First Animal Cloning from a Fully Mature Cell

[Angel]

University of Pittsburgh researchers announced Sunday they have cloned the first animal from a fully mature cell, a feat they say forces scientists to rethink how Dolly the sheep and other mammals were cloned.

Yet independent experts questioned those conclusions, arguing that animals already have been cloned from mature cells -- and in a more convincing fashion.

Pitt stem cell biologist Dr. Tao Cheng and scientists at the University of Connecticut reported in yesterday's online issue of the journal Nature Genetics that they have cloned two live mouse pups from a mature white blood cell. Mature cells -- called fully differentiated cells -- are incapable of dividing or turning into other types of cells, as stem cells can.

Cloning mammals is highly inefficient, usually taking thousands of eggs to generate several live young. Knowing what type of cell is best for cloning would make the process more efficient if it is ever used to treat disease.

"Now we can say with near certainty that a fully differentiated cell retains the genetic capacity for becoming like a seed that can give rise to all cell types necessary for the development of an organism," said Cheng, director of stem cell biology and co-leader of the cancer stem cell program at the University of Pittsburgh Cancer Institute.

Cloning expert Konrad Hochedlinger disagreed.

"I'm not sure if this work really tells us much more about anything," said Hochedlinger, an assistant professor of medicine at Harvard Medical School.

Hochedlinger showed in 2002 that mice can be cloned from mature immune cells, and he said other researchers have cloned mice from fully mature nerve cells.

"All of these papers in high-profile journals made the strong point that terminally differentiated cells retain all the genetic information to give rise to a cloned animal," Hochedlinger said.

Cheng and his colleagues said these other experiments did not produce true clones because they did not directly use the technique used to create Dolly the sheep in 1996. Dolly was the first cloned mammal.

That method involves removing the genetic material from an unfertilized egg and replacing it with DNA taken from a donor cell.

The egg containing the transferred DNA is induced to divide until it forms an early-stage embryo. This embryo is then implanted into a surrogate mother. If carried to term, the cloned animal would be genetically identical to the DNA donor.

The Pitt and UConn researchers performed hundreds of cloning experiments using three types of donor cells isolated from bone marrow: fully differentiated white blood cells called granulocytes, less mature progenitor cells and adult stem cells.

Prevailing scientific thought holds that the adult stem cells should be most effective because they can multiply and turn into more specialized cell types.

But Cheng and co-corresponding author Xiangzhong Yang discovered the opposite was true.

Only the granulocytes gave rise to full-term mouse pups, although both died within two hours of birth.

About 35 percent to 39 percent of clones made with granulocytes developed into early-stage embryos, compared to 11 percent for the progenitor cells and 4 percent for the stem cells, their study reports.

"That was completely a surprise," Cheng said. "We were stumped by our results, so we repeated our experiments six times to make sure there was no chance of contamination of our clones from the stem cells."

Critics say the experiments left a 0.6 percent chance that stem cells created the clones.

"This has been a constant problem in the cloning field, that it is hard to unequivocally prove the cell type you are using," Hochedlinger said.

To be certain the mouse pups were created with granulocytes, the researchers would need to use genetic markers that trace the fate of the donor cells before and after cloning, said Dr. Rudolph Jaenisch, a leader in the cloning field.

"It's a nice thing to get mice from granulocytes, but a few issues are not at rest and are very difficult to address, so I'm not sure if I can accept all these conclusions," said Jaenisch, of the Whitehead Institute for Biomedical Research at Massachusetts Institute of Technology.

In the past decade, scientists have debated what type of cell spawned Dolly and other cloned mammals. Many argued that adult stem cells, because of their presumed versatility, provided the donated DNA.

Yang, who directs the UConn Center for Regenerative Biology, said fully differentiated cells such as granulocytes are more likely candidates because his team could not clone any pups from adult stem cells.

Others aren't so sure.

"We'll never know that," said Dr. Robert Blelloch, a cloning expert at the University of California, San Francisco. "We can't go back in time and figure that out."