In November 1998, the world learned that human embryonic stem cells had been successfully grown in cell culture. The discovery was hailed as a "medical revolution," for here at last was a source of undifferentiated human cells that have the potential to develop into any cell type in the body. People with illnesses or injuries requiring the replacement of tissue—any tissue—need not worry about finding a suitable donor. People need no longer suffer the consequences of heart attacks, Alzheimer's disease or diabetes when compatible heart tissue, neurons and pancreatic cells can be grown in a culture dish. In theory, stem-cell technology could fulfill such a vision.

There is a problem, however. Stem cells were culled from human embryos and fetuses that had the potential to develop into complete human beings. The fear is that research using human embryonic tissue could pull medicine down a slippery slope to a world where unborn human beings are harvested for the cells they could provide to another human being. It's a ghoulish image that conjures up the public's worst fears of science. On the other hand, the human embryonic stem cells that have already been collected may be a sufficient source of cells for future replacement tissue, so that no new embryos are required.

As with many issues raised by the development of new technologies, there is no simple answer to the question of how this research should proceed. Reasonable people on both sides of the argument disagree. As this magazine goes to press, the National Bioethics Advisory Commission (NBAC) is undertaking an extensive review of the ethical, legal and medical issues associated with human embryonic stem-cell research. Their recommendations are slated to appear early this summer. This research is of such fundamental importance that all responsible citizens should be aware of its implications. Here I provide a brief primer that explains the current state of the science and the ethical questions raised by the work.

What Are Embryonic Stem Cells?

Embryonic stem cells form at a very early stage in human development and remain in an undifferentiated state for only a short period of time. They are first clearly recognizable about five to seven days after fertilization, when a human embryo forms a structure called a blastocyst. Consisting of merely 140 cells, this hollow, fluid-filled sphere gives little indication that it could develop into a complete person. There are two types of cells in the blastocyst at this stage. Trophoblast cells, which form the "shell" of the sphere, will become the supporting tissues of the fetus such as the placenta. Inner-cell-mass cells, which are located at one end within the blastocyst, are the undifferentiated cells that will divide and develop into the individual.

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The inner cell mass begins to differentiate when it forms the three fundamental germ layers of the embryo, known as the ectoderm, the mesoderm and the endoderm. Each of these germ layers has a specific destiny as a particular set of tissues in the mature individual, and all organs are ultimately derived from them. The ectoderm will form the skin, the eyes and the nervous system. The mesoderm will form bone, blood and muscle tissue. And the endoderm will develop into the lungs, liver and lining of the gut. Because the inner cell mass gives rise to a complete individual, these cells are considered to be totipotent. When inner-cell-mass cells are cultured in a dish they are called embryonic stem cells.

Isolated mouse embryonic stem cells have been seen to differentiate into various types of cells, including neural cells, hematopoietic cells (the precursors of blood cells) and cardiomyocytes (heart-muscle cells). Remarkably, these cells have a tendency to develop spontaneously into primitive versions of certain structures. For example, under standard culture conditions a certain proportion of the cells will differentiate into embryoids (which bear an eerie resemblance to a small beating heart), and a certain proportion will develop into yolk sacs containing hematopoietic cells. The relative proportion of embryoids or yolk sacs that are produced can be altered by modifying the culture medium, but to date no one has been able to induce the cells to develop into a pure population of differentiated cells.

In principle, mouse embryonic stem cells have the capacity to develop into a functional organ, but this has yet to be seen in the culture dish. Some remarkable structures do form, however, when embryonic stem cells are transplanted into mice with an immune-system disorder (severe combined immunodeficiency, or SCID), which prevents the mouse's body from rejecting the transplanted cells. When placed into a SCID mouse, mouse embryonic stem cells have developed into muscle, cartilage, bone, teeth and hair.

Despite their potential, isolated embryonic stem cells cannot develop into a mouse if returned directly to the uterus because the cells have lost their capacity to form trophoblast cells, which are necessary for implantation. Under these conditions they are considered to be pluripotent, rather than totipotent. However, if the embryonic stem cells are first added to a donor tetraploid embryo that is unable to develop normally and the resulting embryo is transferred to a mouse uterus, a normal mouse is born that is totally derived from the cultured embryonic stem cells. This indicates the incredible totipotent properties of these cells.

