Translating Thomson’s “Induced Pluripotent Stem Cell Lines Derived from Human Somatic Cells”

Yu, Thomson, and all, from Wisconsin published their paper on reprogrammed induced Pluripotent Stem (iPS) cells from adult cells online in Science Express online, yesterday, just after the Yamanaka/Takahashi team from Japan published theirs in the journal, Cell. (The Thomson paper was not scheduled to go live online until the 22nd.)

As discussed on this blog, yesterday, Yamanaka’s group built on their earlier research published in 2006 and 2007, using mouse fibroblasts to prove that four genes, Oct4, Sox2, c-myc, and Klf4, could reprogram those cells to a state that resembled embryonic stem cells in all tests that they tried. Then, they used fibroblasts from commercially available samples from 1) a skin biopsy taken from the face of a 36 year-old Caucasian woman, 2) synovial cells (joint lining) from a 69 year old Caucasian male, and 3) neonate foreskin skin fibroblasts.

(This last is a common source of skin fibroblasts, with easily and ethically accessed skin tissue, collected at the circumcision of newborn boys.)

The Thomson team did not begin the basic research using mouse cells and did not simply go forward using the genes from the earlier experiments on mouse cells. Instead, they started at the beginning, using human Embryonic Stem Cells (hESC) that had been directed to become a special type of white blood cell, CD45+. This type of cell can be manipulated to demonstrate whether they had the functioning gene, Oct4 (a definite marker that is used to prove whether or not a cell is a hESC), by growing them in the presence of gentamycin, an antibacterial.

By adding some genes and removing others, the team determined that they had, “identified a core set of 4 genes, OCT4, SOX2, NANOG, and LIN28, that were capable of reprogramming human ES cell-derived somatic cells.” They also discovered that the cells could be reprogrammed into embryonic-like cells without Nanog, but that Nanog made it possible to recover more reprogrammed cells.

(From the text accompanying Fig.1: "In three independent experiments using different preparations of
mesenchymal cells, individual removal of either OCT4 or SOX2 from reprogramming combinations eliminated the appearance of reprogrammed clones, whereas the individual removal of either NANOG or LIN28 reduced the number of reprogrammed clones, but did not eliminate such clones entirely.")

Next, they tested this combination of genes in a commercially available, genetically modified cell culture, IMR90 fetal fibroblasts. (These cells were cultured from a little girl aborted at 16 weeks gestation. ) These cells are fetal cells, not adult cells, and they were chosen because they have been studied and the genome is well known. They do not grow well in the fluids and conditions that encourage cultures of hESCs and the researchers could identify them by the way that they look.

Next, in order to prove that the genes could reprogram “adult cells,” the team used fibroblast cultures from foreskins to produce 4 different cultures of reprogrammed induced Pluripotent Stem Cells.

The authors conclude,

"The human iPS cells described here meet the defining criteria we originally proposed for human ES cells, with the significant exception that the iPS cells are not derived from embryos. Similar to human ES cells, human iPS cells should prove useful for studying the development and function of human tissues, for discovering and testing new drugs, and for transplantation medicine."

Edited typos 11/21/07 17:30 PM (That could be the next neuroscience break through: why don't we see our typos until later?)

Science Magazine on "Race" to Stem Cell Breakthrough

The Thomson article is online (abstract is free, article is behind a pay wall), but I haven't had a chance to read it.

In the meantime, Science Magazine has a news article on both the publication from Wisconsin's Thomson and the previously discussed Takahashi/Yamanaka article in Cell.

Be sure and read the last sentence!!!!

Now the race to repeat the feat in human cells has ended in a tie: Two groups report today that they have reprogrammed human skin cells into so-called induced pluripotent cells (iPCs). In a paper published online in Cell, Yamanaka and his colleagues show that their mouse technique works with human cells as well. And in a paper published online in Science, James Thomson of the University of Wisconsin, Madison, and his colleagues report success in reprogramming human cells, again by inserting just four genes, two of which are different from those Yamanaka uses.

Thomson's team started from scratch, identifying its own list of 14 candidate reprogramming genes. Like Yamanaka's group, the team used a systematic process of elimination to identify four factors: OCT3 and SOX2, as Yamanaka used, and two different genes, NANOG and LIN28. The group reprogrammed cells from fetal skin and from the foreskin of a newborn boy. The researchers were able to transform about one in 10,000 cells, less than Yamanaka's technique achieved, Thomson says, but still enough to create several cell lines from a single experiment.

Although promising, both techniques share a downside. The retroviruses used to insert the genes could cause tumors in tissues grown from the cells. The crucial next step, everyone agrees, is to find a way to reprogram cells by switching on the genes rather than inserting new copies. The field is moving quickly toward that goal, says stem cell researcher Douglas Melton of Harvard University. "It is not hard to imagine a time when you could add small molecules that would tickle the same networks as these genes" and produce reprogrammed cells without genetic alterations, he says.

Once the kinks are worked out, "the whole field is going to completely change," says stem cell researcher Jose Cibelli of Michigan State University in East Lansing. "People working on ethics will have to find something new to worry about."

(edited November 21, 2007 to adjust the title. I over reacted in calling this statement and "insult.")