outreach will be to include middle school science teachers through the Rocky Mountain-Middle School Math and Science Partnership.