BioWorld International Correspondent

LONDON - Researchers who want to use differentiated cells derived from human embryonic stem cells to treat disease and assess new drug treatments have overcome a major hurdle that has until now stood in their way.

A team at the University of Nottingham in the UK has developed a strategy that allows it to prepare pure populations of differentiated cells from the mixture of cell types that generally results after differentiation of human embryonic stem cells.

The team has shown that it can purify cardiomyocytes (heart muscle cells) from a heterogeneous mix of differentiated cells produced from stem cells. The purification method does not adversely affect cardiomyocyte function. Tests have shown that the behavior of purified and non-purified cardiomyocytes is the same.

Chris Denning, lecturer in stem cell biology at the Wolfson Centre for Stem Cells, Tissue Engineering & Modelling (STEM), at the University of Nottingham, told BioWorld International: "We now have the ability to produce virtually pure cultures of cardiomyocytes from human embryonic stem cells, which was not possible before. This is a vitally important advance, as only by having such pure populations will we be able to use these cells for applications such as drug screening, toxicology and transplantation into patients."

An account of the work appears in the Sept. 25 issue of the journal Molecular Therapy in a paper titled: "Transgenic Enrichment of Cardiomyocytes from Human Embryonic Stem Cells."

Denning and his colleagues have been working for some time on producing cardiomyocytes from human embryonic stem cells. He said: "The disappointing aspect has been that although researchers can get beautiful beating clusters of cells in a culture dish, only 1 to 20 percent of the cells are actually heart cells. The majority of the cells are other types, such as brain cells, liver cells and so on."

To address the problem, team members devised a genetic selection strategy. They engineered human embryonic stem cells to include a promoter that would only be switched on in cells that became cardiomyocytes. The promoter, in turn, was linked to a gene encoding a protein that conferred resistance to a drug. When they added the drug to a culture of mixed differentiated cells, therefore, only the cardiomyocytes would have the appropriate enzyme to break down the drug and survive.

"Using this strategy, we can get close to 100 percent purity of heart cells," Denning said. "We have also gone on to demonstrate that these drug-selected cells are completely functional in terms of their cardiac activity. Their electrophysiology is indistinguishable from the cardiomyocytes that have not been through the drug selection procedure."

Having pure cultures of cardiomyocytes means that scientists now can go on to do many of the experiments that previously they only had been able to talk about.

Denning pointed out, for example, that little is known about the molecular biology of many human genetic diseases that affect muscle and heart muscle. That is partly because the phenotypes of mice in which the relevant genes have been knocked out do not always correspond to the human disease phenotypes.

"In such diseases, which include Duchenne muscular dystrophy," he said, "we would like to be able to recreate the disease in the culture dish. This would allow us to understand more about the biology of the disease and consider ways of devising new treatments."

The group already has begun testing approved drugs on their purified cardiomyocyte cultures. Using equipment that allows them to measure the electrophysiology of the cells over long periods of time, they have, for example, added drugs that are already in clinical use, and some that have been banned because they induce electrophysiological abnormalities such as a long Q-T syndrome.

"We have been trying to validate the effects of these drugs on the cultured cells, and we are linking up with some industrial partners to evaluate the effects of novel heart drugs," Denning said.

In the long term, the group would like to progress to using the cells therapeutically, for example to treat patients who have had heart attacks. Before scientists can achieve this goal, he said, two further hurdles need to be overcome. One is how to produce enough cells to be able to treat a patient. Although some laboratories currently can produce more than 1 million cardiomyocytes at a time, the number required to treat a heart attack patient will, Denning suggested, be more like 1 billion.

Secondly, there will need to be some way of ensuring that the cells used stay in the place where they are needed. "We are working with our tissue engineering colleagues within the STEM Centre to develop different matrices that we can load the cells onto, and transplant into a human heart," he added.

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