In vivo gene manipulations of epithelial cell sheets: a novel model to study epithelial-to-mesenchymal transition

Takashi Yoshino, Daisuke Saito, Ryosuke Tadokoro, Yoshiko Takahashi
Development, Growth & Differentiation 2011, 53 (3): 378-88
Embryonic cells are classified into two types of cells by their morphology, epithelial and mesenchymal cells. During dynamic morphogenesis in development, epithelial cells often switch to mesenchymal by the process known as epithelial-to-mesenchymal transition (EMT). EMT is a central issue in cancer metastasis where epithelial-derived tumor cells are converted to mesenchymal with high mobility. Although many molecules have been identified to be involved in the EMT mostly by in vitro studies, in vivo model systems have been limited. We here established a novel model with which EMT can be analyzed directly in the living body. By an electroporation technique, we targeted a portion of the lateral plate mesoderm that forms epithelial cell sheets delineating the kidney region, called nephric coelomic epithelium (Neph-CE). Enhanced green fluorescent protein-electroporated Neph-CE retained the epithelial integrity without invading into the underling stroma (mesonephros). The Neph-CE transgenesis further allowed us to explore EMT inducers in vivo, and to find that Ras-Raf and RhoA signals were potent inducers. Live-imaging confocal microscopy revealed that during EMT processes cells started extending cellular protrusions toward the stroma, followed by translocation of their cell bodies. Furthermore, we established a long-term tracing of EMT-induced cells, which were dynamically relocated within the kidney stroma. The Neph-CE-transgenesis will open a way to study cellular and molecular mechanisms underlying EMT directly in actual body.

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