In July 2009, UCLA and Broad Institute researchers analyzed induced pluripotent stem cell lines derived with newer methods that didn't require integration of the reprogramming factors. Their analysis showed different molecular signatures between iPS cells and their embryo-derived counterparts, and these signatures showed a significant degree of overlap with those generated with integrative methods.
"We can’t explain this, but it appears something is different about iPS cells and embryonic stem cells,” said Bill Lowry, a researcher with the Broad Stem Cell Research Center. “The differences are there, no matter whose lab the cells come from, whether they’re human or mouse cells or the method used to derive the iPS cells. Perhaps most importantly, many of these differences are shared amongst lines made in various ways.”
Then in September came work regarding the differences between induced and embryonic stem cells done by UCSD and the Salk Institute.
By creating iPSCs from human neural stem cells without the use of viruses, UCSD and Salk scientists learned something that moved toward an explaination of some of the questions raised by the UCLA/Broad Institute results. What they found was this: While the genetic transcriptional profile of the new iPSCs was closer to that of embryonic stem cells than to human neural stem cells, the iPSCs still carried a transcriptional "signature" of the original neural cell.
"While most of the original genetic memory was erased when the cells were reprogrammed, some was retained," said lead researcher Alysson R. Muotri, UCSD Stem Cell Program researcher who added that in the past it wasn't known if this retinue was caused by the use of viral vectors.
Although genetically identical to the mature body cells from which they are derived, induced pluripotent stem cells (iPSCs) are notably special in their ability to self-renew and differentiate into all kinds of cells. Now researchers from Johns Hopkins and Harvard have detected a remarkable if subtle molecular disparity between the two: Turns out they have distinct "epigenetic" signatures; that is, they differ in what gets copied when the cell divides, even though these differences aren't part of the DNA sequence.
"Relatively little study has been done on the epigenetic nature of stem cells," says Andrew Feinberg, M.D., M.P.H., a professor of medicine at the Johns Hopkins University School of Medicine. He was joined in the research by George Daley, M.D., Ph.D., and colleagues from Harvard University.
To compare and contrast mature connective tissue cells called fibroblasts with the pluripotent stem cells into which they were reprogrammed, the investigators focused on a chemical change known as methylation. This chemical change which, associated with silencing genes, is classified as epigenetic because, although not part of the DNA sequence, it is copied when a cell divides. They identified and then measured so-called differentially methylated regions (DMRs) of genes whose expression was changed in the process of being reprogrammed from a parent cell to a stem cell.
Building on previous research that looked at where differently methylated sites were located in cancer cells, as well as on research that had shown these same sites matching up with many of the methylated areas that had been implicated in the differentiation of normal brain, liver and spleen tissues, the team discovered that the reprogramming of a cell to become a stem cell apparently involves many of the very same DMRs and genes.
"The surprise," says Feinberg, "is that there is such a degree of overlap between the differently methylated regions and genes that are involved in turning a fibroblast into a stem cell and turning a normal cell into a cancer cell."
The researchers suggest in the study that certain sites throughout the genome appear to be generally involved in distinguishing DNA methylation among different cell types and cancers, and these same sites are involved in reprogramming fibroblasts back into stem cells.
The scientists used the CHARM method (comprehensive high-throughput arrays for relative methylation) to survey where, across the genomes of nine human iPS cell lines, genes had been silenced, or turned off, and then compared these DNA methylation sites with those of the fibroblasts the iPS cells were derived from.
"This type of research gets to the fabric of the fundamental differences between stem cells and their parental cells," says Akiko Doi, a doctoral candidate in the graduate program in Cellular and Molecular Medicine at Johns Hopkins. "Clearly, that fabric involves these DMRs, which are essential to our understanding the nature of these potentially therapeutic iPS cells."
As scientists learn more about the epigenetics of reprogrammed cells, they may find new ways of creating them or using them. "If we discover that certain genes or regions are altered in iPS cells," says Feinberg, "then we might be able to target these and come up with new ways of approaching stem cell therapy.
"We can try to correlate these differences with the ways these iPS cells behave, and answer questions such as which ones are more stable and which ones form tumors. If we can use the epigenetic information to characterize these cells, this could inform how we might use them therapeutically."
Adds George Daley, also director of the Stem Cell Transplantation Program at HHMI/Children's Hospital in Boston, "Our data also point to differences between iPS cells and embryonic stem (ES) cells, which everyone has felt were similar if not identical. Such differences may prove important in the behavior of iPS cells in studies on tissue formation and may complicate therapies based on iPS cells. We need to develop ways of generating iPS cells that are a closer match to ES cells in their methylation patterns. Only then will we be confident that iPS cells are a safe replacement for ES cells in research and therapy."
Adapted from earlier Stem Cell Digest Posts and the Johns Hopkins Medical announcement.
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