Pluripotent stem cells can be created in several ways, depending on the type.

Genetic reprogramming (induced pluripotent cells): Several labs, including that of George Q. Daley, MD, PhD, Director of Stem Cell Transplantation Program, have shown that it requires only a handful of genes to reprogram an ordinary cell from the body, such as a skin cell, into whats known as an induced pluripotent cell (iPS cell). Currently, these genes (Oct4, Sox2, Myc, and Klf4) are most commonly brought into the cell using viruses, but there are newer methods that do not use viruses.

Although skin cells are probably the number-one source of iPS cells currently, lines are also being created from blood cells and mesenchymal stem cells (a type of multipotent adult stem cell that gives rise to a variety of connective tissues). Laboratories in the Stem Cell Program at Childrens Hospital Boston are exploring whether iPS lines made from different kinds of patient cells are easier to work with, or can more readily form the particular kind of cell a patient might need for treatment.

Childrens researchers are also continuing to experiment with more efficient programming techniques, so they can get a higher yield of true pluripotent stem cells.

IVF donations of unused/discarded embryos (ES cells): Another major source of pluripotent stem cells for research purposes is unused embryo donated by couples undergoing in vitro fertilization (IVF). Some of these may be poor-quality embryos that would otherwise be discarded. The resulting cells are considered to be true embryonic stem cells (ES cells).

The donated embryos are placed in a media preparation in special dishes and allowed to develop for a few days. At about the fifth day the embryo reaches the blastocyst stage and forms a ball of 100-200 cells. At this stage, ES cells are derived from the blastocysts inner cell mass. In some cases, the ES cells can be isolated even before the blastocyst stage.

To date, Childrens has created more than a dozen new ES cell lines using this approach, which we are now making available to other scientists. These ES cells are not genetically matched to a particular patient, but instead are used to advance our knowledge of how stem cells behave and differentiate.

Some people question the ethics of using discarded IVF embryos for research. For more discussion, see Policy and Ethics.

Somatic cell nuclear transfer: The process called nuclear transfer involves combining a donated human egg with a cell from the body (typically a skin cell) to create a type of embryonic stem cell, sometimes called an ntES cell. Nuclear transfer requires an egg donor.

First, an incredibly thin microscopic needle is used to remove the eggs nucleus, which contains all the eggs genetic material, and replace it with the nucleus from the body cell. The process of transferring the nucleus into the egg reprograms it, reactivating the full set of genes for making all the tissues of the body. How this happens isnt well understood yet, and researchers in the Stem Cell Program at Boston Childrens Hospital are trying to understand it better.

Next, the resulting reprogrammed cell is encouraged to develop and divide in the lab, and by about day five, it forms a blastocyst, a ball of 100-200 cells. The inner cells of the blastocyst are then isolated to create ntES cells.

Of all the techniques for making pluripotent cells, nuclear transfer is the most technically demanding and therefore the least efficient. To date, it has only been successful in lower animals, not in humans. But because the stem cells created would be an exact genetic match to the patient, nuclear transfer may eliminate the tissue matching and tissue rejection problems that are currently a serious obstacle to successful tissue transplantation. For this reason, nuclear transfer is an important area of research at Childrens.

Because ntES cells created from human patients would match them genetically, nuclear transfer is sometimes called therapeutic cloningnot to be confused with the concept of reproductive cloning.

Parthenogenesis (unfertilized eggs): Using a series of chemical treatments, its possible to trick an egg into developing into an embryo without being fertilized by sperm. This process, called parthenogenesis, sometimes happens in nature, allowing many plants and some animals to reproduce without the contribution of a male.

By inducing parthenogenesis artificially, researchers have been able to create parthenogenetic embryonic stem cells, or pES cells, in mice. The embryos created, known as parthenotes, are grown for about five days until they reach the blastocyst stage. Development is then stopped and pES cells were derived from the blastocysts inner core of cells.

Parthenogenesis hasnt been accomplished in human eggs yet, at least not by choice (a Korean team is thought to have created human pES cells accidentally in 2007). But researchers at Childrens are trying to do so, since pES cells, if carefully typed genetically, could potentially be used to create master banks of pluripotent stem cells. Doctors could then choose a cell line thats genetically compatible with the patients immune system. (For details, see How do pluripotent stem cells get turned into treatments?).

Of more immediate concern is the possibility that parthenogenesis could be used to make pES cells for the egg donor herself or a sibling. However, before using these cells in patients, researchers need to know more about the safety of this approach.

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How Do We Get Pluripotent Stem Cells? | Boston Children's ...

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