Stem Cell Program | Pluripotent Stem Cell Research

Scientists have discovered ways to take an ordinary cell, such as a skin cell, and reprogram it by introducing several genes that convert it into a pluripotent cell. These genetically reprogrammed cells are known as induced pluripotent, or iPS, cells. The Stem Cell Program at Boston Children's was one of the first three labs to do this in human cells, an accomplishment cited as the Breakthrough of the Year in 2008 by the journal, Science.

iPS cells offer great therapeutic potential, because they come from a patient’s own cells. They are genetically matched to that patient, so they can eliminate tissue matching and tissue rejection problems that currently hinder successful cell and tissue transplantation. iPS cells are also a valuable research tool for understanding how different diseases develop.

Because iPS cells are derived from skin or other body cells, it is sometimes preferred over embryonic stem cells from embryos or eggs. This process must be carefully controlled and tested for safety before it’s used to create treatments. In animal studies, some of the genes and the viruses used to introduce them have been observed to cause cancer. Further research is also needed to make the process of creating iPS cells more efficient.

iPS cells are under further investigation at Boston Children’s in the lab of George Q. Daley, MD, PhD, Director of Stem Cell Transplantation Program — who reported creating 10 disease-specific iPS lines, which is part of a growing repository of iPScell lines.

Scientists use “embryonic stem cell” as a general term for pluripotent stem cells that are made using embryos or eggs, rather than for cells genetically reprogrammed from the body. There are several types of embryonic stem cells:

Perhaps the most widely-known type of pluripotent stem cell, made from unused embryos that are donated by couples who have undergone in vitro fertilization(IVF). The IVF process — in which the egg and sperm are brought together in a lab dish — frequently generates more embryos than a couple needs to achieve a pregnancy.

These unused embryos are sometimes frozen for future use, sometimes made available to other couples undergoing fertility treatment, and sometimes simply discarded, but some choose to donate them.

Pluripotent stem cells made from embryos aren’t genetically matched to either parent, and are unlikely to be used to create cells for treatment. They are used to advance our knowledge of how stem cells behave and differentiate.

The term somatic cell nuclear transfer (SCNT) means the transferring of nucleus from a somatic cell — any cell of the body — to another cell. This type of pluripotent stem cell, sometimes called an ntES cell, has only been made successful in animals under lab conditions. To make ntES cells in human patients, an egg donor would be needed, as well as a cell from the patient (typically a skin cell).

The process of transferring a different nucleus into the egg reprograms it to a pluripotent state, reactivating the full set of genes for making all the tissues of the body. The egg is then allowed to develop in the lab for several days, and pluripotent stem cells are derived from it.

Like iPS cells, ntES cells match the patient genetically. If created successfully in humans, and if proven safe, ntES cells could completely eliminate tissue matching and tissue rejection problems. For this reason, they are actively being researched at Boston Children’s.

Through chemical treatments, unfertilized eggs can develop into embryos without being fertilized by sperm, a process called parthenogenesis. The embryos are allowed to develop in the lab for several days, and then pluripotent stem cells can be derived from them.

If this technique is proven safe, a person may be able to donate their own eggs to create pluripotent stem cells matching them genetically that in turn could be used to make cells that wouldn’t be rejected by their immune system.

Through careful genetic typing, it may be possible to use pES cells to create treatments for patients beyond the egg donor themselves by creating “master banks” of cells matched to different tissue types. In 2006, working with mice, Boston Children’s researchers were the first to demonstrate the potential feasibility of this approach.

Because pES cells can be made more easily and more efficiently than ntES cells, they could potentially be ready for clinical use sooner. However, more needs to be known about their safety. Concerns have been raised that tissues derived from them might not function normally.

The work of 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 what’s 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 multi potent adult stem cell that gives rise to a variety of connective tissues). Laboratories in the Stem Cell Program 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.

Researchers are also continuing to experiment with more efficient programming techniques to get a higher yield of true pluripotent stem cells.

Another major source of pluripotent stem cells for research purposes is unused embryo donated by couples under going 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 blastocyst’s inner cell mass. In some cases, the ES cells can be isolated even before the blastocyst stage.

To date, Boston Children’s 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.

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 egg’s nucleus, which contains all the egg’s 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 isn’t well understood yet, and researchers in the Stem Cell Program 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 Boston Children’s.

Because ntES cells created from human patients would match them genetically, nuclear transfer is sometimes called therapeutic cloning — not to be confused with the concept of reproductive cloning.

Using a series of chemical treatments, it’s 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 are derived from the blastocyst’s inner core of cells.

Parthenogenesis hasn’t 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 Boston Children’s 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 that’s genetically compatible with the patient’s immune system. 

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