VISUALIZATION OF RE-CELLULARIZING A CARDIAC EXTRACELLULAR MATRIX

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 窗口小室系统将允许可视化细胞与C-ECM成分在全厚度支架中的共定位，其中间充质干细胞(MSCs)有望与I型胶原共定位，内皮祖细胞(EPC)与IV型胶原共定位。 第一个目标是建立一个窗口室，以可视化全厚度C-ECM及其与内皮祖细胞和间充质干细胞的再细胞化。然后，种子细胞与C-ECM蛋白(I型和IV型胶原)的共定位将被鉴定。最后，将优化去细胞方案，以保留与细胞共定位的C-ECM蛋白，通过增加细胞与C-ECM的附着来提高再细胞化效率。 更新： 多光子显微镜将使我们能够以比我实验室现有的共焦显微镜更深的深度进行成像。二次谐波产生的胶原蛋白将用于鉴定间充质细胞与细胞外基质成分的共定位。外部荧光将使用荧光内皮细胞和间充质干细胞以及针对IV型胶原层粘连蛋白的荧光抗体进行成像。实时成像部分将在我们自己的实验室使用共焦显微镜进行。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. A window chamber system will allow visualization of cell co-localization with C-ECM components in a full thickness scaffold, in which mesenchymal stem cells (MSCs) are expected to co-localize with collagen I and endothelial progenitor cells (EPC) with collagen IV. The first goal will be to build a window chamber to visualize a full thickness C-ECM and its re-cellularization with EPCs and MSCs. Then, co-localization of seeded cells with C-ECM proteins (collagen I and IV) will be characterized. Lastly, the de-cellularization protocol will be optimized to preserve the C-ECM proteins with which cells were shown to co-localize, increasing the re-cellularization efficiency by increasing the attachment of cells to the C-ECM. Updated: The multi-photon microscopy will enable us to image at a greater depth than is currently possible with the confocal microscopes available in my lab. Second harmonic generation of collagen will be used to identify co-localization of mesenchymal cells with ECM components. External fluorescence will be imaged using fluorescent endothelial and mesenchymal stem cells and fluorescent antibodies specific to collagen IV laminin. The real-time imaging portion will be performed in our own lab using a confocal microscope.