Nanoflare-based isolation of cardiac progenitor cells derived from murine and human pluripotent stem cells

点击翻译按钮获取中文摘要

Today, the isolation of cardiac progenitor cells derived during in-vitro differentiation of pluripotent stem cells is limited by the lack of appropriate cell surface marker. The isolation of cardiac progenitor cells is an indispensible requirement for cardiac regenerative strategies, research in cardiac development as well as the use of iPS based in vitro disease models. Currently, cardiac progenitor cell populations are defined by their specific expression profile of different transcription factors like the parallel expression of Tbx5 and Nkx2.5 for the primary heart field (PHF) and Isl1 for the second heart field (SHF). In the proposed project we will use the nanoflare technology for a specific isolation of PHF and SHF progenitor cells derived during in vitro differentiation of pluripotent stem cells. The nanoflare technology is a new approach for endogenous live mRNA detection. Nanoflares consists of a central gold nanoparticle with conjugated fluorescent-labeled DNA probes for cell specific mRNA detection. The binding of the mRNA to the nanoflare probe leads to the release of the fluorescence signal. Afterwards the target cells can be isolated by this specific and transient fluorescence signal using FACS sorting. The proposed project for specific cardiac progenitor cell isolation using the nanoflare technology consists of the following milestones: 1) Efficient cell labeling during in vitro differentiation of muirne and human pluripotent stem cells by nanoflares 2) Detection of murine cardiac progenitor cells using specific nanoflares for Tbx5, Nkx2.5 and Isl1 during in vitro differentiation of appropriate transgenic pluripotent stem cell lines 3) Detailed characterization of nanoflare based isolated murine cardiac progenitor cells 4) Detection and characterization of human cardiac progenitor cells isolated by nanoflares during in vitro differentiation of human pluripotent stem cells In summary, the proposed project will build the indispensible requirement for a specific, efficient, easy and the physiological properties preserving technology for the isolation of cardiac progenitor cells derived from pluripotent stem cells. Beside this, the proposed technology could be easily transferred to different research areas, which use cell derivates derived from pluripotent stem cells and therefore harbors also a great general potential for the scientific community.