Induced Pluripotent Stem Cell Transplant Claims Debunked

When this report first appeared, it seemed too good to be true and it turns out it probably was. Nobel Laureate Shinya Yamanaka at Kyoto University announced his remarkable discovery of induced pluripotent stem (iPS) cells in 2006. However, another Japanese researcher, Hisashi Moriguchi, made an even more earth-shaking claim earlier this year. Moriguchi, who was a visiting researcher at the University of Tokyo, claimed to have modified iPS technology to treat a person with terminal heart failure. The patient was allegedly surgically treated in February, 2012, according to a front-page article in the Japanese newspaper Yomiuri Shimbun. The article also said that the patient was healthy. If this was true, this would certainly be an earth-shaking result. An unidentified head of a Tokyo-based organization devoted to helping children with heart problems, told Yomiuri Shimbun, “I hope this therapy is realized in Japan as soon as possible.”

The Nippon News Network had posted a video of Moriguchi presenting his research at the New York Stem Cell Foundation, but they have since removed this video.

Unfortunately, once the journal Nature was altered to this report, they contacted Harvard Medical School and Massachusetts General Hospital (MGH), where Moriguchi claimed to have performed this work. Both institutions denied that Moriguchi had even done such a procedure. According David Cameron, a spokesperson for Harvard Medical School, “No clinical trials related to Dr Moriguchi’s work have been approved by institutional review boards at either Harvard University or MGH.” Likewise, the public affairs officer for MGH, Ryan Donovan, said “The work he is reporting was not done at MGH.”

There are other problems with Moriguchi’s work. Moriguchi reported that he had invented a method to reprogram cells using just two chemicals: a small molecule that inhibits a small RNA called “microRNA-145” and another molecule that binds the TGF-β receptor. However, a University of Tokyo stem-cell researcher, Hiromitsu Nakauchi, said that he has never “heard of success with that method.” Nakauchi even said that before this week he had never heard of Moriguchi.

Another bizarre claim made by Moriguchi was that he could differentiate iPS cells into heart muscle cells by utilizing a ‘supercooling’ method that he had invented. Nakauchi said that this was “another weird thing.”

Moriguchi never published his technique in a peer-reviewed journal, but in a book about advances in stem-cell research (see Moriguchi, H., Mihara, M., Sato, C. & Chung, R. T. in Embryonic Stem Cells — Recent Advances in Pluripotent Stem Cell-Based Regenerative Medicine (ed. Atwood, C.) 359–370 (InTech, 2011)). In this book, there are paragraphs copied almost verbatim from other papers. For example, a section under the heading “2.3 Western blotting” is identical to a passage from a 2007 paper by Yamanaka (see Takahashi, K. et al. Cell 131, 861–872 (2007)). Furthermore, section 2.1.1 describes human liver biopsies but the information in this section matches the number of patients and timing of specimen extractions described in an earlier article, but the name of the institution has been changed (see Thenappan, A. et al. Hepatology 51, 1373–1382 (2010)).

Nature contacted Moriguchi and he stood by his publication. He told Nature, “We are all doing similar things so it makes sense that we’d use similar words.” However, he did admit to using other papers “as reference.”

With respect to his reported supercooling technique, Moriguchi cited a paper of his own in Scientific Reports, which is published by the Nature Publishing Group. Nature, however, noted that this paper describes supercooling of human ovaries for preservation (Moriguchi, H., Zhang, Y., Mihara, M. & Sato, C. Sci. Rep. 2, 537 (2012)). The paper has nothing to do with the differentiation of iPS cells into cardiac cells. Moriguchi said that a journal referee had recommended that he leave the latter experiment out of the paper “because it’s basically the same technology”.

Moriguchi said that he did most of the contentious work himself, including safety research in pigs. However, the initial surgery and some of a further five similar procedures in other patients that took place from August onwards, and while, according to Moriguchi, other researchers were supposedly involved in some of these procedures, he would not provide any names.

Where did Moriguchi acquire the surgical expertise to perform these procedures? Moriguchi initially told Nature that earned a medical degree at the Tokyo Medical and Dental University, and that he learned surgery there. However, in his later conversation with Nature, Moriguchi said that he has a nursing degree from the institution and not a medical degree.

The University of Tokyo confirmed that Moriguchi held a position there from 2006 to 2009, during which he studied “medical economics” and “evaluation of clinical technologies.” Currently, he is a visiting researcher at the university, working in the laboratory of Makoto Mihara in the university hospital’s cosmetic-surgery section, where, according to a secretary, he “comes in once or twice a week.”

Moriguchi also claimed to have a laboratory at MGH and Harvard Medical School, but these institutions only confirmed that Moriguchi was a visiting fellow at MGH in 1999–2000, but he has not been associated with the hospital or the medical school since then.

Nature asked Moriguchi who had funded his iPS cell procedures and where they had been carried out, where his ethical review had taken place and which good manufacturing practice (GMP) facility had produced the necessary clinical-grade iPS cells, Moriguchi referred again to MGH and Harvard Medical School, but he could not name the head of the ethical review board or any contacts at the GMP facility.

Jerome Ritz, co-director of the Connell O’Reilly Cell Manipulation Core Facility at Harvard Medical School, told Nature, “We have not produced any iPS cells for any patients in our facility. I can’t imagine what other facility might have produced these cells.”

