Culture of Embryonic Cells for Analysis of Amphibian and Mammalian Early Embryogenesis
Part of the
Bodega Marine Laboratory Marine Science Series
book series (BMSS)
Culture of embryonic cells isolated from early embryos allows detailed analysis of the cell motility, interactions between cells, and between cells and the extracellular matrix (ECM). There is always the risk that cell behavior in culture might be artifacts not related to the behavior inside embryos. In many situations, one must use cells immediately after isolation from embryos, and try to prepare culture conditions similar to those inside the embryo.
Migration of the presumptive mesodermal cells during amphibian gastrulation was studied by such methods. It was first necessary to find culture conditions to allow mesodermal cells to behave in a manner similar to that found inside embryos. By the use of culture medium with a pH and calcium ion concentration found in embryos, as well as coating of the substratum with extracellular matrix (ECM) components, we were able to perform detailed analysis of cell behavior. Our analysis revealed an important role for the ECM fibril network containing fibronectin as a substratum which guided mesodermal cell migration by contact guidance.
A similar strategy was used for the analysis of mammalian gastrulation. Mesodermal cells isolated from the primitive-streak-stage mouse embryos attached to and migrated on the ECM produced by endothelial cells. This culture system revealed a deficiency in the mesodermal cells isolated from Brachyury (T) mutant embryos. Cell attachment to ECM was further analyzed by using antibodies against fibronectin or laminin and synthetic peptides, or by coating of the culture substratum with various ECM components. These studies indicated that both fibronectin and laminin play roles in adhesion and migration of the mesodermal cells.
For longer range analysis, however, it would be advantageous if isolated embryonic cells could be re-introduced into embryos after manipulations in culture. Genetical manipulation of the isolated cells would produce insight into the molecular basis of embryogenesis, but it requires a long term culture of the embryonic cells without losing the ability to re-integrate into the embryo. Mouse embryonic stem (ES) cells have such characteristics. We introduced a marker gene (Lac Z) into ES cells and analyzed early embryogenesis by producing chimaeric mouse embryos.
KeywordsMouse Embryo Embryonic Cell Inner Cell Mass Primitive Streak Mesodermal Cell
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.
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© Plenum Press, New York 1991