[Study on Flk1⁺ Cells during Mouse Early Embryogenesis by Lineage Tracing]

To understand the differentiation of mesoderm-derived Flk1⁺ cells on different locations of the early mouse embryonic development and to explore the potential of Flk1⁺ cells to differentiate into mesenchymal lineage, like pericytes during vascular development.

Based on the Cre-LoxP system conditional knockout study strategy, the Flk1-Cre mice and ROSA26 reporter mice were used for lineage-tracing studies. The fate of the Flk1⁺ progenitor cells was traced with the GFP⁺ population. The detection of mesoderm marker Flk1, hematopoietic cell-specific marker CD45, endothelial cell-specific markers CD31, CD144, and Emcn (endomucin), pericyte specific markers PDGFRβ and NG2, using the methods of immunohistochemistry, immunofluorescence, and flow cytometry should be combined to solve the concerned problems.

Immunohistochemical staining of different fractions of E8.5-10.5 in the early embryogenesis of Flk1-Cre; ROSA26-EYFP mouse lineage showed that there were multiple populations in the Flk1⁺ cell-derived GFP⁺ population surrounding hematopoietic sites, such as dorsal aortas, limb buds and yolk sac. In addition to hematopoietic cells, the CD31⁺/Emcn⁺ typical endothelial cells distributed specifically along the blood vessel wall, there were many types of cell populations, such as mesenchymal-like cells. The immunofluorescence demonstrated that the cells of this group are neither hematopoietic, non-endothelial cells around the blood vessels, which are NG2+ pericytes. FACS analysis also confirmed that Flk1⁺ cells contributed to pericytes. In addition, in different hematopoietic sites of the embryo, a small population of CD31+CD140B+ cell populations with a mesenchymal-like morphology was observed in the GFP⁺ population.

In the early stages of embryogenesis, mesoderm-derived Flk1⁺ populations not only contribute to hematopoietic, endothelial, and muscle lineages, but also have a differentiation potential for mesenchymal lineage, like pericytes. The presumably observed CD31⁺CD140B⁺ cell population may be a group of endothelial cells differentiated from Flk1⁺ progenitor cells and undergoing an endothelium-to-mesenchymal transition, EndMT, gradually losing the endothelial surface-specific marker and also starting to express a pericyte surface-specific marker.