Induced Pluripotent Stem Cells from Reprogramming Adult Differentiated Skin Cells
By Gershom Chua
Introduction
Way back in 1995, a then relatively unknown molecular biologist James Thomson succeeds in isolating and deriving stem cells, or cells that have the ability to differentiate further into any desired kind of cell to facilitate development, from non-human primates. This success would lead him to apply the same breakthrough to human embryonic stem cells in 1998 which caused a global buzz around the possibilities of stem cell research in curing diseases such as Parkinson’s disease, leukemia, diabetes, cancer, etc., against the ethical and moral dilemmas surrounding the process by which he obtained this breakthrough. Since then, stem cell research has proven controversial because of the implications it presents to reproductive human cloning and the moral value of human life fought for by pro-life movements. Scientists have been pushing for continued research despite these grounds because of what they deem to be far beneficial results that stem cell research may eventually bring to treating diseases, clinical applications, drug development, transplant medicine, and beyond. This debate has been mirrored in American state policies concerning it, shifting from the withdrawal of support via federal or government funds given to further stem cell research. This withdrawal then limited the development of stem cell research this past decade within the confines of support and funding from private institutions and companies.
Recently, developments from four significant teams have at last uncovered a new method into stem cell research that may just prove to be able to pass the ethical and moral debates due to its refined process and push the potentials of this significant breakthrough. Since debates and controversy arise only because of the involvement of extracting stem cells from human embryos regardless of the method by which it was done (as controversy still followed refined processes of obtaining stem cells from excess embryos from in vitro fertilization, embryos from abortions, etc which are relatively more humane than the original procedure of destroying live human embryos), the possibilities that open up through the findings of four significant teams from various universities in the past three years allow this ethical hurdle to be pushed aside to give way to progress. The findings have shown induced pluripotent stem (iPS) cells derived from reprogrammed adult differentiated skin cells to share quite similar characteristics with traditional embryonic stem cells, therefore presenting an alternative to the more problematic source of stem cell study.
Background
As early as three years ago, a Japanese team headed by Shinya Yamanaka and Kazutoshi Takahashi from Kyoto University have found out that by reprogramming adult differentiated mouse skin cells through somatic cell nuclear transfer or SCNT, they could create skin cells that were characteristically undifferentiated and that closely resembled traditional embryonic stem cells. These cells were then dubbed as induced pluripotent stem cells or iPS cells because of their similarities with the embryonic stem cell. The team used a set of four genes, namely OCT3/4, SOX2, c-MYC, and KLF4, to induce this embryonic stem cell-like behavior in the mouse adult skin cells.
From this study, the same team worked on the possibilities of applying their findings to human skin cells and eventually succeeded by replicating the process with some minor changes.
This lead to other teams from US universities confirming and refining on their process by changing the particular genes they used to induce this stem cell-like behavior in skin cells , the number of genes used, and by changing the type of vector used to deliver these genes into the cell.
Timeline
2006 Shinya Yamanaka, Kazutoshi Takahashi, and their team from Kyoto University successfully reprogrammed mature differentiated mouse cells to create cells similar to embryonic undifferentiated stem cells (or later known as iPS or induced pluripotent stem cells)
2007 Shinya Yamanaka and the same team from Kyoto University successfully adapts the process to human adult differentiated skin cells
James Thomson, Junying Yu, and their team from the University of Wisconsin confirm the Japanese team’s findings and are successful at refining the process, replacing two of the four genes used by the previous team that may prove to be eventually dangerous
Shinya Yamanaka and the same team from Kyoto University refine their process even further by reducing the genes used from four to three, significantly reducing risks of dangers brought by using more genes
2008 Kathrin Plath, William Lowry, and their team from the University of California, Los Angeles confirm the previous processes of Yamanaka’s and Thomson’s findings which the team finished just as Yamanaka’s and Thomson’s papers have been published
Kathrin Plath and the same team from UCLA successfully created cardiac cells from reprogramming differentiated mouse cells, a concrete step towards the possibilities of iPS cells
Matthias Stadtfeld, Konrad Hochedlinger, and colleagues from the Massachusetts General Hospital, Boston created iPS cells from differentiated mouse skin cells using a non-DNA altering virus or adenovirus over previously used retroviruses
Significant Contributions
Shinya Yamanaka, Kazutoshi Takahashi, and the Kyoto University Team
Their early venture into the study of reprogrammed mouse adult skin cells saw them utilizing retroviruses in delivering the set of four genes together with the nucleus from mammalian oocytes or immature ovum into the adult skin cells. The four genes were used because of their ability to induce this embryonic stem cell-like behavior in the adult skin cells that were then cultured to produce iPS cells.