What do we have? We have a Japanese researcher who is a liar and who has as much of a problem telling the truth as Barak Obama. This clinical trial clearly never happened and Moriguchi should be banned from further stem cell work.

Human Ovaries Harbor Egg-Making Stem Cell Population

We have read it before, countless times, that women are born with a particular number of eggs and after they die during ovulation or are ovulated, the women is out of eggs and goes through menopause. She does not have the ability to make any more eggs.

Well, another dogma falls by the wayside. As it turns out, egg-making stem cells exist in the woman’s ovaries. An article published in Nature Medicine by Jonathan Tilly and his colleagues who work in a laboratory at Massachusetts General Hospital in Boston, confirms earlier work by Tilly in 2004 that found ovarian stem cells exist in mouse ovaries (Joshua Johnson, et al., Germline stem cells and follicular renewal in the postnatal mammalian ovary. Nature 428, 145-150 (11 March 2004) | doi:10.1038/nature02316), and 2009 publication by a laboratory in Shanghai, China, that confirmed Tilly’s controversial publication (Kang Zou, et al., Production of offspring from a germline stem cell line derived from neonatal ovaries. Nature Cell Biology 11 (2009): 631 – 636).

Despite the rigor of earlier experiments by Tilly’s group and Ji Wu’s, many ovarian experts remained quite skeptical that such a stem cell population existed in humans. This present publications, however, seems to seal the deal. According the Tilly, “This is unequivocal proof that not only was the mouse biology correct, but what we proposed eight years ago was also correct — that there was a human population of stem cells in young adult tissue.” says Tilly.

By listening to their critics, Tilly and is group developed techniques to address the concerns to those skeptical of their findings. They a protocol by which they could isolate and identify mouse ovarian stem cells. This new methods use “fluorescence-activated cell sorting” or FACS. FACS requires the attachment of a fluorescent tag onto the surface of those cells you wish to isolate. Tilly and his group used antibodies linked to a fluorescent dye that could bind tightly to a surface protein called “Ddx4.” Ddx4 is found on the surfaces of ovarian stem cells, but is quickly lost once the stem cells differentiate into egg cells. Treating ovarian cells with the fluorescently-labeled antibodies essentially “painted” them a glowing color. Then the cells were given to a cell sorter than placed cells into one container or another, based on whether or not their surfaces glowed. The cell sorter also distinguishes between whole cells and dead or damaged ones that might fluoresce by accident. In this way, Tilly’s lab invested a protocol for isolating and identifying ovarian stem cells that was highly selective and sensitive.

This new protocol confirmed that Ddx-4-expressing stem cells were present in mouse ovaries. The group did not stop there. They asked if human ovaries had the same stem cell population. They turned to Yasushi Takai, who is a former research fellow in Tilly’s lab, but now works as a reproductive biologist at Saitama Medical University in Japan. Takai provided Tilly with frozen, whole ovaries that had been removed from young women during sexual-reassignment procedures. Tilly says, “It was 9 November when we did the first human FACS sort and I knew immediately that it had worked. I cannot even put into words the excitement — and, to some degree, the relief — I felt.”

From the FACS experiment, Tilly’s group isolated human oogonial stem cells (OSCs). When cultured in the laboratory these OSCs spontaneously produced normal immature oocytes. How did the OSCs do this? To get an inside view of OSC differentiation, Tilly’s team labeled OSCs with green fluorescent protein in order to trace them. Then they injected the green-labeled OSCs into bits of cultured human ovarian tissue, and then transplanted the whole thing under the skin of mice. One to two weeks after transplantation, they found that the OSCs had formed green-glowing cells that greatly resembled ovaries and expressed two ovary-specific genes.

Tilly sounded a cautious note: “There’s no confirmation that we have baby-making eggs yet, but every other indication is that these cells are the real deal — bona fide oocyte precursor cells.” To do this, Tilly must show that the OSC-derived oocytes can be fertilized and form an early embryo. Such work must be done with private funding, since federal funding cannot legally be used for any research that will result in the destruction of a human embryo, regardless of the source of the embryo. Another strategy might be to procure a license from the UK Human Fertilisation and Embryology Authority to do the work with collaborators in the United Kingdom.

Tilly’s experiments have actually converted one scientist who was rather skeptical of his results. Evelyn Telfer, a reproductive biologist at the University of Edinburgh, UK, did not believe Tilly’s initial results. Now, however, she has become a believer. Telfer testified, “I’ve visited [Tilly’s] lab, seen these cells and how they behave. They’re convincing and impressive.” Telfer, has studied in vitro maturation of human eggs, and she wants to work with Tilly to try to grow the OSC-derived eggs to the point at which they are ready for fertilization.

Telfer noted that even though OSCs can form egg-like cells in culture, there is presently no evidence that they can do so in the ovary or that they actually do form new eggs in the ovary. However, the ability to convert OSCs into eggs in vitro might make them usable for in vitro fertilization (IVF), and this achievement would change assisted reproduction forever.

However, Tilly admonishes, “That’s a huge ‘if’.” However, it could means that women who under cancer treatments and experience early menopause could have OSCs removed before treatment and for later fertility use. In fact, according to Tilly, follow-up experiments have shown that OSCs actually exist in the ovaries of women well into their 40s. Even giving women another five years would cover most women affected by IVF.