Yamanaka has a conscience

“When I saw the embryo, I suddenly realized there was such a small difference between it and my daughters,” said Dr. Yamanaka.

The New York Times article on Shinya Yamanaka, "Risk taking is in his genes," (free one time registration necessary) should get the headline-writer in trouble for a sad pun.

Instead, Dr. Yamanaka might be in trouble with the objectors to conscience. (No links, just look at today's posts - or the last two months of posts - the subject keeps popping up.)

People like John Gearhart, MD will want to "put pressure" on Yamanaka to write letters to Nancy Pelossi and the rest of the US legislators making the usual reactionary case for Federal funding for embryonic stem cell research in light of the successes with non-destructive research.

The NYT reporter, Martin Fackler, can't be too popular in the next few days for pointing out that the US laws and funding are not nearly as tight as those in Japan, due to moral objections in that country:

In 1999, his career got a break when he was hired by other universities, including Kyoto University in 2004, that were willing to give him a laboratory and more money. At about the same time, he said, he visited his friend’s fertility clinic. That visit inspired him to find a way around the moral issues that had bogged down stem cell research, not just in the United States but also Japan, where the Education Ministry put tough restrictions on embryo use.

In fact, restrictions are so tight that he says he cannot use human embryos at his laboratories here. Instead, research using human embryos is done at U.C. San Francisco, where he maintains a small two-person laboratory. He said he had never handled actual embryonic cells himself, and the American lab uses them only to verify that the reprogrammed adult cells are behaving as true stem cells.

“There is no way now to get around some use of embryos,” he said. “But my goal is to avoid using them.”

For a look at the science and bioethics slant on these revelations, see Wired Science (see the comments on this one), Blog.bioethics.net, Wesley Smith's Secondhand Smoke, and Jennifer Lahl's blog, "The Human Future."

Wash this reactionary's mouth out with soap!

Bioethics.net compares the Bush administration's happiness about reprogrammed adult stem cells with that man, Mr. Clinton's, "I did not have sex with that woman!" and President Bush's statement "Mission accomplished," after our US troops captured Baghdad.

I'll accept the latter (at some future date, if the evidence supports it), but the first is at least as false as Clinton's wagging finger - and (speaking of Yuk factors) did we really need to be reminded of that?

The author, James W. Fossett (who is anything but "non-partisan") states that Yamanaka, the first to report reprogrammed adult cells in humans and mice is from Japan and wasn't affected by the US Federal funding limitations. He doesn't mention that Yamanaka's research didn't rely on the use of new embryonic cells, at all. Yamanaka took the information gleaned from animal research and the currently funded cells and moved to the front of all other stem cell researchers by pointing the way to the key to the production of stem cells from each patient who needs them - from his or her own cells.

Instead, Fossett is running scared due to the "rhetorical parity" from cell reprogramming and the possibility that the success in reprogramming cells will result in more reprogramming research!

Fossett doesn't mention that James Thomson's research using human Embryonic Stem cells (hESC). then human fetal cells harvested after abortions - and finally in skin cells harvested at circumcision of little boys - was funded by the National Institutes of Health, and that those hESC are the ones that supposedly are of no use.

Fossett also fails to mention of the new report by Yamanaka on the technique using only 3 inserted genes to the prior 4, and that the eliminated gene is the one that had scientists concerned about cancers.

I'm sure that he doesn't recall the "first transplantable lung cells" from hESC's by Texas researchers last year. These cells were developed by viral "transfection," also, and were lauded as "a platform that could potentially be useful in the development of spinal cord cells, heart cells, nerve cells and others.” These were neither the first or transplantable, but they did get much more notice than similar cells developed from umbilical cord blood cells without viral transfection.

That may be the problem: the proponents of hESC research are used to getting many times the publicity from hESC research than that received by the non-hESC researchers. And so, we get the concerns about "rhetoric."

There's those deceitful knee jerk reactionaries practicing their projection, again.

Reactionary Scientists

I wonder how often our friend from Kyoto is planning to publish and what tweaks we'll hear about next week?