Human Embryonic Stem Cells

Since 1978, the federal government has banned the use of federal funds for human embryo research. However, the Public Health Service Act authorizes federal funding of human fetal-tissue research and provides conditions for its conduct. Members of several scientific societies, such as the Society for Developmental Biology and the Federation of American Societies for Experimental Biology (FASEB), have claimed a voluntary moratorium on human embryo research. In 1994 the Human Embryo Research Panel presented a report to the director of the National Institutes of Health (NIH) suggesting that research deriving human embryonic stem cells is acceptable as long as embryos are not created expressly for research purposes. A broadly written federal law bans the use of federal funds for the derivation of human embryonic stem cells [Public Law 105-78, Section 513(a)], but it is legal for private funds to be used to perform human embryo research.

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Private funds from Geron Corporation in Menlo Park, California are currently supporting three laboratories engaged in human embryonic stem-cell research: John Gearhart's team at Johns Hopkins University, Roger Pedersen's team at the University of California, San Francisco, and James A. Thomson's team at the University of Wisconsin, Madison. All three laboratories have strictly segregated their research on human embryonic stem cells so that it is supported only by the private funds.

Gearhart's laboratory has used a novel approach to obtain undifferentiated human cells. Rather than relying on the inner cell mass of the blastocyst, Gearhart reasoned that primordial germ cells (the precursors of egg and sperm cells) should be undifferentiated, much like embryonic stem cells. Embryonic germ cells are derived from embryos or fetuses when they are about five to nine weeks old, when the undifferentiated cells are migrating from the yolk sac to the gonads. In his experiments, Gearhart's team acquired the cells from embryos and fetuses that were aborted for therapeutic reasons. When grown on a "feeder" layer of mouse fibroblasts (which release factors that inhibit stem-cell differentiation), embryonic germ cells have properties similar to embryonic stem cells: They can be cultured for at least seven months without differentiating, they retain normal chromosome structure, and when properly stimulated they can differentiate into embryoids similar to those observed in mouse embryonic stem-cell cultures. The embryoids contain several types of differentiated cells including derivatives of all three germ layers. Thus, human embryonic germ cells are considered pluripotent.

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Roger Pedersen's and James Thomson's research groups acquired human embryonic stem cells from the inner cell mass of the blastocyst. These experiments have proved to be extremely challenging. Obtaining human embryonic stem cells from blastocysts is difficult because the embryos are acquired as "leftovers" from in vitro fertilization clinics. The quality of these embryos is usually poor because the attending physician implants the best embryos into the mother. Moreover, blastocysts are in limited supply, and few are donated for research as it costs about $10,000 to produce a batch by in vitro fertilization. There is an added technical problem because it is difficult to get in vitro fertilized human embryos to develop to the blastocyst stage.

Despite these difficulties, Thomson's laboratory (which is very experienced with non-human primate embryonic stem cells) has met with much success. They obtained human embryos produced by in vitro fertilization from couples who provided informed consent. Of 14 inner-cell masses (representing 14 embryos of both sexes), five embryonic stem-cell lines have been established. All five cell lines appear to retain the characteristics of human embryonic stem cells. Their pluripotency is demonstrated by experiments in which the stem cells are injected into SCID mice. In each instance the injected cells form a "teratoma" consisting of cells derived from all three germ layers. These stem-cell lines also have the capacity to renew themselves while remaining undifferentiated. This is because the cells have high levels of telomerase, an RNA-dependent DNA polymerase enzyme that allows cells to divide indefinitely. Cells that do not express telomerase have a limited capacity to replicate. In principle Thomson's cell lines provide a virtually unlimited supply of normal, pluripotent human embryonic stem cells.

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It is unclear whether these pluripotent cells are also totipotent, because the only way to prove totipotency is to derive a human being from the cells. This has not been done for ethical and legal reasons. Many scientists claim that human embryonic stem cells cannot develop into a human being if implanted directly into a woman's uterus because the cells have lost the ability to develop into trophoblast cells. However, if the same technology that is used for mouse embryos is applied to human embryos, the embryonic stem cells could be repackaged with trophoblast cells from a donor human embryo. In theory, this embryo could develop into a normal human being if implanted into a woman's uterus.

