MEDICINE

A new era for stem cells

The news that human skin cells had been converted into stem cells hit the world headlines in late 2007. Hailed by the experts as the Holy Grail, this discovery has revived both hope for new medical therapies and controversy over the legitimacy of research using embryonic stem cells.

© Jeff Miller/University of Wisconsin In 1998, James Thomson from the University of Wisconsin-Madison (USA), led a research group that isolated the first human embryonic stem cells. In 2007, working with a Japanese team, his laboratory successfully extracted the first pluripotent cell lines from human skin. © Jeff Miller/University of Wisconsin
© Lay Glennon/Thomson Lab/University of Wisconsin-Madison As yet undifferentiated embryonic stem cells (shown in blue, in the centre). They represent the very first stage of cell development and can later turn into 22 different types of functional cell throughout the human body. © Lay Glennon/Thomson Lab/University of Wisconsin-Madison
© University of  Wisconsin-Madison Seen under a microscope, the colonies of embryonic stem cells appear as dense, rounded masses. The longer, flatter cells with which they are enmeshed (fibroblasts) act as feeder cells to enable them to develop. © University of Wisconsin-Madison
© Su-Chun Zhang/University of Wisconsin-Madison Neural precursor cells are grown in the laboratory from embryonic stem cells, creating already mature neurons (shown in red) and glial cells (in green). © Su-Chun Zhang/University of Wisconsin-Madison

In late November 2007, a mere handful of the world’s news media failed to report the major breakthrough announced simultaneously by two research teams, one American and the other Japanese. There could not have been more of a stir if the scientists had claimed to have turned lead into gold. Both groups of researchers had managed to convert human skin cells into stem cells, opening up potentially unlimited opportunities for replacing damaged tissues and organs. The new technique allows stem cells to be produced without destroying embryos. This is important, as the idea of sacrificing human lives for scientific research has been an issue of heated debate for years.

But how did the researchers manage to rejuvenate skin cells to make them so akin to embryonic stem cells? Although the techniques employed by the two teams were similar, they differed in a number of aspects.

Four rejuvenating genes

Shinya Yamanaka’s team, from the University of Kyoto in Japan, worked on skin cells from the face of a 36-year-old woman. After collecting these fibroblasts, the scientists injected them with genes coding for four transcription factors. This genetic manipulation enabled them to activate certain genes in the skin cells that are normally active only at the embryonic development stage. This tricks the fibroblasts into believing that they are unspecialised embryonic stem cells. The means used by the Japanese team to trick the skin cells were: Oct3/4 and Klf4, two transcription factors involved in maintaining the pluripotency of stem cells (i.e. their ability to differentiate into any type of foetal or adult cell); Sox2, a protein present in embryonic stem cells; and c-Myc, which is essential for cell growth and division as well as for slowing cell differentiation.

Shinya Yamanaka and his team succeeded in producing a stem cell line from 5 000 skin cells. “Although such efficacy may seem low, it means that multiple lines of pluripotent stem cells can be obtained from a single 10-centi - metre sample,” explains Shinya Yamanaka. The results of this study were published in the journal Cell (1) in November 2007.

Across the Pacific, the recipe was a little different. While James Thomson’s team also used genes coding for four transcription factors, they did not use the same candidate proteins as the Japanese team. Although the University of Wisconsin-Madison researchers also used Oct3/4 and Sox2, they chose NANOG and LIN28 for their other two transcription factors. Another difference between the two teams was that Thomson’s worked on cells from the foreskin of a newborn and succeeded in rejuvenating one cell in every 10 000. Their results were published in the journal Science(2).

So, researchers in both Japan and the United States have managed to reprogramme cells that are already fully differentiated. Both teams used a retrovirus to introduce selected genes into skin cells.

These two techniques make it possible to produce stem cells that contain the patient’s own genetic information. This would be a significant advantage in avoiding the risk of rejection in transplant patients. However, there is one point in favour of the American technique. “The big difference between the two teams is that the Japanese researchers used c-Myc, which is a cancer gene,” explains Daniel Brison, co-Director of the North West Embryonic Stem Cell Centre at the Universityof Manchester (UK). “As the Americans did not use c-Myc, their cells are clinically more advantageous.”

What’s the situation on using embryos?

The stem cells obtained from reprogramming human adult cells are called “induced pluripotent stem cells” (iPS). Their basic physical, genetic and biological properties are very similar to those of embryonic stem cells (ES cells). “However, years of research are needed before we know whether they are as effective as embryonic stem cells or for certain differences to become apparent. Embryonic stem cells have been studied for more than 20 years in mice, and 10 years in humans,” says Daniel Brison.

Although the results of these recent iPS studies are of enormous biological and clinical interest, the research has also triggered an ethical debate. What is the advantage of such a technique over current techniques? While some people present iPS as the solution to many ethical issues posed by embryonic stem cells and therapeutic cloning, not all the experts are in agreement. “Stem cells that have been reprogrammed from adult cells are more interesting from every standpoint”, argues Jonas Frisén from the Department of Cell and Molecular Biology (CMB) of Sweden’s Karolinska Institutet. “The technique doesnot require the use of embryos and produces cells that are genetically identical to the patient’s own cells.”

