Scripps Research Institute scientists are reporting a breakthrough in stem cell research in which they successfully created live mice from mouse skin cells, without using embryonic stem cells or cloning techniques that require eggs. This milestone opens the door to the development of exciting therapies, such as using a patient's own cells to grow replacement organs.
The research is reported in the August 2, 2009, advance, online issue of the journal Nature in a paper titled "Adult mice generated from induced pluripotent stem cells."
In this report, a group of Scripps Research scientists, led by Assistant Professor Kristin Baldwin, Ph.D., describes the creation of mice from what are known as induced pluripotent (iPS) cells-stem cells created by reprogramming normal cells taken non-destructively from living animals. While for several years no research team had been able to generate live adult animals from iPS cell lines, the study is part of a trio of new papers showing the feat is possible. Also reporting similar results are two Chinese groups, who published their findings online in Nature and Cell Stem Cell just days ago. Each group used different methods, with the Scripps Research team's protocols offering more successful results by some measures.
"Reprogramming by iPS cell technology is one of the most exciting areas of research right now," says Baldwin, "because these experiments challenge fundamental paradigms in basic biology and, at the same time, contribute to a technology that offers enormous potential for therapeutic advances."
From Skin Cell to Mouse
Since iPS cells were first generated, several years ago, multiple groups have tried to produce mice from them but no live mice were ever born. For unknown reasons, iPS mouse embryos stopped developing about two-thirds of the way through gestation. This led to concerns that iPS cells might be inferior to embryonic stem cells and hinted that reprogramming with four factors might not be the best method to produce pluripotent cell lines from patients. But the Baldwin team was up to the challenge.
The first part of the team's new study involved gathering cells that were already "differentiated," i.e. developed into a particular cell type, such as skin, nerve, or muscle. In this case, the scientists worked with skin cells from fetal mice, though other cell types may also work. Viruses were then used to insert genes coding for four proteins, called reprogramming factors, into these cells' DNA. These reprogramming factors shifted the cells out of their normal differentiated state to a "pluripotent" state resembling that of embryonic stem cells, which allows the cells to produce a wide variety of cell types. This cellular rewiring caused the cells to change their size and shape so that after only 7 to 10 days they could not be visually distinguished from embryonic stem cells.
"It's actually quite remarkable to see," says Baldwin.
Once skin cells were converted into cells that looked like embryonic stem cells, the researchers needed to find out whether the cells acted like embryonic stem cells. If they didn't, the researchers reasoned, the cells might not be useful therapeutically in generating important cell types such as neurons, heart cells, or liver cells.
The ultimate test of this developmental puripotency was to generate live mice entirely from iPS cells. This week, the Baldwin team and two Chinese groups all reported that they had independently grown live mice from iPS cells all the way to fertile adulthood.
Many questions remain about what determines whether live mice will result from a given line of iPS cells. Of the reported successes, the Baldwin group's techniques appear most effective, with the best line producing live pups about 13 percent of the time compared to 3.5 percent or 1 percent reported in the Zhou and Gao groups' studies. The Scripps Research team also was able to generate live mice from four of 15 lines generated in one experiment (four of four tested) while the Zhou and Gao groups reported success with three of 37 (three of six tested) and one of five, respectively (one of two tested). "We can't say for sure yet, but it is possible that we may have identified a protocol more likely to produce mice that survive until birth than current methods in the field," says Baldwin.
Now that researchers have a number of cell lines that do and do not generate live mice, comparisons among them should make it possible to zero in on exactly which parameters mark the production of successful lines. This in turn should provide invaluable information to aid in advancing iPS studies. Baldwin is also interested in the possibility of comparing mice raised normally with those generated using cloning, embryonic stem cells, or iPS cells in order to better understand how tissues derived from iPS cells might behave in human cell transplant experiments.
Clearly one advantage of iPS cell lines, assuming no issues with their stability emerge, is that they can be created without the need to harm the donor, meaning they offer the potential to avoid most of the ethical issues associated with embryonic stem cells. But iPS cells offer other advantages as well, which are tied to the key hopes for future work.
Researchers, and perhaps eventually physicians, can take cells from mice or humans at any age to generate an iPS cell line. This opens the enticing possibility that iPS cells might be manipulated to grow replacement organs such as hearts and livers, or to provide healthy replacements for damaged cells, such as neurons needed to cure paralysis, Parkinson's, or Alzheimer's disease, all possibilities research groups around the world are vigorously pursuing. Because such cells would be derived directly from the patient, the rejection problems that plague conventional transplant therapies would be eliminated. Another hope is that iPS cells will be used to create new disease models that will foster better understanding of disease causes and more rapid identification of potential treatments.
Baldwin credits a combination of hard work and teamwork for her group's success. The reprogramming experiments were carried out by a talented trio of co-first authors, postdoctoral researcher Michael Boland, Ph.D., and Scripps Research Kellogg School graduate students Kristopher Nazor and Jennifer Hazen, with assistance from Wesley Gifford, from the University of California, San Diego. The technically challenging tetraploid complementation experiments and blastocyst injections were performed in close collaboration with Sergey Kuprianov, Ph.D., and his team from the Scripps Research Mouse Genetics core, including Alberto Rodriguez and Greg Martin.
This work was supported by a Pew Scholars award and through funding from the California Institute of Regenerative Medicine, the Whitehall Foundation, the O'Keefe Foundation, and the Shapiro Family Foundation.