Simple method turns blood cells into virus-free beating heart cells
12 April 2011
A simplified, cheaper, all-purpose method that can safely turn
blood cells into heart cells has been developed at Johns Hopkins
University in the US. The method is virus-free and produces heart cells
that beat with nearly 100% efficiency, according to the research team
To get stem cells taken from one source (such as blood) and
develop them into a cell of another type (such as heart), scientists
generally use viruses to deliver a package of genes into cells to,
first, get them to turn into stem cells.
However, viruses can mutate genes and initiate cancers in newly
transformed cells. To insert the genes without using a virus,
Zambidis’ team turned to plasmids, rings of DNA that replicate
briefly inside cells and eventually degrade.
Adding to the complexity of coaxing stem cells into other cell
types is the expensive and varied recipe of growth factors,
nutrients and conditions that bathe stem cells during their
transformation. The recipe of this “broth” differs from lab to lab
and cell line to cell line.
“We took the recipe for this process from a complex minestrone to
a simple miso soup,” says Elias Zambidis MD PhD, assistant professor
of oncology and pediatrics at the Johns Hopkins Institute for Cell
Engineering and the Kimmel Cancer Center.
Zambidis says, “Many scientists previously thought that a
nonviral method of inducing blood cells to turn into highly
functioning cardiac cells was not within reach, but we’ve found a
way to do it very efficiently and we want other scientists to test
the method in their own labs.” However, he cautions that the cells
are not yet ready for human testing.
Reporting in the April 8 issue of PLoS ONE, Zambidis’
team described what he called a “painstaking, two-year process” to
simplify the recipe and environmental conditions that house cells
undergoing transformation into heart cells. They found that their
recipe worked consistently for at least 11 different stem cell lines
tested and worked equally well for the more controversial embryonic
stem cells, as well as stem cell lines generated from adult blood
stem cells, their main focus.
The process began with Johns Hopkins postdoctoral scientist Paul
Burridge PhD, who studied some 30 papers on techniques to create
cardiac cells. He drew charts of 48 different variables used to
create heart cells, including buffers, enzymes, growth factors,
timing, and the size of compartments in cell culture plates. After
testing hundreds of combinations of these variables, Burridge
narrowed the choices down to between four to nine essential
ingredients at each of three stages of cardiac development.
Beyond simplification, an added benefit is reduced cost. Burridge
used a cheaper growth media that is one-tenth the price of standard
media for these cells at $250 per bottle lasting about one week.
Zambidis says that he wants other scientists to test the method
on their stem cell lines, but also notes that the growth “soup” is
still a work in progress. “We have recently optimized the conditions
for complete removal of the fetal bovine serum from one brief step
of the procedure — it’s made from an animal product and could
introduce unwanted viruses,” he says.
In their experiments with the new growth medium, the Hopkins team
began with cord blood stem cells and a plasmid to transfer seven
genes into the stem cells. They delivered an electric pulse to the
cells, making tiny holes in the surface through which plasmids can
slip inside. Once inside, the plasmids trigger the cells to revert
to a more primitive cell state that can be coaxed into various cell
types. At this stage, the cells are called induced pluripotent stem
Burridge then bathed the newly formed iPSCs in the now simplified
recipe of growth media, which they named “universal cardiac
differentiation system.” The growth media recipe is specific to
creating cardiac cells from any iPSC line.
Finally, they incubated the cells in containers that removed
oxygen down to a quarter of ordinary atmospheric levels. “The idea
is to recreate conditions experienced by an embryo when these
primitive cells are developing into different cell types,” says
Burridge. They also added a chemical called PVA, which works like
glue to make cells stick together.
Nine days later, the nonviral iPSCs turned into functional,
beating cardiac cells, each the size of a needlepoint.
A 'beating' cardiomyocyte
Burridge manually counted how often iPSCs formed into cardiac
cells in petri dishes by peering into a microscope and identifying
each beating cluster of cells. In each of 11 cell lines tested, each
plate of cells had an average of 94.5% beating heart cells. “Most
scientists get 10% efficiency for IPSC lines if they’re lucky,” says
Zambidis and Burridge also worked with Johns Hopkins University
bioengineering experts to apply a miniversion of an
electrocardiograph to the cells, which tests how cardiac cells use
calcium and transmit a voltage. The resulting rhythm showed
characteristic pulses seen in a normal human heart.
Virus-free, iPSC-derived cardiac cells could be used in
laboratories to test drugs that treat arrhythmia and other
conditions. Eventually, bioengineers could develop grafts of the
cells that are implanted into patients who suffered heart attacks.
Zambidis’ team has recently developed similar techniques for
turning these blood-derived iPSC lines into retinal, neural and
The research was funded by the Maryland Stem Cell Research Fund
and the National Institutes of Health.