Blood island formation in attached cultures of murine embryonic stem cells

Differentiation of murine embryonic stem cells in suspension culture results in the formation of cystic embryoid bodies that develop blood islands. In this study pre‐cystic embryoid bodies were attached to a substratum, and the program of differentiation was monitored. The attached ES cell cultures formed blood islands on a cell layer that migrated out from the center of attachment and beneath a mesothelial‐like cell layer. Morphological and in situ marker analysis showed benzidine‐positive hematopoietic cells surrounded by vascular endothelial cells that expressed PECAM and took up DiI‐Ac‐LDL. Waves of morphological differentiation were evident, suggesting a graded response to differentiation signals. Electron microscopy of the blood islands showed that they were similar to blood islands of cystic embryoid bodies and mouse yolk sacs, and cell‐cell junctions were evident among the blood island cells. RNA expression analysis was consistent with the presence of hematopoietic precursor cells of several lineages and a primitive vascular endothelium in the cultures. Thus a program of vascular and hematopoietic development can be elaborated in attached ES cell cultures, and these blood islands are accessible to experimental manipulation. © 1996 Wiley‐Liss, Inc.

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