They later applied this to human adult skin cells. They engineered cultured human skin cells or fibroblasts to enable ease in introducing new genes. The genes that were then used in the process they did in mouse adult skin cells were also delivered into these cultured fibroblasts via a retrovirus vector. After weeks, the team found the reprogrammed adult skin cells to have produced colonies that closely resembled human embryonic stem cells. After much testing, the team concluded that the colonies or iPS cells really were virtually similar to traditional stem cells that would have been otherwise obtained controversially. Further test revealed the ability of these iPS cells to be later induced to differentiating into any of the three major cell types, namely ectoderm, mesoderm, and endoderm.
The same team later proved the use of four genes could actually be reduced to a mere three. The finding, even though not as spectacular as the prior one, is still significant in that they were able to eliminate c-MYC from the set of genes the used. Eliminating c-MYC from the set of genes in the formula meant eliminating a possible threat of inducing cancer in the cultured iPS cells. This finding thus refines on the process and is continually refined until present.
James Thomson, Junying Yu, and the University of Wisconsin Team
Nearly a decade after pioneering stem cell research, James Thomson once again enters the public terrain by confirming the results of the Japanese team but using a different set of genes to induce the stem cell-like behavior in adult skin cells. Instead of using the genes KLF4 and the potentially cancer-inducing c-MYC, they used NANOG and LIN28, both of which do not seem to present relatively the same degree of danger as the previous two. This study is significant in that it was made after the Japanese team’s initial success with the four original genes and prior to the same team’s discovery of using three genes instead.
Kathrin Plath, William Lowry, and the University of California, Los Angeles Team
Though having published the results of their study much later than the two previous teams’, the team led by Kathrin Plath of UCLA proves significant because they have not only confirmed the two previous teams’ findings but also made concrete progress soon after. Reverting back to the source of this breakthrough after having success with adult skin cells, the team was able to successfully create cardiovascular from iPS cells first derived from mouse adult skin cells. This was achieved through culturing iPS cells on a protein matrix that directs traditional embryonic stem cells into differentiating into cardiovascular progenitor cells or immature heart cells that can later further develop to mature heart cells that will be able to perform various purposes. The created progenitor cells were then isolated using KDR, a growth factor receptor found on the surface of progenitor cells. They were then induced to developing into cardiomyocytes or mature heart cells that manage the heartbeat, endothelial cells or cells that line the walls of blood vessels, etc. The team succeeded to the point of observing the created cardiomyocytes beating inside the Petri dish in which they were cultured in.
Now, the team is studying if their success in creating mouse cardiovascular cells from iPS cells can also be translated to human cardiovascular cells from iPS cells cultured from human adult skin cells.
Matthias Stadtfeld, Konrad Hochedlinger, and the Massachusetts General Hospital, Boston Team
The team led by Matthias Stadtfeld from the Massachusetts General Hospital confirms other team’s previous findings but have gone on to refine the process even further on the level of the vector they used to reprogram the adult skin cell. Working on mouse adult skin cells, they have been able to produce iPS cells with the same set of genes delivered not by retroviruses as were used by previous studies but by adenoviruses. They changed vectors primarily because retroviruses are known to activate cancers and tumors which would render the whole premise of stem cell research useless. By using adenoviruses as vectors, they have observed that these vectors eventually do not leave any trace in the cells produced after a few cycles of cell division in the created iPS cells. This clearing of the vector problem paves the way for truly testing the method to human skill cell application.
