Comparing two cellular reprogramming methods from one donor’s cells yields good news for iPSCs

In 2012, a mere six years after his discovery of induced pluripotent stem cells (iPSCs), Shinya Yamanaka was awarded the Nobel Prize in Medicine. Many Nobel winners aren’t recognized until decades after their initial groundbreaking studies. That goes to show you the importance of Yamanaka’s technique, which can reprogram a person’s cells, for example skin or blood, into embryonic stem cell-like iPSCs by just adding a small set of reprogramming factors.

These iPSCs are pluripotent, meaning they can be specialized, or differentiated, into virtually any cell type in the body. With these cells in hand, researchers have a powerful tool to study human disease and to develop treatments using human cells directly from patients. And at the same time, this cell source helps avoid the ethical concerns related to embryonic stem cells.


Induced pluripotent stem cell (iPSC) colonies.
Image Credit: Joseph Wu

Still, there has been lingering uneasiness about how well iPSCs match up to embryonic stem cells (ESCs), considered the gold-standard of pluripotent stem cells. One source of those concerns is that the iPSC method doesn’t completely reprogram cells and they retain memory of their original cell source, in the form of chemical – also called epigenetic – modifications of the cells’ DNA structure. So, if a researcher were to make, say, heart muscle cells from iPSCs that have an epigenetic memory of its skin cell origins, any resulting conclusions about a given disease study or cell therapy could be less accurate than ESC-related results. But a report published yesterday in PNAS should help relieve these worries.

The CIRM-funded study – a collaboration between the labs of Joseph Wu and Michael Synder at Stanford University and Shoukhrat Mitalipov at Oregon Health & Science University – carried out an exhaustive series of experiments that carefully compared the gene activity and cell functions of iPSC-derived cells with cells derived from embryonic stem cells. The teams sought to compare cells generated from the same person to be sure any differences were not the result of genetics. To make this “apples-to-apples” comparison, they generated embryonic stem cells using another reprogramming technique called somatic cell nuclear transfer (SCNT).

With SCNT, a nucleus from an adult cell is transferred to an egg which has its own nucleus removed. The resulting cell becomes reprogrammed back into an embryo from which embryonic stem cells are generated – the researchers call them NT-ESCs for short. In this study, the skin cell sample used for making the iPSCs and the cell nucleus used for making the NT-ESCs came from the same person. In scientific lingo, the iPSCs and SCNT stem cells are considered isogenic.

Now, it turns out the NT-ESC reprogramming process is more complete and eliminates epigenetic memory of the original cell source. So why even bother with iPSCs if you have NT-ESCs? There are big disadvantages with SCNT: it’s a complex technique – only a limited number of labs pull it off – and it requires donated human eggs which carries ethical issues. So, if a direct comparison iPSCs and SNCT stem cells shows little difference then it would be fair to argue that iPSCs can replace NT-ESCs for deriving patient-specific stem cells.

And that’s exactly what the teams found, as Dr. Wu summarized it to me in an interview:

“Direct comparison between differentiated cells derived from iPSCs and SCNT had never been performed because it had been difficult to generate patient-specific ESCs by the SCNT method. Collaborating with Dr. Shoukhrat Mitalipov at Oregon Health & Science University and Dr. Michael Snyder at Stanford University, we compared patient-specific cardiomocytes (heart muscle cells) and endothelial (blood vessel) cells derived by these two reprogramming methods (SCNT and iPSCs) and found they were relatively equivalent regarding molecular and functional features.”

PSC-ECs2 copy

Blood vessel cells derived by iPSC (left) and SCNT (right) reprogramming methods.
Image credit: Joseph Wu

Because the heart muscle and blood vessel cells were similar regardless of reprogramming method, it suggests that the epigenetic memory that remained in the iPSCs is less of a worry. Dr. Wu explained to me this way:


Joseph Wu

“If iPSCs carry substantial epigenetic memory of the cell-of-origin, it is unlikely these iPSCs can differentiate to a functional cardiac cell or blood vessel cell. Only the stem cells free of significant epigenetic memory can differentiate into functional cells.”


Hopefully these results hold up over time because it will bode well for the countless iPSC-related disease studies as well as the growing number of iPSC-related projects that are nearing clinical trials.

