(February 4th, 2014) Since January 29th, 2014, biology has to be split into before and after the discoveries of Haruko Obokata. Because, if it is true what the Guardian and many other science channels report, her work is much more than just a new method for cell reprogramming to pluripotency.
Haruko Obokata’s unconventional method, recently published in Nature, is about turning leukocytes, positive for membrane protein CD45, or in fact any cell from a young (one week old) mouse, into pluripotent stem cells by putting them into an acidic (pH 5.7) environment for 30 minutes. Sheer-stress through pipetting apparently has the same effect. And here is the main difference to known reprogramming methods originally developed by Shinya Yamanaka’s lab: no forced gene expression is required. In fact, while Yamanaka was following a widely accepted notion that cells can be reprogrammed by activating or repressing certain genes – large scale screens helped to zero in on the key pluripotency genes – Haruko Obokata has persistently followed her own idea that acidity and squeezing can, one is tempted to say magically, turn terminally differentiated cells into pluripotent stem cells.
The so-called STAP (stimulus-triggered acquisition of pluripotency) cells survive and retain their pluripotency for one or two weeks only, yet, under special culture conditions, they can be made into pluripotent stem cell lines. Already at day 7 after the 30 min acid bath, STAP cells show all key criteria of pluripotency: they express nearly all relevant genes and form teratomas, typical non-invasive tumours, which only pluripotent stem cells are able to form. Most importantly: when injected into a blastocyst, STAP cells readily integrate with the blastocyst embryonic stem cells to form chimeric embryos. In fact, through special assays, the author could prove that these cells can produce entire embryos by themselves, without any chimeras.
If you are impressed now, hold your breath for more. In her second publication in the same Nature issue, Haruko Obokata showed that STAP cells can do much more than all known pluripotent cells: not only could STAP cells reconstruct the entire embryo but also extra-embryonic structures, like placenta, which normally derive from the blastocyst’s trophoblast. During formation of the blastocyst, some embryonic cells become the trophoblast, while others become the pluripotent stem cells of the inner cell mass and form the embryo. Normally, the inner cell mass cells, neither embryonic nor induced pluripotent, can ever contribute to trophoblast formation. Yet, STAP cells proved to express simultaneously both the markers of pluripotent stem cells and trophoblast cells, allegedly explaining their super-pluripotency. For a stem cell biologist, this means in theory that STAP cells are – you’d better sit down for this - totipotent (though Haruko Obokata never uses this word). Totipotency is the Holy Grail of stem cell research, never seen outside the fertilised oocytes and the very early stages of embryogenesis, and certainly never reproduced in a dish.
As a stem cell researcher, I am indeed dumbstruck, as this discovery is so great that the Nobel Prize must be a mere formality now. Every cell can, apparently, at any moment and all by itself revert to the early-embryonic, quasi-totipotent state, only prompted by a brief change in the environment. Truly, the take-home messages of these two publications are so incredible that they actually do appear…unbelievable. Of course, the experimental data seem solid and they have to in order to get accepted by Nature. Having said this, the biological background is still rather puzzling.
Firstly, there are these incomprehensible time windows and cell types: only one week old mice were used, and no adult animals. This makes sense to a stem cell scientist, as the stem cell potential in young organisms is much more pronounced than in adult ones. On the other hand, why doesn’t the author use new-born mice or post-gastrulation embryos? One would expect even more impressive stem cell efficiency from embryos. Should STAP reprogramming only work in one week old mice but not in older or younger animals or embryos? This would be very difficult to explain biologically. All tested cells, mostly terminally differentiated and rarely dividing cells from the brain, muscle, liver etc. became pluripotent after acid treatment, with comparable efficiency. Yet, the author used exclusively the CD45-positive leukocytes. These cells have no stem cell potential and divide poorly. This should have convinced sceptical referees, concerned about contaminations by somatic stem cells (actually it would be just as exciting if these cells were to achieve quasi-toti-potency). Yet, the hottest candidates, the CD34-positive hematopoietic stem cells practically failed to become STAP (<2%), another huge surprise.
Secondly, Haruko Obokata states that STAP cells are strongly affected by apoptosis at day 7 and rapidly lose their pluripotency. Moreover, their proliferation rate is hardly measurable anyway. Yet, these dying and differentiating cells are capable of producing entire embryos together with a trophoblast, while many common laboratory induced pluripotent stem cell lines, unlimitedly proliferating and viable, are often not even capable of forming chimeric embryos. Because STAP cells did not survive enzymatic dissociation, the author cut their colonies with a small scalpel and injected the chunks into the blastocysts. Common embryonic or induced pluripotent stem cells would not cope with this procedure and certainly would not build chimeras. Yet, the sickly STAP cells had no problem at all assembling a complete embryo, together with placenta, from these crude bits and pieces.
Thirdly, as STAP cells quickly die and differentiate, the author could rescue them through the use of the adrenocorticotropic hormone, ACTH, (a method for stem cell derivation, published earlier by one of the study’s co-authors). Unfortunately though, STAP cells lose their unique quasi-totipotency and become a common, boring pluripotent cell line. The consequence is that you cannot borrow the STAP cells through scientific exchange and analyse their unique totipotency features in your own lab. You would have to start everything anew, in order to reproduce Haruko Obokata’s experiments. I think I will wait until other labs do this before I jump on the current tidal wave of media excitement and decide to throw my current understanding of biology overboard.
Fourthly: if short term changes in pH or sheer stress were to have such a tremendous effect on somatic cells, would this not imply an equally tremendous danger of cancer? Bluntly asked: why don’t STAP-derived teratomas constantly form inside the stomach (low pH) or inside the aorta (high sheer stress)? In fact, not even stomach cancers bear any similarities to teratomas. Maybe it is indeed the case that only CD45-positive leukocytes of one-week-old mice are susceptible and the next day the mice are already safe from teratoma formation. Nothing is impossible anymore.
Photo: Fotolia/PRILL Mediendesign
A similar version of this article first appeared in German at www.laborjournal.de.