Dolly the cloned ewe has been at the center of controversy since she was announced to the world in 1997. Beneath the philosophical considerations, the science of the cloning feat--in particular the type of cell used to kick off the process--has been an issue of some debate. Some have argued that so-called adult stem cells--root cells in most tissue that kick into action to replace damaged tissue--must have been involved. But a new test in mice shows that adult stem cells are actually worse than regular cells for the purposes of cloning with current techniques. Moreover, the experiment delivered two cloned pups from the genetic material contained in fully formed white blood cells.
Cloning relies on a process known as somatic cell nuclear transfer, in which the nucleus of a donor cell is transferred into a fertilized egg that has been emptied of its chromosomes. That egg now contains an exact duplicate of the donor's genome, and if all goes well when it is implanted into a surrogate mother, a clone will result. Xiangzhong Yang of the University of Connecticut and Tao Cheng of the University of Pittsburgh as well as a host of colleagues examined the cloning potential of three different types of cells: hematopoietic stem cells, progenitor cells and granulocytes. Each represents a different stage in the differentiation process of blood cells; stem cells can become any kind of blood cell, progenitor cells are already on a particular track and granulocytes are a specific type of white blood cell (further specified as neutrophils).
The researchers expected that nuclei transferred from stem cells would be the best in creating clones. "We thought that adult stem cells would give a much higher efficiency while terminally differentiated cells would give the lowest or zero," Yang explains. "To our surprise, when we compared cells from the same animal, instead of decreasing from stem cells with differentiated cells, we see a tremendous increase." In fact, only 4 percent of stem cell transfers produced nascent embryos compared with 8 percent of progenitor cells and 35 percent of granulocytes. Out of 1,368 granulocyte transfers at least 34 percent reached the blastocyst stage, and two actually resulted in living pups (who did not survive for long), according to the paper presenting the result published online in Nature Genetics on October 1.
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This is an important confirmation on several levels: most critically, adult stem cells are notoriously difficult to collect and store, whereas differentiated cells are plentiful and stable. "You can get [differentiated cells] anywhere in your body," Yang notes. "Most adult stem cells cannot be cultured in vitro without differentiation." Plus, hopes for therapeutic cloning rest on the ability to produce embryonic stem cells from cells harvested from diseased patients.
Stem cells harvested from embryos rather than adults remain the most powerful for cloning and other purposes; Yang's team showed that cloning from such cells succeeded in 49 percent of attempts and led to 18 mouse pups. Of course, such embryonic stem cells are not available in adult patients, so being able to create them from regular cells is an important step. But mystery still surrounds the best way to clone. "We think that adult stem cells are more quiescent; we think that it's like a locked, closed door," Yang explains. "We think that differentiation is a way that opens some doors and makes it easier for nuclear transfer programming to go back into embryonic stem cells."
"We have some very encouraging data from an ongoing study that you can modify the status of cells and increase the efficiency of cloning," he adds. "If we modify the epigenetic status," that is, turning certain genes on or off, "it could be that you could increase the efficiency. This opens the door for a lot of different studies." The odds are that Dolly's genome came from a differentiated cell, but the quest for the best way to create cloned animals continues.