More than Meets the Eye: Stem Cells Generated using Different Methods Produce Different Types of Cells

What’s the best way to make a fully versatile, ‘pluripotent,’ stem cell? Three different methods each have their pluses and minuses. But now new research has found that the stem cells created by each method, while similar on the surface, show vast differences.

The findings, published online today in the journal Nature, reveal new insights into stem cells’ underlying cellular machinery—which is of utmost importance as researchers transform their discoveries from the lab and into much-needed therapies for patients.

Scanning electron micrograph of cultured human neuron from induced pluripotent stem cell.  [Credit: Mark Ellisman and Thomas Deerinck, National Center for Microscopy and Imaging Research, UC San Diego]

Scanning electron micrograph of cultured human neuron from induced pluripotent stem cell. [Credit: Mark Ellisman and Thomas Deerinck, National Center for Microscopy and Imaging Research, UC San Diego]

Stem cells have held promise for regenerating tissues, or even organs, lost or damaged by injury or disease. This is due to stem cells’ ‘pluripotency’—their ability to transform into virtually any cell in the body. Initially, scientists used stem cells extracted from unused embryos that consenting couples had donated to research. But the use of these so-called embryonic stem cells, or ES cells, has since been limited due to ethical considerations and early limits to federal funding.

So scientists have been on the hunt for an alternative method of creating pluripotent cells. And so far, they have come up with two.

One, called somatic cell nuclear transfer (SCNT) takes the genetic material of an adult cell and transplants it into an unfertilized egg. The second method transforms adult cells, such as skin or blood, back into embryonic-like stem cells—called induced pluripotent stem cells, or iPS cells—by manipulating various genes.

Each of the newer methods has its pluses and minuses—but which produces cells that most closely resemble ES cells, still considered the “Gold Standard” in stem cell biology? Since the success of the SCNT technique is so recent, no one had taken a close look until now. So a collaboration of researchers from the University of California, San Diego (UCSD), The Salk Institute for Biological Sciences and Oregon Health & Science University (OHSU), compared the two methods side by side. And what they found was surprising.

Dr. Louise Laurent, co-senior author from UCSD, explained in today’s news release:

“The nuclear transfer ES cells are much more similar to real ES cells than the iPS cells. They are more completely reprogrammed and have fewer alterations in gene expression and DNA methylation levels that are attributable to the reprogramming process itself.”

iPS cell technology, which was pioneered in 2006 by Shinya Yamanaka, offers a series of advantages over traditional ES cells. As Laurent continued:

“The ability to make personalized iPS cells from a patient that could be transplanted back into that patient has generated excitement because it would eliminate the need for immunosuppression.”

iPS cells have generated so much excitement, in fact, that Yamanaka was awarded the 2012 Nobel Prize in Physiology or Medicine for developing this technique.

The SCNT method was developed more recently by OHSU’s Dr. Shoukhrat Mitalipov. The current researchers generated lines of cells using both methods. After confirming that each line was, in fact, pluripotent, they used advanced genomics techniques to examine the biochemical process called ‘DNA methylation’ in each line.

DNA methylation is a fundamental chemical process within each cell. It’s responsible for switching key genes on and off at precise intervals. In recent years, researchers have discovered that the order and timing of this process is vital for the correct development of the cell. As Dr. Joseph Ecker, co-senior author from the Salk Institute, explained:

“If you believe that gene expression and DNA methylation are important, which we do, the closer you get to the patterns of embryonic cells, the better. Right now, nuclear transfer cells look closer to the embryonic stem cells than do the iPS cells.”

However, while the scientists confirmed that SCNT cells more closely resemble ES cells, the process of producing them is far from ideal. First, the SCNT method is technically difficult. And second, federal funds still cannot be used in this procedure—representing a significant hurdle to being widely adopted.

On the other hand, iPS cell generation is, by comparison, a much easier process technically. So perhaps these findings can spur the development of an improved method, taking the technological ease of iPS cell generation and marrying it with the accuracy of the SCNT method. Laurent argues that this could yield a new and improved approach:

“Our results have shown that widely used iPS cell reprogramming methods make cells that are similar to standard ES cells in broad strokes, but there are important differences when you look really closely. By using the egg cell to do the job, we can get much closer to the real thing. If we can figure out what factors in the egg drive the reprogramming process, maybe we can design a better iPS cell reprogramming method.”