It offers the possibility of a personalised supply of all kinds of tissue types, without cloning, donated eggs or the destruction of embryos. If the latest breakthrough in stem cell technology in mice is repeated in humans – and żěè¶ĚĘÓƵ has learned that experiments on human cells are now under way – it could demolish the ethical objections that have dogged the field.
Shinya Yamanaka and Kazutoshi Takahashi of Kyoto University in Japan have produced what are effectively embryonic stem cells (ESCs) from mouse skin cells by exposing them to four messenger chemicals that are found in embryonic but not adult cells. The researchers also derived ESCs from differentiated cells in mouse embryos, but the work with skin cells is most significant because it overcomes ethical objections relating to embryos.
A personalised, ethically watertight treatment would represent the long dreamed-of pinnacle of stem cell research. Replacement tissues generated this way would overcome at a stroke the desperate shortages of donated organs. Since the tissues would be genetically identical to the recipient, patients would also be spared a lifetime of taking immunosuppressive drugs to stop donated organs being rejected.
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To transform the skin cells, Yamanaka and Takahashi first traced 24 genes that are highly active in mouse ESCs but not adult cells. They then ferried combinations of these genes into the skin cells aboard viruses, and found that some of the skin cells appeared to be transformed into ESCs. The researchers worked out that four genes, called Oct3/4, Sox2, c-Myc and Klf4, were vital to this (Cell, vol 126, p 663).
These four genes make chemical factors that rewind the the skin cells’ developmental clock. The resulting ESC-like cells are called induced pluripotent cells (iPCs), and by implanting them into mice with no immune system, Yamanaka and Takahashi proved the cells could change into all the tissues of the body.
The big question is whether these four chemicals can perform the same trick in human adult cells. “If this technique were to work in humans, it would be ideal, since scaling-up of such a procedure to produce adequate numbers of cells for a real therapy could be relatively straightforward,” says Chris Mason of University College London in the UK.
“Human embryonic stem cells might be used to treat a host of diseases, such as Parkinson’s disease, spinal cord injury and diabetes,” says Yamanaka. “Our finding is an important step in controlling pluripotency, which may eventually allow the creation of pluripotent cells directly from [adult] cells of patients.”
“Human embryonic stem cells might be used to treat a host of diseases, such as Parkinson’s, spinal cord injury and diabetes”
There is another key question about using viruses to load genes into a patient’s cells: would the virus be safe when transplanted back into the patient inside those cells? Also, the product of the c-Myc gene is linked with progression of cancer, so Yamanaka himself warns there is a risk that the transplanted cells could go haywire and form cancers. Overall, however, stem cell researchers have reacted enthusiastically to the findings. “It opens up entirely new opportunities for the production of cells from patients either to study inherited disease or for therapy,” says Ian Wilmut of the University of Edinburgh, UK, the creator of Dolly the sheep.
“If it really works, it would be remarkable and immensely useful, both scientifically and therapeutically,” says John Gurdon, head of the Gurdon Institute in Cambridge, UK. “It’s surprising to me that it works, and I’m sure there will be lots of labs trying to repeat and refine it.”
One of the Gurdon Institute’s projects is to find out how chemical factors in eggs perform the same trick of rewinding adult nuclei back to an embryonic state. This was exploited to make Dolly the sheep, who was created by fusing the nucleus of an adult sheep cell with an egg cell emptied of its own chromosomes (see Graphic).
Using the same approach to make human ESCs is controversial. The idea is to make a “temporary” embryo by fusing the nucleus of a patient’s cell with a human egg emptied of its own chromosomes. After growing for a few days, it should be possible to harvest and grow ESC lines – although no one has yet achieved this. Objectors say that as with using “spare” embryos from IVF, the procedure is unethical because it destroys a human embryo.
Gurdon says that two of the factors used in the Cell paper – made by the genes Oct3/4 and Sox2 – are already well known as “markers” unique to the surface of pluripotent stem cells, so it makes sense that production of these chemicals reverts adult cells to a pluripotent state. “They are characteristic of pluripotency and induce it themselves,” he says.
Gurdon also points out that although only tiny numbers of the experiment’s skin cells – less than 0.1 per cent – became pluripotent, he says that it doesn’t matter as long as it works in a few. And, of course, it requires no eggs or embryos – which some feel may be too good to be true. “It seems a bit too easy,” says stem cell expert Stephen Minger of King’s College London.
Yamanaka says that now they have demonstrated the principle of using messenger chemicals to make ESCs, it might be possible to refine the technique by tweaking the chemical recipe. Only some combinations of the 24 genes were tested, so others might turn out to be more efficient. One surprise was that one of the messengers called Nanog, well known for its ability to stop ESCs maturing, wasn’t in the final recipe. But Nanog levels increased as a knock-on effect of the other factors, so it might not be required as a primary ingredient.
Another puzzle is why mouse embryos injected with the iPCs and implanted in the wombs of mice all died before birth. Dissections showed that many of the dead fetuses were chimeras, composed of a combination of their own tissues and those created from the pluripotent cells.
“Although none developed to term, there are many factors that could account for this,” says Ian Wilmut. “They include the continued expression of the viral vectors that produced the specific factors to produce pluripotent cells.” The result would have been that not all the cells matured correctly into different tissue types, but kept growing as stem cells.
More encouragingly for hopes of creating human tissue, the engineered cells successfully formed a multitude of tissues and organs in the fetuses, including nerves, muscles, cartilage, heart, liver, spine and gonads.
“The engineered cells successfully formed a multitude of tissues and organs, including nerves, muscles, heart, liver and spine”
“The big thing is whether you can do this with human cells,” says Minger. “I would guess not,” he says. “I think many people thought it would be much more complicated than this, and whether the cells are stable, safe, and have full differentiation potential of embryonic stem cells remains to be seen.”
But despite recent high-profile cases where promising ESC research failed to overcome all the ethical objections (żěè¶ĚĘÓƵ, 2 September 2006, p 4), not everyone is pessimistic. “I’d assume it could work in humans,” says Gurdon, but he cautions that it could take time. Although IVF initially worked in animals, it was years before the first test-tube baby was born, he warns.
“We are now trying with human cells,” says Yamanaka, “but we don’t know the answer yet. We do not yet know when we will know the answer, either.”
