Induced Pluripotent Stem Cells Made at 100% Efficiency
October 12, 2013
In case you missed it, an important news item in Nature came across the wire recently: Scientists were able to convert skin stem cells into induced pluripotent stem cells at near 100% efficiency. What does this mean?
Let’s take a step back and discuss induced pluripotent stem cells for a moment. These cells come from another kind of stem cell in the body, termed “adult stem cells.” Shinya Yamanaka and John Gurdon, two scientists awarded the Nobel Prize in Physiology and Medicine for their work with induced pluripotent stem cells, used skin cells as their source. We know that our skin has stem cells because our skin renews itself all the time. We shed several pounds of skin per year, only to have it replenished by skin stem cells. Scientists took these skin stem cells and converted them into induced pluripotent stem cells. And we are not limited to skin as a source of adult stem cells; we have adult stem cells to re-supply our blood cells, heart cells, and many other kinds of cells in the body. Bone marrow transplants, which have been used in the clinic for many years, just involve transferring healthy adult stem cells to someone who has diseased blood cells.
Unlike adult stem cells, which yield specific types of cells, pluripotent stem cells are primordial in the sense that they can become almost any cell type in the body. Skin stem cells became more skin cells, but when they are converted to pluripotent stem cells, they are able to become many different types (heart, liver, lung, etc.).
So how do scientists convert skin cells into pluripotent stem cells? The standard procedure for making induced pluripotent stem cells involves activating certain genes that are associated with pluripotency. Four important genes for establishing pluripotency are the OSKM factors (OCT3/4, SOX3, KLF4, and c-MYC), also known as Yamanaka factors. These genes are involved in tagging epigenetic regions in the cell. When scientists over-express these genes (switch them on and turn up the dial, so to speak), they will induce pluripotency in a skin stem cell. There are a few problems with the original techniques, though. The standard procedures for developing induced pluripotent stem cells have about 0.1% efficiency; in other words, only about 0.1% of the starting cells convert into pluripotent stem cells. Furthermore, some of the cells become induced pluripotent stem cells faster than others, resulting in a mixture of adult stem cells and pluripotent stem cells. This is not terribly efficient, particularly given the amount of effort that goes into making these cells – typical procedures require several additions of RNA into the cells over the course of several weeks.
The Nature article linked above reports the development of a procedure that yields significantly higher efficiency in less time, resulting in a more homogeneous mixture of cells. A team of scientists from the Weizmann Institute of Science in Israel discovered that if they turned off a certain gene, Mbd3*, the skin stem cells converted to induced pluripotent stem cells at nearly 100% efficiency within 8 days. As far as the researchers can surmise, “nearly 100% of cells tested had expressed key endogenous pluripotency markers.”
This discovery of the effects of Mbd3 was far from a fluke. Prior research had shown that a complex with Mbd3 as one of its genes was somehow involved in pluripotency, but exactly how this complex was involved was a bit of a mystery. As it turns out, when a cell naturally transitions from a pluripotent stem cell to an adult stem cell, Mbd3 is activated. Mbd3 essentially tells cells to stop being pluripotent through epigenetic signals. The old procedure kept Mbd3 activated while also activating the pluripotent”go” genes, OSKM; the research article authors likened it to having your foot on both the break and the gas at the same time. When the new technique turns off Mbd3, it is like taking your foot off the break. Turning on the OSKM factors while also turning off Mbd3 makes for a much more efficient process to produce induced pluripotent stem cells.
Most of the bioethics issues surrounding induced pluripotent stem cells center on what steps in the process might pose potential health hazards if this technique were used in clinical applications.
One of the key markers verifying that you have induced pluripotent stem cells is that they form teratomas, tumors that are made of multiple cell types. However, tumor formation is a major problem when it comes to using these cells in the clinic. The question is how to make and use induced pluripotent stem cells such that they will produce healthy new cells to replace damaged or diseased ones and then stop when they are done. Perhaps additional research on how Mbd3 operates can provide better ways to ensure the stem cells do not continue making cells to the point that they form tumors.
Furthermore, the procedure reported here involved the use of lentivirus, a viral vector which is used to insert the pluripotent factors into the cell’s genes. This is the standard procedure for making induced pluripotent stem cells, but has been a source of controversy because the viral vectors, particularly the one used for c-MYC, are linked to cancer. Finding alternative methods for introducing the OSKM factors is an ongoing pursuit of research.
Lastly, with induced pluripotent stem cells, it is important to control the cell’s destination. Because these cells are capable of becoming any cell type, once you have induced pluripotent stem cells, you want to be able to convert them to the right cell type. Researchers do not yet fully understand the “signals” that tell the pluripotent stem cells what kind of cells to become, so this part of the process is dependent on the cells picking up the right signals from the body itself. From an ethics standpoint, the intended cell destination is also important. It is one thing to make stem cells to repair the heart. It is another to make stem cells that will form gametes (eggs or sperm), which can potentially be used to form embryos.
“Deterministic Direct Reprogramming of Somatic Cells to Pluripotency,” by Rais, et al, Nature vol. 502, October 2013.
*This is actually an Mbd3/NuRD repressor complex, with Mbd3 being the key player for this particular experiment.