Year 2

Cardiovascular disease is a major concern for medicine and is caused by damage to blood vessels. We have begun a project to generate endothelial cells, the cells which line the blood vessels, from human embryonic stem cells (hESCs) using gene transfer technology and regulated gene expression.

Little is known about the early stages of blood vessel endothelial differentiation of the human embryo. It is imperative that we understand normal development in order to mimic it in the laboratory. We have used hESCs to model embryonic development and determine the pattern of gene expression in the early stages of differentiation. Using what is known about mouse embryonic development as a model, we have determined that gene expression in differentiating human cells closely follows that of differentiating mouse cells. In particular, we have determined the timing of the expression pattern of a gene that is required for the generation of endothelial cells. This knowledge will allow us to induce expression of this gene at the proper time during differentiation in the cells in the laboratory to increase the number of blood vessel cells we can generate.

We have established a method in the laboratory to reliably generate endothelial cells from unmodified hESCs. Timing of gene expression during development is extremely important and improper timing can result in cells being unable to respond to the signal generated by the gene or unable to progress further in development. We have found that introduction of a single factor into the differentiating hESCs results in either as little as two or as much as six times more endothelial cells depending upon the time of administration than in cells without this added factor. These cells behave similarly to cells generated without the addition of the factor in all tests that we have performed on the cells.

To test the ability of the cells that we have generated in the laboratory to aid in human condition, we have been testing mouse models of retinopathy of prematurity (ROP). Premature infants are often placed in a very high oxygen environment to help with their underdeveloped lungs. While this aids their survival, the high levels of can disrupt the vessels in the retina and result in blindness. We are using this model to test the ability of administered hESC derived endothelial cells to aid in the recovery of retinal vessels from exposure to a high oxygen environment. So far we have found that endothelial cells derived from hESCs with and without the addition of the single factor mentioned above result in an improved vessel network in the eyes of tested mice.