Induced pluripotent stem cells III

There is still some news from here that we need to look at. It has to do with reducing the risk of tumorigenicity by using a signaling protein Wnt3a – a member of the Wnt family of proteins – in place of the c-Myc transcription factor for inducing pluripotency in differentiated adult cells.

This press release gives the executive summary:

Embryonic-like Stem Cells Can Be Created Without Cancer-causing Gene (8/6/08)

Currently, IPS cells can be created by reprogramming adult cells through the use of viruses to transfer four genes (Oct4, Sox2, c-Myc and Klf4) into the cells' DNA. The activated genes then override the adult state and convert the cells to embryonic-like IPS cells.

However, this method poses significant risks for potential use in humans.

First, the viruses employed in the process, called retroviruses, are associated with cancer because they insert DNA anywhere in a cell's genome, thereby potentially triggering the expression of cancer-causing genes, or oncogenes. Second, c-Myc is a known oncogene whose overexpression can also cause cancer. For IPS cells to be employed to treat human diseases such as Parkinson's, researchers must find safe alternatives to reprogramming with retroviruses and oncogenes.

Earlier research has shown that c-Myc is not strictly required for the generation of IPS cells. However, its absence makes the reprogramming process time-consuming and highly inefficient.

To bypass these obstacles, the Whitehead researchers replaced c-Myc and its retrovirus with a naturally occurring signaling molecule called Wnt3a. When added to the fluid surrounding the cells being reprogrammed, Wnt3a promotes the conversion of adult cells into IPS cells.

What amounts to a crude form of gene therapy has been used to make IPS cells. The idea is to insert extra copies of genes for 4 different transcription factors into a cell's DNA in order to raise the expression level of those factors. The problem is that every insertion of a gene into a cell's DNA risks damage to some other random gene in the DNA. Wnt3a, on the other hand, is a signaling protein that normally affects cells only by attaching to receptors on the cell surface.

So what has been accomplished here is that the number of transcription factor genes that need to be inserted into the DNA is reduced from 4 to 3. In addition, the factor that is eliminated, c-Myc, has tumorigenicity risks of its own. Therefore, this research represents a small but useful improvement. However, it is probably only a first step.

More about the present research: here

We discussed earlier research aimed at eliminating use of c-Myc in making IPS cells here.

In fact, just a little bit earlier than the research discussed above, a team from Germany reported, on July 31 in Nature, making IPS cells by adding just two transcription factors. However, they didn't start with adult somatic cells, but with neural stem cells that already had higher expression levels of Sox2 and c-Myc. Given that, they needed to add only Oct4 and either Klf4 or more c-Myc. Here's the abstract:

Pluripotent stem cells induced from adult neural stem cells by reprogramming with two factors

Here we show that adult mouse neural stem cells express higher endogenous levels of Sox2 and c-Myc than embryonic stem cells, and that exogenous Oct4 together with either Klf4 or c-Myc is sufficient to generate iPS cells from neural stem cells. These two-factor iPS cells are similar to embryonic stem cells at the molecular level, contribute to development of the germ line, and form chimaeras. We propose that, in inducing pluripotency, the number of reprogramming factors can be reduced when using somatic cells that endogenously express appropriate levels of complementing factors.

Keep in mind that there's a further variable that's important here: the efficiency of the process, i. e. the yield of IPS cells obtained as a percentage of original cells at the beginning. It should be obvious that a fair amount of work still needs to be done to find a method of making IPS cells that's both efficient and produces cells that are potentially safe to use in therapeutic applications (as opposed to pure research).

OK, enough of that. Let's move on to something new.

One of the interesting questions about IPS cells is about exactly how close they are to actual embryonic stem cells, which are pluripotent by definition. The best way to measure the degree of closeness is by comparing gene expression levels between embryonic stem cells and IPS cells.

The next research has done exactly that. In fact, it studies gene expression levels for stem-like cells obtained from a wide variety of sources:

A new test distinguishes embryonic stem cells and those with equal therapeutic potential (8/24/08)

To distinguish adult stem cells from pluripotent cells, Loring’s team compared the gene activity of about 150 stem cell samples of various types, including reprogrammed cells, embryonic stem cells and neural stem cells. Out of this comparison popped 299 interacting genes that form what the researchers call a pluripotency network, or PluriNet. Measuring the activity of these genes could reliably distinguish the different kinds of stem cells, the team reports.

Here's the abstract for this research:

Regulatory networks define phenotypic classes of human stem cell lines

We report here the creation and analysis of a database of global gene expression profiles (which we call the 'stem cell matrix') that enables the classification of cultured human stem cells in the context of a wide variety of pluripotent, multipotent and differentiated cell types. Using an unsupervised clustering method to categorize a collection of ∼150 cell samples, we discovered that pluripotent stem cell lines group together, whereas other cell types, including brain-derived neural stem cell lines, are very diverse. Using further bioinformatic analysis we uncovered a protein–protein network (PluriNet) that is shared by the pluripotent cells (embryonic stem cells, embryonal carcinomas and induced pluripotent cells). Analysis of published data showed that the PluriNet seems to be a common characteristic of pluripotent cells, including mouse embryonic stem and induced pluripotent cells and human oocytes. Our results offer a new strategy for classifying stem cells and support the idea that pluripotency and self-renewal are under tight control by specific molecular networks.

Tags: stem cells, embryonic stem cells, induced pluripotent stem cells