Funding the Research

Because of the regenerative capacity of human embryonic stem cells, the currently established cell lines could be used to supply other scientists without the need to collect new embryos or fetuses. In light of the legislative ban on supporting human embryo research with government funds, Harold Varmus, director of the NIH, asked for a legal opinion regarding the use of these cell lines from the NIH's cabinet-level parent agency, the Department of Health and Human Services (DHHS), on December 2, 1998. After a thorough discussion of the law and the human embryonic stem-cell research, the DHHS concluded that "the congressional ban on human embryo research does not apply to research on human embryonic stem cells because the cells are not an embryo as defined by statute." In addition, "since the human embryonic stem cells do not have the capacity to develop into a human being" on their own if implanted into a woman's uterus, "they cannot be considered human embryos." Therefore research using pluripotent stem cells derived from human embryos can be funded by the DHHS.

With regard to the human embryonic germ cells, "the legal opinion indicated that because the embryonic germ cells are derived from non-living fetuses, they fall within the legal definition of human fetal tissue." Thus the use of embryonic germ cells is subject to the federal restrictions on fetal-tissue research, and work that generates and uses pluripotent stem cells from non-living fetuses can also be supported by the DHHS.

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The NIH plans to fund research using human embryonic stem cells. However, the NIH assures that no government funds will be used for the creation of human embryos expressly for research purposes. Although the NIH understands and respects the compelling ethical, legal and moral issues surrounding human pluripotent stem-cell research, it has also asked Congress, the NBAC and the Council of Public Representatives for additional advice and will not fund any human embryonic stem-cell research until guidelines are developed and disseminated to the research community. In addition, President Clinton has asked the NBAC to undertake a thorough review of the issues associated with human stem-cell research, including ethical, legal and medical considerations (as it had done with the human cloning issue in 1997).

As reported in recent issues of The New York Times (January 20, 1999, page A29 and February 17, 1999, page A12), reaction to the DHHS ruling has been mixed. A group of 73 scientists, 67 of them Nobel laureates, support the DHHS position, calling it both laudable and forward-thinking (Science 283:1849). The American Society for Reproductive Medicine praised the decision. In a letter on December 2, 1998, FASEB President William R. Brinkley of the Baylor College of Medicine strongly urged members of the Senate Labor, Health and Human Services Appropriations Committee to allow federal funding of the stem-cell research to continue because of the tremendous medical potential. He is pleased about the ruling but at the same time warns that we should recognize the complex ethical issues involved and the need to examine them fully. Other scientific societies, including the American Society for Cell Biology and the Society for Developmental Biology, also support the ruling on Federal funding for human embryonic stem-cell research. Senator Tom Harkin, an Iowa Democrat, called for speedy issuance of the ruling and indicated that the decision would bring scientists closer to finding cures for many diseases, saying that "the government should not issue blanket bans on medical research." Harold Varmus indicated that "the prospect of doing amazingly interesting science is really quite wonderful." However, he cautioned that it would still be illegal for scientists to use federal funds to derive their own embryonic stem cells. On the other hand, they can now use federal monies to work on the cell lines already obtained by Thomson and Gearhart, if their projects are funded by NIH.

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Other responses have not been so favorable to the new ruling. Richard M. Doerflinger, associate director for policy development at the Secretariat for Pro-Life Activities at the National Conference of Catholic Bishops, denounced the ruling as a loophole that violates the spirit of current law: "[T]hey will destroy the embryos with private funds and experiment on the tissue with public funds." On February 16, 1999, 70 members of the House of Representatives asked to rescind the ruling allowing federal money for research on human embryonic stem cells in a letter to Donna E. Shalala, the Secretary of Health and Human Services. The letter indicated that "the ruling would violate both the letter and spirit of the Federal law that bars Government support for research in which human embryos are destroyed." Other people reacted to the research itself. Judy Brown, president of the American Right to Life League, protested the use of stem cells since they were taken from potential human beings who should be protected by law (Time, November 16, 1998, page 96).

Several people, including Harold Varmus, are troubled that stem-cell research is performed only by private companies, because the oversight contributed by an institutional review board for NIH-supported research is omitted from the process. The oversight is designed to analyze the scientific merits of the research and to ensure that human and animal subjects are protected. Although private companies have their own review boards, they are not required to follow NIH guidelines. Some also worry that if biotechnology companies alone support the research, independent verification of the results will not be possible. This could result in medical products that may be marketed before their effects are fully known. Others argue that the review of NIH-funded research by an institutional review board is not sufficient, because NIH itself commercializes research by patenting discoveries made by its scientists and by encouraging scientists to enter into joint ventures with private companies. Some suggest that more federal regulations are needed to address the debate on stem-cell research. According to Lori Andrews, director of the Institute for Science, Law and Technology at the Illinois Institute of Technology, "In 1997, Senator John Glenn introduced a bill to extend federal protection to the subjects of research supported by private funds. Ironically, the bill died in committee in October, just before the stem-cell debate began." (The Chronicle of Higher Education 45(21):B4–B5)

An Alternative?