I also wonder how many of the comments about "must fund all" come from - or actually are a type of - the application of the sort of pressure that Gearhart told his audience in DC that he and others applied to induce Atala to write Pelosi?

I'm trying to get used to the idea of scientists as reactionaries.

Politics aside, I don't think any of the research could have gone any faster even without the "pressure" from either side. The basic science had to be done, and was.

Wait 'Till Next Week! (More iPS good news)

Dr. Yamanaka of Japan, the MD who made history last week by announcing that he had been able to obtain embryo-like stem cells fom adult skin cells called fibrobalsts. On Friday, November 30, has published a new report in Nature Biotechnology telling us how he was able to skip inserting the potential cancer causing gene, c-Myc.

At this rate, who knows what we'll have next week?

From The Scientist (online here):

In the current study, the researchers showed that pluripotent cells can be made from both mouse and human adult cells without introducing the c-Myc gene, by transducing just the other three. It's not that Myc isn't needed in the process, the authors noted in the paper; rather, they suggest that the other three genes may be spurring endogenous Myc activity. None of the 26 chimeras made from cells generated without c-Myc developed tumors within 100 days, compared to six out of 36 chimeras made from cells using all four genes.
*************
So far, too, efficiency with this triple-gene method is much lower than with the original four genes; half of the experiments without c-Myc did not produce pluripotent cells at all, while experiments using the four genes almost always yielded pluripotent colonies. "Does this mean that it now only works with a rare cell type?" Lovell-Badge wrote. "As always, many more questions are posed than answered."

The question is whether all fibroblasts are alike, or whether there is a smaller group of fibroblasts that are easier to induce to become "induced Pluripotent Stem" Cells.

If there are specialized cells in the skin that are easier to manipulate than others, this is good news for researchers and the patients who are looking toward stem cell research for treatments and cures.

Translation of Yamanaka, Yu "induced Pluripotent Stem Cells" (Revised)

Scientists who report their findings are expected to discuss the problems as well as the outcome of their research. This is usually found in the "Discussion," "Conclusions" or "Results" section of the paper. This is the best place to figure out what the researches intended, what they did and what the report means. (Then you go back and check to see if they proved what they "discussed." And then, you wait for other labs to confirm it.)

The actual (Takahashi et al., "Induction of Pluripotent Stem Cells from Adult Human Fibroblasts by Defined Factors," Cell (2007).) Cell article on reprogrammed adult fibroblast skin cells, the "induced Pluripotent Stem Cells) or "iPS," is available for free, here. The Science Magazine report about similar work by James Thomson from Wisconsin (the researcher who reported the production of human embryonic stem cells in the first place) is supposed to be published November 22, 2007. (Editorial note 11/30/07 – Science published the Thompson and Yu report the same day that Tamanaka's report was published, two days ahead of schedule. See my “translation,” here.)

To the best of my understanding, here's a translation into layman's terms about what the Takahashi/Yamaka report means:

While it took a lot of cells and more time than the researchers first expected because the human iPS grew much slower than the mouse iPS,

1. The cells that grew looked and functioned like human embryonic stem cells with a few minor differences,
2. They believe they proved that their technique is responsible for all the new pluripotent cells that were found in their cultures(there weren't any cells from another culture introduced accidentally or on purpose and which would make them look more successful than they were),
3. The cells could be directed to develop nerve cells and heart cells,
4. They were able to use several types of adult specialized cells to achieve iPS, and
5. The researchers suggest several possible ways to overcome the drawbacks of the process.

The authors believe that the inefficiency or the need to begin with lots of adult cells and wait a little longer for a substantial amount of human iPS should not be a "practical" problem because the adult cells are easy to obtain and labs all over the world should be able to reproduce their results. Since the technique should be well-funded (it qualifies for US Federal funding and is ethical, since no human beings have to die), the authors believe it will be possible for lots of researchers to work on them.

If I were to predict the future, I would anticipate banks of iPS - or even specialized or intermediate forms of cells that are produced from iPS - being stored for each of us, just in case. In the very long term, we will learn more about stimulating our on bodies' stem cells from research on these cells, so that we can repair or prevent damage without transplants or waiting for cultures to grow in the lab.

