Stem Cell Reprogramming Technique is
Safer than Previously Thought
Friday, 07 October 2011
Stem cells made by reprogramming
patients' own cells might one day be used as therapies for a host of diseases,
but scientists have feared that dangerous mutations within these cells might be
caused by current reprogramming techniques. A sophisticated new analysis of
stem cells' DNA finds that such fears may be unwarranted.
 |
|
Kristin Baldwin, Ph.D., is an associate
professor in the Scripps Research
Institute's Dorris Neuroscience
Center. Credit: photo courtesy of
The Scripps Research Institute.
|
The induced pluripotent stem cell
(iPSC) technique was first described in 2006. It requires the insertion into an
ordinary non-stem cell of four special genes, whose activities cause the cell
to revert to a state like that of embryonic stem cell. In principle, iPSCs may
be used to repair diseased or damaged tissues, and because they are made from a
patient's own cells, they shouldn't provoke an immune reaction. But recent
studies have found unacceptably high levels of mutations in iPSCs derived from
adult human cells. That has led to widespread suspicion that the reprogramming
process is largely to blame.
In the new study, the Scripps Research
and University of Virginia researchers set out to investigate this issue using
the latest chromosomal error-mapping methods.
"The
techniques that our University of Virginia colleagues brought to this study are
much more sensitive than anything else that's available right now," said Michael J. Boland, a research associate in the Scripps
Research Baldwin lab and co-first author of the paper with Aaron R. Quinlan, a
postdoctoral researcher in Hall's lab. The new methods included a
high-resolution version of a DNA-error-finding technique known as paired-end
mapping, and an advanced algorithm, "HYDRA," for handling the
voluminous mapping data.
To generate the iPSCs, the Scripps
Research team followed the standard, four-gene reprogramming procedure, but
sought to minimize other potential sources of DNA mutations that might have
influenced some previously reported results. The donor cells they selected were
not decades-old human skin cells, but relatively error-free fibroblast cells
from fetal mice. The researchers also kept these fibroblast cells only briefly
in lab dishes before reprogramming them.
When the team members analyzed these
iPSCs they used two strategies to distinguish which mutations were present in
rare donor fibroblast cells and which were newly acquired during reprogramming.
Their advanced techniques also allowed them to find more kinds of mutations,
across a wider range of the genome, than ever before. Yet instead of finding
more mutations, they found almost none.
"We
sequenced three iPSC lines at very high resolution, and were surprised to find
that very few changes to the chromosomal sequence had appeared during
reprogramming," said Boland.
Each of the iPSC lines contained only a
single mutation that probably originated from the reprogramming process; two
affected genes while the other appeared not to. Mutations inherited from the
donor fibroblast cell were present in one pair of lines, while a second line "inherited" none. The
researchers were particularly cheered by the complete absence of new "retro-element transpositions"
— mutations caused by retrovirus-like sequences that burrowed into the
mammalian genome long ago that can become active again in certain cell types.
All cells have ways to suppress these retro-elements, but the suppression
mechanisms in normal cells are different from those in stem cells, so the
researchers had worried that retro-elements would be allowed to escape suppression
during the transition to a stem cell state. While no previous surveys of iPSCs
could detect these mutations, this study showed that despite very sensitive
detection of controls, no retro-elements had become active during
reprogramming.
"That
was is very encouraging, because retro-element mutations can be very damaging
to the genome," Boland said.
Some of the mutations seen in human
iPSCs in previous studies might have been due to incomplete reprogramming that
impaired the cells' DNA-maintenance mechanisms. In this study using mouse
iPSCs, however, there was no doubt that a complete reprogramming to an
embryonic state had occurred: all three iPSC lines were used to produce live,
fertile mice, in work that Boland, Baldwin, and their colleagues described in
Nature in 2009.
"The
mice generated from these cells have survived to a normal lab-mouse lifespan
without obvious diseases that might arise from new DNA mutations," said Baldwin.
Her lab now is trying to determine
whether a reprogramming method similar to the one used with mouse iPSCs in this
study could also yield relatively error-free human iPSCs.
"If
our results with these mouse cells are applicable to human cells, then
selecting better donor cells and using more sensitive genome-survey techniques should
allow us to identify reprogramming methods that can produce human iPSCs that
will be safer or more useful for therapies than current lines," she said.
Source:
Scripps Research Institute
Contact:
Mika Ono
Reference:
Genome Sequencing of Mouse Induced
Pluripotent Stem Cells Reveals Retroelement Stability and Infrequent DNA
Rearrangement during ReprogrammingAaron R. Quinlan, Michael J. Boland, Mitchell L. Leibowitz, Svetlana Shumilina, Sidney M. Pehrson, Kristin K. Baldwin, Ira M. Hall
Cell Stem Cell, Volume 9, Issue 4, 366-373, 4 October 2011, 10.1016/j.stem.2011.07.018
.........
For more on stem cells and cloning, go to CellNEWS at
Posts
No comments:
Post a Comment