Technique removes need for eggs or embryos.
Research reported this week by three different groups shows that normal skin cells can be reprogrammed to an embryonic state in mice1,2,3. The race is now on to apply the surprisingly straightforward procedure to human cells.
If researchers succeed, it will make it relatively easy to produce cells that seem indistinguishable from embryonic stem cells, and that are genetically matched to individual patients. There are limits to how useful and safe these would be for therapeutic use in the near term, but they should quickly prove a boon in the lab.
“It would change the way we see things quite dramatically,” says Alan Trounson of Monash University in Victoria, Australia. Trounson wasn't involved in the new work but says he plans to start using the technique “tomorrow”. “I can think of a dozen experiments right now — and they're all good ones,” he says.
In theory, embryonic stem cells can propagate themselves indefinitely and are able to become any type of cell in the body. But so far, the only way to obtain embryonic stem cells involves destroying an embryo, and to get a genetic match for a patient would mean, in effect, cloning that person — all of which raise difficult ethical questions.
As well as having potential ethical difficulties, the 'cloning' procedure is technically difficult. It involves obtaining unfertilized eggs, replacing their genetic material with that from an adult cell and then forcing the cell to divide to create an early-stage embryo, from which the stem cells can be harvested. Those barriers may have now been broken down.
“Neither eggs nor embryos are necessary. I've never worked with either,” says Shinya Yamanaka of Kyoto University, who has pioneered the new technique.
Last year, Yamanaka introduced a system that uses mouse fibroblasts, a common cell type that can easily be harvested from skin, instead of eggs4. Four genes, which code for four specific proteins known as transcription factors, are transferred into the cells using retroviruses. The proteins trigger the expression of other genes that lead the cells to become pluripotent, meaning that they could potentially become any of the body's cells. Yamanaka calls them induced pluripotent stem cells (iPS cells). “It's easy. There's no trick, no magic,” says Yamanaka.
The results were met with amazement, along with a good dose of scepticism. Four factors seemed too simple. And although the cells had some characteristics of embryonic cells — they formed colonies, could propagate continuously and could form cancerous growths called teratomas — they lacked others. Introduction of iPS cells into a developing embryo, for example, did not produce a 'chimaera' — a mouse carrying a mix of DNA from both the original embryo and the iPS cells throughout its body. “I was not comfortable with the term 'pluripotent' last year,” says Hans Schöler, a stem-cell specialist at the Max Planck Institute for Molecular Biomedicine in Münster who is not involved with any of the three articles.
This week, Yamanaka presents a second generation of iPS cells1, which pass all these tests. In addition, a group led by Rudolf Jaenisch2 at the Whitehead Institute for Biomedical Research in Cambridge, Massachusetts, and a collaborative effort3 between Konrad Hochedlinger of the Harvard Stem Cell Institute and Kathrin Plath of the University of California, Los Angeles, used the same four factors and got strikingly similar results.
“It's a relief as some people questioned our results, especially after the Hwang scandal,” says Yamanaka, referring to the irreproducible cloning work of Woo Suk Hwang, which turned out to be fraudulent. Schöler agrees: “Now we can be confident that this is something worth building on.”
The improvement over last year's results was simple. The four transcription factors used by Yamanaka reprogramme cells inconsistently and inefficiently, so that less than 0.1% of the million cells in a simple skin biopsy will be fully reprogrammed. The difficulty is isolating those in which reprogramming has been successful. Researchers do this by inserting a gene for antibiotic resistance that is activated only when proteins characteristic of stem cells are expressed. The cells can then be doused with antibiotics, killing off the failures.
The protein Yamanaka used as a marker for stem cells last year was not terribly good at identifying reprogrammed cells. This time, all three groups used two other protein markers — Nanog and Oct4 — to great effect. All three groups were able to produce chimaeric mice using iPS cells isolated in this way; and the mice passed iPS DNA on to their offspring.
Jaenisch also used a special embryo to produce fetuses whose cells were derived entirely from iPS cells. “Only the best embryonic stem cells can do this,” he says.
“It's unbelievable, just amazing. It's like Dolly. It's that type of accomplishment.”
“It's unbelievable, just amazing,” says Schöler, who heard Jaenisch present his results at a meeting on 31 May in Bavaria. “For me it's like Dolly [the first cloned mammal]. It's that type of accomplishment.”
The method is inviting. Whereas cloning with humans was limited by the number of available eggs and by a tricky technique that takes some six months to master, Yamanaka's method can use the most basic cells and can be accomplished with simple lab techniques.
But applying the method to human cells has yet to be successful. “We are working very hard — day and night,” says Yamanaka. It will probably require more transcription factors, he adds.
If it works, researchers could produce iPS cells from patients with conditions such as Parkinson's disease or diabetes and observe the molecular changes in the cells as they develop. This 'disease in a dish' would offer the chance to see how different environmental factors contribute to the condition, and to test the ability of drugs to check disease progression.
But the iPS cells aren't perfect, and could not be used safely to make genetically matched cells for transplant in, for example, spinal-cord injuries. Yamanaka found that one of the factors seems to contribute to cancer in 20% of his chimaeric mice. He thinks this can be fixed, but the retroviruses used may themselves also cause mutations and cancer. “This is really dangerous. We would never transplant these into a patient,” says Jaenisch. In his view, research into embryonic stem cells made by cloning remains “absolutely essential”.
If the past year is anything to judge by, change will come quickly. “I'm not sure if it will be us, or Jaenisch, or someone else, but I expect some big success with humans in the next year,” says Yamanaka.
Okita, K., Ichisaka, T. & Yamanaka, S. Nature doi:10.1038/nature05934 (2007).
Wernig, M. et al. Nature doi:10.1038/nature05944 (2007).
Maherali, N. et al. Cell Stem Cell doi:10.1016/j.stem.2007.05.014 (2007).
Takahashi, K. & Yamanaka, S. Cell 126, 663–676 (2006).
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