Advantages and Disadvantages
The creation of induced pluripotent stem cells or iPS cells as an alternative for traditional embryonic stem cells present various advantages that drive scientific research on it to progress as fast as it does now. It is advantageous that iPS cells are derived from reprogramming adult skin cells, therefore not having to tamper with human embryos that has been the cause for the controversy after all. “This is the beginning of the end of the controversy”, as James Thomson commented. This would not only end controversy on stem cell research but push forward other breakthroughs because of a change in perception of science that this would bring. This is also advantageous because of the degree of which iPS cells are similar to embryonic stem cells. As ScienceDaily.com reported, “The reprogrammed cells were not just functionally identical to embryonic stem cells. They also had identical biological structure, expressed the same genes and could be coaxed into giving rise to the same cell types as human embryonic stem cells.” With this similarity, iPS cells then promises to cure diseases that are believed to be cured by embryonic stem cells such as leukemia, Parkinson’s disease, diabetes, spinal cord injuries, cancers, etc. without any ethical or moral issue to prevent it from doing so. Another advantage of iPS cells is that they are obtained from the same patient who will be treated by using reprogramming them and reinserting them, therefore taking out the rejection of the iPS cells as a possible issue. Lastly, by using partially specific cells derived from iPS cells such as the cardiovascular cells cultured from mouse iPS cells, these partially specific cells will lessen the chances of developing tumors and other undesired growths because of the specificity of their nature when they are reinserted into patients.
There are, however, disadvantages that should be weighed in considering iPS cells. First, despite this shift from controversial sources of stem cells such as obtaining them from living human embryos, researchers will still need to have samples of embryonic stem cells to continually assess the progress of iPS cells. This is because stem cells are well considered to be “the golden standard” by which iPS cells are to be compared to, therefore, to fully progress with iPS cells, continuous input of stem cell samples are still needed. Second, in using iPS cells to treat diseases, one encounters the same problems of that in gene therapy where one should fully understand the disease and which genes should be specifically corrected and replaced. Lastly, much progress has been seen in culturing iPS cells in mouse skin cells and iPS cell culture has only just begun in humans, therefore presenting us with the dilemma of how exactly to translate findings in mouse skin cell samples to human skin cell applications. This has been proven possible by the actual creation of iPS cells from human skin cells, but the problem remains that further more complicated findings may prove hard to translate to human cells.
Future
Having established the creation of iPS cells in both mouse skin cells and human skin cell to be possible by having succeeded in producing samples from the four main teams that have done significant work in this area, the future lies ahead with infinite possibilities. Since the vector carriers of the genes to induce the stem cell-like behavior in iPS cells have been established to be able to shift to adenoviruses that are completely safe in delivering genes into skin cells without trace, now what is left to study is which genes would be perfect in further reprogramming. Since only a handful of genes have been tapped into this technology, the possibility of finding the best combination that would yield the least risk of dangers and undesired effects after reprogramming is there but would take some time before it is found. Also, the fact that cardiovascular cells have been cultured from mouse iPS cells show just how much farther we can go with the possibilities that will arise in the future after much development and progress is reached in this area.
Implications
The implications this alternative to using embryonic stem cells proves to have range to a lot of areas. Scientifically, this discovery proves that further development in our understanding of the possibilities found in our bodies need not go against ethics and morals. Ethically, this discovery paves a way for science and breakthroughs and peoples’ ethic and moral standards to come together in a state of harmony, where one does not have to compromise to meet the other’s needs. Human ethics and morals will not have to be compromised to see progress that may eventually benefit us all. Politically, this breakthrough will once again open the governments’ doors to funding scientific research as nothing is compromised. This is seen by the 09 March 2009 signing of US President Barack Obama of a new agreement for federal funds to be put into supporting stem cell research mainly because it proves not to go against ethic and moral concerns, as opposed to prior this breakthrough when the Bush administration vetoed the Stem Cell Research Enhancement Act of 2007 for reasons. iPS cell technology then can be seen as paving a new way for science to be recognized and accepted throughout societies as it does not need to go against present values these peoples hold on. Economically, since the technology is surely at its infancy and if in the near future proves to be safe and directly applicable, it will further push existing hegemony of class and wealth because this would surely entail quite a hefty sum because of its novelty. It will take another breakthrough after this one to lower the costs for clinical and medical application on patients.
References
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Baker, Monya. 2007. From Skin Cell to Stem Cell. Nature Reports.
Smith, Michael. Adult Skin Cells Reprogrammed into Stem Cells for Disease Research. Med Page Today.
Falco, Miriam. Human Skin Cells Reprogrammed to Act Like Stem Cells. CNN Health.com.
Baker, Monya. 2007. Adult Cells Reprogrammed to Pluripotency, Without Tumors. Nature Reports.
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Colen, B.D. Advance in Pluripotent Cell Creation. Harvard University Gazette Online.
Quill, Elizabeth. 2009. Stem Cell Efforts Take Steps. Science News. Volume 175. Issue 1.Washington: Science for Society and the Public. p.20.
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