It may be possible to produce made-to-order, person-specific, pluripotent embryonic stem cells using somatic-cell nuclear-transfer technology, the technology used to produce the cloned sheep Dolly. With this method, any human somatic cell, such as a cheek cell, could be fused with an enucleated donor human egg cell that is then stimulated to develop into a blastocyst embryo. The inner-cell-mass cells could then be harvested from the blastocyst. A specific tissue could then be grown from the embryonic stem cells, which could be implanted into the original donor. The advantage of the nuclear-transfer method is that the embryonic stem cells would be compatible with the donor of the somatic cell. This would prevent any graft rejections due to immune-system incompatibility. To date no one has derived embryonic stem cells using this technique, but the feasibility of this approach has been demonstrated.

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In 1996 Jim Robl, of the University of Massachusetts, and Jose Cibelli, now at Advanced Cell Technology in Worcester, Massachusetts, fused one of Cibelli's cheek cells with an enucleated cow egg using nuclear-transfer technology. The resulting hybrid, or chimeric, embryo successfully divided at least five times to the 32-cell stage, but was not allowed to develop further. In principle, if the chimera were allowed to develop to the blastocyst stage, it could be used to harvest embryonic stem cells.

Could such an embryo develop into a person? The nuclear DNA certainly would direct the embryo's cells to make human proteins, but the cells would contain both human and bovine mitochondria, which have their own DNA. Because mitochondria express DNA primarily for mitochondrial function in the production of energy, it is thought that they do not contribute greatly to the overall appearance of the cell. However, the cell's nucleus contributes proteins for mitochondrial function. If human proteins are incompatible with bovine mitochondria, presumably the human mitochondria would then provide the cell's energy needs. If the bovine cytoplasm acts mainly as a source for nutrients and energy, the human DNA should direct the cells to have the characteristics of a human cell. Based on research with other organisms, the cells should have a human appearance. However, there is insufficient evidence to determine whether such an embryo could provide viable embryonic stem cells and whether it could develop into a child.

Those who support the cow/human-chimera research claim that state and federal bans on embryo research are avoided because these laws only apply to the "product of fertilization," and the nuclear-transfer technique does not involve fertilization or human eggs. Others argue that because the embryos contain some bovine DNA they are not human embryos. There is insufficient scientific evidence to know whether such an embryo could develop into a person, and therefore it is unclear whether the cow/human chimera could be considered human. A possible use of such a chimeric embryo is that it may provide a means to overcome rejections of transplanted tissues without creating human embryos.

President Clinton is ". . . deeply troubled by the news of experiments involving the mingling of human and non-human species." As a result, the President has asked members of the NBAC to investigate the bioethical, medical and legal ramifications of the research and report back to him as soon as possible. The report is expected early this summer.

The Potential

If scientists are ultimately successful in inducing human embryonic stem cells to routinely differentiate into specific cell types, the potential applications are enormous. Any disorder that involves the loss of normal cells could be targeted for tissue-transplantation therapy. New neurons could be grown for neurodegenerative disorders such as Parkinson's disease and Alzheimer's disease, new pancreatic cells for diabetics, and new cardiac muscle for rebuilding the heart.

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Human embryonic stem cells could also be genetically altered to be universal donors, so that the resulting graft tissue will not be rejected. A bank of cell types could be generated for transplantation medicine. Embryonic stem cells could be grown as universal graft tissue for blood, bone marrow, lung, liver, kidney, tendons, ligaments, muscle, skin, hair, teeth, the retina and the lens of the eye. The possibilities are endless.

Another way to prevent graft rejection is to substitute the embryonic stem-cell nucleus for a nucleus from a healthy cell of the patient. The resulting graft tissue would genetically match the patient's tissue. This would be especially important for organ transplantation. The modified embryonic stem cells could be grown into a new organ that is derived from the genetic instructions provided by the patient's donor cell nucleus. The trick would be to induce the cells to become a functioning organ. Some companies (such as Advanced Cell Technology) are currently working with pig and cow embryonic stem cells as sources of animal organs for human transplants. They are trying to genetically alter the embryonic stem cells so that the tissues derived from them are compatible with the human immune system. The resulting animals could then be used for human transplants. To date, transplanting hearts or kidneys from pigs into people has been unsuccessful because the human immune system recognizes the tissue as foreign. By using genetically engineered embryonic stem cells to generate embryos, these grafts may become successful.