North West Embryonic Stem Cell Centre co-Director, Daniel Brison, is less convinced of the advantages of iPS: “The embryos used to produce stem cells are surplus embryos creatresearch ed for in vitro fertilisation treatments. They are doomed in any case. Embryonic stem cells are still the most natural source of pluripotent cells because they are able to form a heart, muscles, brain tissues and so on, which adult skin cells are not supposed to do.” Daniel Brison also believes that, even though therapeutic cloning has not yet been tested in humans, it is a more natural method than iPS. “The egg is the normal environment in which the nucleus is programmed to be pluripotent. Although therapeutic cloning is unnatural in that it involves placing an adult cell nucleus into an egg to allow it to be reprogrammed, it is still a more natural method than one using iPS, which are genetically modified cells.” Daniel Brison even predicts a public outcry against iPS-derived therapies, as happened with genetically modified food.

Scotsman, Ian Wilmut, who cloned Dolly the sheep in 1997, expresses a contrasting view. He finds the new technique highly promising and has decided to abandon his research on embryo cloning to devote himself to iPS, in the belief that it heralds “a new era” for biology.

Sparking a whole new ethical debate…

One of the chief advantages of reprogramming skin cells is its simplicity. Any standard laboratory can manipulate the four genes needed to make skin cells regress into stem cells. Furthermore, skin cells are much easier to collect than embryos.

The researchers who made this extraordinary but controversial discovery are cautious, however. “The research has only just begun and we have very little idea how these cells function”, explains James Thomson, leader of the American research team, who still considers embryonic stem cells to be the research gold standard. Japanese lead researcher, Shinya Yamanaka, believes that it will take at least one year to prove the safety of the new technique.

Although the two research teams’ results have been welcomed because they do not use embryos, they have triggered a whole new ethical debate. Indeed, some scientists are already pointing to the possibility of scientific misconduct in the use of this technique. They claim that, in theory, a single person’s DNA could be used to create both eggs and spermatozoa from iPS. In practice this could wreak havoc with the reproductive process.

No legislation on the subject yet exists. A spokesperson for the British Human Fertilisation and Embryology Authority (HFEA) told the Daily Telegraph newspaper that legally it is a “grey area”. In an interview published by leading French newspaper, Le Figaro, the chairman of France’s National Ethics Advisory Committee for the Life Sciences and Health (CCNE) played down the risk of scientific misconduct. Legislation to ban this practice is expected very soon.

While it is wise to foresee the risks of new practices, it would appear difficult to create gametes (sex cells) from iPS. Indeed, sex cells can be formed only in their niche (ovarian follicles in women and seminiferous tubules in men). Furthermore, as they mature, gametes undergo meiotic reduction. This process specific to sex cells results in cells with half the number of chromosomes. So it is pure fantasy to imagine that gametes could be produced from iPS without going through meiosis!

Audrey Binet

  1. Takahashi, K., Tanabe, K., Ohnuki, M., Narita , M., Ichisaka, T., Tomoda, K., Yamanaka, S. (2007). Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell, 131, 1-12
  2. Yu, J., Vodyanik, M., Smuga-Otto, K., Antosiewicz-Bourget, J., Frane, J., Tian, S., Jonsdottir, G., Ruotti, V., Nie, J., Thomson, J. (2007). Induced pluripotent stem cell lines derived from human somatic cells. Science.

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How long before we see clinical applications?

The recent discoveries of the American and Japanese research teams do not seem to challenge the scientific worth of embryonic stem cells and are far from ending the ethical debate. However, they do hold out new hope for treating ailments like cancer, diabetes, arthritis, spinal cord lesions, heart disease, burns, Parkinson’s disease and Alzheimer’s. With regard to organ transplants, the new technique could enable doctors to create stem cells using the patient’s own DNA, thus eliminating the risk of rejection.

It is easy to understand why this innovative method is being greeted with so much fervour. However, there is still a long way to go before the method can be tested for clinical applications. First of all scientists will need to determine how identical iPS are to embryonic stem cells.

What is more, the mechanisms involved in cell reprogramming are still very poorly understood. Another outstanding issue that needs to be resolved before any medical applications can begin is to find an alternative to retroviruses for introducing “rejuvenating genes” into a cell. The use of such viruses could cause harmful genetic mutations. A key stage in developing the technique is therefore to remove the need for a retrovirus.

The flaw in the Japanese research method is that it employs a gene with such powerful carcinogenicity. However, Shinya Yamanaka’s team have apparently already modified its protocol and is said to have obtained pluripotent stem cells from mouse and human adult skin cells using only three genes. During this experiment, none of the 26 mice created using stem cells devoid of the c-Myc gene died from a tumour. By contrast, six of the 37 mice created from stem cells containing the cancer gene died from one.

The experts are resuming their quest for the Grail with renewed vigour. Two weeks following media coverage of the American and Japanese research results, more researchers announced that they had succeeded in applying the technique. Jacob Hannah and his team from the WhiteheadInstituteforBiomedicalResearchin Cambridge, Massachusetts (USA) successfullytreated anaemic mice using iPS derived from skin cells in their tail (Science, 7 December 2007). This could represent the first small step towards human iPS applications …



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