The major hurdle is that the cells were produced by the Recombinant DNA technique, using retroviruses in plasmids.

The retroviruses are a class of viruses that actually insert themselves into the DNA strands of animal or plant cells to become a part of that cell’s DNA and are copied when the cell reproduces. They are manufactured in the lab in the form of plasmids in order to carry genes into the experimental cells.

Plasmids are little bits of DNA, a mini-virus in a circle. Think of a chain with pairs of magnets or interlocking puzzle pieces that connect the ends and make a loop. When open, the plasmid becomes a strand of DNA which has ends that are "sticky.” When placed in a culture with mouse or human cells, the plasmids infect the cells and then move into the nuclei of the cells. The retroviral DNA is inserted or inserts itself into the DNA of the host cell because the sticky ends of the plasmid strand match or mate to certain areas of the host DNA.

Plasmids can be manufactured to carry copies of genes that researchers want to insert into the DNA of experimental cells. The technique is common in commercial and experimental labs for at least the last 30 years. In fact, "Recombinant DNA" is used to induce strains of bacteria and yeast cells in cultures to manufacture vaccines like the flu and Hepatitis B vaccine and the insulin used by diabetics these days. The particular retroviruses used by Tamanaka are said to be "strongly silenced in humans." In other words, they don't normally get reproduced as viruses when the cell divides. Once they are taken up in the cell DNA, the viruses used in research don't break out to become infectious viruses, again. However, some of them can induce the cells to form tumors or cancers if injected in an animal or human.


One of the possible problems that the article notes is that the new iPS cells each had several copies of the retrovirus included in their DNA. There is a concern that these bits may be responsible for the tumors that were seen in the mice used in the experiments. Before iPS can be used in humans, it will be necessary to learn to remove all the viral particles or to learn to make the cells without viruses that can cause tumors. Otherwise, there is a risk of causing cancer in patients.

The researchers note that another group of scientists have already reported that it is possible to insert one of the genes without using retroviruses and that the hope is to either find a way to insert the other three genes or to remove all traces of the virus.

There's also a suggestion that what they are actually inducing to grow is a sub-set of fibroblasts with the tendency to become embryonic-like stem cells.

Biographic article on Yamanaka

Here's a cute biography of Shinya Yamanaka, lead researcher from the Japanese team that reported reprogramming of adult cells into embryonic-like stem cells.

As an M.D. myself, I find it interesting that, unlike veterinarians James Thomson of Wisconsin and Time Magazine Person of the Year, Hu Suk Hwang, Dr. Yamanaka is a human doctor, trained in orthopedics:

Yamanaka has spent most of his life in western Japan. A native of Osaka, he earned his medical degree at Kobe University and a doctorate in pharmacology at Osaka City University.

After completing his residency in orthopedic surgery, Yamanaka headed to the University of California, San Francisco, to do postdoctoral studies that laid the groundwork for his current research.

He does express concern about the possible uses of his research by unethical researchers:

Yamanaka worries about the road some people might take.

"We need to come up with some sort of rules about what kind of cells can be used and to what ends. Otherwise, someone may put this technology to use in troubling ways,” Yamanaka said.

The research's ethical and social implications are never far from the table in Yamanaka's laboratory, said Kazutoshi Takahashi, a junior professor who participated in the project.

"The potential problems are cut down when you use this method given that we don't have to use embryonic stem cells, and that's a good thing,” Takahashi said.

Since the debate isn't yet over about ethical vs. unethical stem cells and since some people (like embryonic and fetal stem cell researcher and sometimes guest Science editor, John Gearhart, MD) have admitted to putting pressure on researchers to make sure that they follow the official line to pursue "all promising areas" (echoed here by the stem cell industry trade association organization, BIO) kinds of stem cell research, I hesitated to post this link and the quotes. But someone should record the true "debate."

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.")

Translation of "Induced Pluripotent (Human) Stem Cells"

Please see the revised version of this post, published November 30, 2007.