Another exciting application of human embryonic stem cells is in the development of pharmaceuticals. The cells would provide a means to screen and test potential new drugs and toxicological agents. The standard methods use traditional cell lines or cells from other species, but these do not always give a true indication of the normal human cell's response to the chemicals. By using human embryonic stem cells, the numbers of animals required for drug testing would be greatly reduced. Human embryonic stem cells may also provide a means to study the progression of human disease and find effective, permanent treatments. Human embryonic stem cells will also be useful for studying the normal differentiation process to enhance our understanding of human development.

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It is important to keep in mind that these potential applications require further research with the embryonic stem cells before any of them are realized. One of the big hurdles—culturing the cells—has been overcome. Major difficulties remain, however, such as finding the magic combination of growth factors that will induce the cells to differentiate into a specific tissue type. Some believe that it will be sufficient if the embryonic stem cells are induced to specialize into a precursor cell. Transplanting the precursor cells into the correct environment should be sufficient to generate the required tissue type, because the body has all the factors necessary to do this, and scientists do not need to know exactly what the factors are. Another difficulty will be engineering a universal donor tissue type to prevent graft rejections. It should be possible to swap the embryonic stem cell nucleus with the patient's own cell nucleus, but it is unclear if the resulting cell would still behave like an embryonic stem cell. Transferring the patient's nucleus into a donor egg would produce embryonic stem cells with a genetic composition that matches the patient's tissues, but here we are limited by the availability of donor eggs. It is also unclear whether the technique would induce chromosomal abnormalities. Moreover, further research is required to determine whether differentiated embryonic stem cells are prone to cancer.

How long will it be before human embryonic stem cells are used for replacement tissue? Ethical, religious and legal issues aside, some claim that significant advances will be seen within the next ten years.

Ethical Concerns

The tremendous medical potential of human embryonic stem cells notwithstanding, there are serious concerns about setting the boundaries for this research. Many questions surround the source of the cells and their potential applications. For example, human embryonic stem cells derived from in vitro fertilized embryos would develop into a child if given the chance. What if the cells are derived from a pregnancy that was intentionally terminated? Is it ethical to use these cells when an embryo was killed to obtain them? Does a good end justify an evil means? Is it right to use the cells obtained from an embryo that was spontaneously aborted or killed in an accident? Several human embryonic stem cell lines exist now. They may be sufficient to supply all the embryonic stem cells that will be needed. Should these cells be used to benefit us now that they have been collected? Some suggest that adult stem-cell research should be pursued further and embryonic stem-cell research should be abandoned. Unfortunately, the versatility of adult stem cells is limited.

Some argue that it is unethical to harvest embryonic stem cells from human embryos because human life has not been respected. Others claim that since scientists have not killed the cells, only changed their fate, it is ethical to use them. Some fear that companies will support in vitro fertilization clinics and abortion clinics for the acquisition of embryos and aborted fetal tissue, assuming more cells will need to be collected.

If the embryonic stem cells and embryonic germ cells became commercially available as a cell line, would it be ethical for scientists to use them? What type of research would be acceptable? Should scientists be allowed to grow individual tissues and organs to study the process of development or to build medical replacement tissue? Since it is currently acceptable for human genes to be inserted into non-human animal cells, is it ethical to insert human embryonic stem cells into domestic animal embryos to create a chimera for the purposes of harvesting human organs for transplantation? Is it ethical for human embryonic stem cells from an embryo with a genetic disease (such as cystic fibrosis) to be genetically altered for therapeutic purposes and allowed to continue development into a healthy newborn? Potential ramifications of using the cells reach into adulthood as well. If human replacement tissues become readily available, do people need to live responsibly any more? Might we open the door for people to engage in high-risk behaviors?

These are not easy questions to answer. As a society we must identify the ethical, social, legal, medical, theological and moral issues that surround this research. People from all walks of life—scientists, lawyers, ethicists, clergy and the general public—should be involved in making the decision. We are also at a crossroads where further scientific evidence is needed to explore the full potential of these cells, and yet many of the necessary experiments raise further ethical issues. The question of how we should use these powerful cells remains a challenging problem for the next century.