Until now, there has been an insurmountable barrier to cloning primate cells. Now the barrier is down, thanks to a new technique developed using rhesus monkey cells.
Credit: WIkipedia
CAIRNS: When Shoukhrat Mitalipov ended his unscheduled talk at the end of Monday’s session of the 5th International Society for Stem Cell Research Meeting held in Cairns, Australia, this week, there were audible gasps and exclamations of "wow!" from the audience.
Mitalipov talked of his latest potentially groundbreaking discovery: an efficient and reliable method for cloning primate embryos from adult cells - a technique known as therapeutic cloning. This breaks through what appeared to be an insurmountable barrier to producing embryonic stem cells from cloned primate embryos.
The technique brings scientists closer than ever before to producing human embryonic stem cells from cloned adult body cells. Producing such stem cells from a person's own body tissue would make it possible to sidestep the problem of rejection inherent in using stem cells from other sources.
The development also brings experts the closest they've been to cloning a full-term adult primate, such as a human.
Audible gasps
Stem cell biologist Alan Trounson, who chaired the session, had promised the audience they would not be disappointed by hanging around for Mitalipov’s 11th hour presentation.
They weren’t.
In a world first Mitalipov of the Oregon National Primate Research Centre in Beaverton, USA, provided evidence that he had successfully achieved somatic cell nuclear transfer in a primate. This means he managed to clone a rhesus monkey embryo from adult cells and generate embryonic stem cells from them. Previously it has proved impossible to derive embryonic cells from cloned embryos in primates. Mitalipov made two batches of embryonic stem cells from 20 cloned embryos.
“We’ve been looking for this evidence for a long time,” said Trounson who heads the Monash Immunology and Stem Cell Laboratory at Monash University in Melbourne, Australia.
Cloning from adult cells has been achieved in many species, such as mice, sheep, pigs, cows and dogs, but primates seemed to present an insurmountable barrier. Disgraced Korean 'stem cell king' Woo Sook Hwang failed to achieve the technique even through he had access to over 2,000 human eggs.
But now the primate barrier is down, thanks to a modified technique that Mitalipov used.
How he did it
Using skin cells from a ten year old male rhesus monkey, Mitalipov generated cloned embryos and ultimately two lines of embryonic stem cells, which he has given the acronym CRES, for cloned rhesus embryonic stem cells.
He revealed evidence that the chromosomes of these stem cell lines were indeed derived from the donor monkey. This is because the donor monkey’s chromosomes carried a very distinctive strip of DNA called a 42 XY translocation.
The cell lines also seemed to pass preliminary tests for bone fide embryonic stem cells. They stained positive for the canonical simian embryonic stem markers: Oct-4 , SSEA4, TRA1-60 and TRa-1-80 – which are proteins only produced by embryonic stem cells.
And as Mitalipov dramatically demonstrated in his presentation with a slide of beating cells, the CRES cells passed another test of embryonic stem cells; they where able to transform into either throbbing heart cells or neurons. They also formed teratomas – cancers that produce different types of tissues – when injected into mouse testis, just as other embryonic stem cells do.
However, when Irving Weissman, a developmental biologist of Stanford University in California, asked whether the cells had yet been tested for their ability to contribute to a monkey chimaera – an embryo made by mixing stem cells with cells of another embryo – Natalipov, replied in the negative. The chimaera is used as another definitive verification of embryonic stem cells.
"Beautiful work"
Mitalipov believes that standard cloning techniques have been the reason why experts have had such little luck in cloning adult primate cells before now.
Removing chromosomes from the egg (to prepare it to accept chromosomes from the adult cell being cloned) relies on visualising them using a dye and ultraviolet light – a procedure that damages essential factors that allow the resultant cell to reconfigure itself as an embryo, he said.
Furthermore, said Mitalipov, the use of electricity to fuse the donor skin cell to the chromosome-free egg, may cause the egg to activate or reconfigure itself prematurely.
Mitalipov's alternative technique visualises an egg's chromosomes without the dye or UV light, but through using polarised light to detect fibres that carry the chromosomes instead. He details the technique in the June 11 edition of the journal Human Reproduction.
Details of Mitalipov's cloning success have not yet been published in a journal, but it's likely to be in the press, given his presentation in a public arena. Until then, and until it is repeated by other experts, caution is warranted, comments Monash University's Trounson.
“Nevertheless, this is a beautiful work, really well done,” says Megan Munsie, of the Australian Stem Cell Centre in Melbourne and the first person ever to make embryonic stem cells from a cloned mouse embryo.
