Growth factor from endothelial cells enables mass production of
adult stem cells
21 April 2010
In a leap toward making stem cell therapy widely available,
researchers at Weill Cornell Medical College have discovered that
endothelial cells, the most basic building blocks of blood vessels,
produce growth factors that can grow copious amounts of adult stem cells
and their progeny over the course of weeks. Until now, adult stem cell
cultures would die within four or five days despite best efforts to grow
This new finding sets forth the innovative concept that blood
vessels are not just passive conduits for delivery of oxygen and
nutrients, but are also programmed to maintain and proliferate stem
cells and their mature forms in adult organs. Using a novel approach
to harness the potential of endothelial cells by "co-culturing" them
with stem cells, the researchers discovered the means to manufacture
an unlimited supply of blood-related stem cells that may eventually
ensure that anyone who needs a bone marrow transplant can get one.
The vascular-cell model established in this study could also be
used to grow abundant functional stem cells from other organs such
as the brain, heart, skin and lungs. An article detailing these
findings appears in the March 5 issue of the journal Cell Stem
In adult organs, there are few naturally occurring stem cells, so
using them for organ regeneration is impractical. Until now,
strategies to expand cultures of adult stem cells, which invariably
used animal-based growth factors, serum, and genetically manipulated
feeder cells, have only been marginally successful. This study,
which employs endothelial cells to propagate stem cells without
added growth factors and serum, will likely revolutionize the use of
adult stem cells for organ regeneration, as well as decipher the
complex physiology of the adult stem cells.
"This study will have a major impact on the treatment of any
blood-related disorder that requires a stem cell transplant," says
the study's senior author, Dr Shahin Rafii, the Arthur B Belfer
Professor in Genetic Medicine, co-director of the Ansary Stem Cell
Institute and a Howard Hughes Medical Institute Investigator, at
Weill Cornell Medical College. Currently, stem cells derived from
bone marrow or umbilical cord blood are used to treat patients who
require bone marrow transplants. Most stem cell transplants are
successful, but because of the shortage of genetically matched bone
marrow and umbilical cord blood cells, many patients cannot benefit
from the procedure.
"Over the last few decades, substantial funding has been spent to
develop platforms to expand adult stem cell cultures, but these
efforts have never been able to coax an authentic adult stem cell to
self-renew beyond a few days," continues Dr Rafii. "Most stem cells,
even in the presence of multiple growth factors, serum, and support
from generic non-endothelial stromal cells, die after a few days.
Now, employing our endothelial stem cell co-cultures, we can
propagate bona fide adult stem cells in the absence of external
factors and serum beyond 21 days with an expansion index of more
If this vascular-based stem cell expansion strategy continues to
be validated, physicians could use any source of hematopoietic
(blood-producing) stem cells, propagate them exponentially, and bank
the cells for transplantation into patients.
In a true first, the study demonstrates how this novel vascular
cell platform or "vascular niche" can self-renew adult hematopoietic
stem cells for weeks, both in vitro and in vivo, by co-culturing
them on a bed of endothelial cells. The researchers chose
endothelial cells because they are in close contact with blood stem
cells, and previous work from Dr Rafii's lab had demonstrated that
endothelial cells produce novel stem-cell-active growth factors.
However, maintenance of the endothelial cells is cumbersome and
if they are not "fed" specific substances, such as growth factors
known as "angiogenic factors," they immediately die. To get around
this problem, the researchers genetically engineered the endothelial
cells to stay in a long-term survival state by inserting a recently
discovered gene cloned from adenoviruses, which does not promote
oncogenic transformation of the human cells.
This earlier discovery, using a single gene to put endothelial
cells into a long-lasting "suspended animation" state without
harming their ability to produce blood vessels, was also discovered
in Dr Rafii's lab and published in the journal Proceedings of
National Academy Sciences in 2008.
Endothelial cells could generate stem cells and their
In this study, the researchers also discovered that endothelial
cells not only could expand stem cells, but also instruct stem cells
to generate mature differentiated progeny that could form immune
cells, platelets, and red and white blood cells, all of which
constitute functioning blood.
"We are the first group to demonstrate that endothelial cells
elaborate a repertoire of stem-cell-active growth factors that not
only stimulate stem cell expansion but also orchestrate
differentiation of these stem cells into their mature progeny," says
Dr. Jason Butler, a senior investigator at Weill Cornell Medical
College and first author of the study.
"For example, we have found that expression of specific
stem-cell-active factors, namely Notch-ligands, by the endothelial
cells lining the wall of working blood vessels promote proliferation
of the blood-forming stem cells. Inhibition of these specific
factors on the endothelial cells resulted in the failure of the
regeneration of the blood-forming stem cells. These findings suggest
that endothelial cells directly, through expression of
stem-cell-active cytokines, promote stem cell reconstitution."
Further describing this innovative concept, in a high-impact
article published in the January 2010 issue of Nature Reviews
Cancer, Drs. Rafii and Butler, and Dr. Hideki Kobayashi, who is also
a co-author of the current study, have elaborated on specific
endothelial cell-produced growth factors that promote the growth of
tumor cells besides stem cells.
Development of the vascular-cell technology that supports
long-lasting growth of stem cells will also allow scientists to
generate abundant sources of functional and malignant stem cells for
genetic and basic studies.
This study has also resolved a long-standing controversy in which
several groups had claimed that bone-forming cells (osteoblasts)
exclusively support the expansion of blood-forming stem cells.
"However, using a highly sophisticated molecular imaging
approach, we show that regenerating blood-forming stem cells in the
bone marrow are in intimate contact with the blood vessels,
indicating that endothelial cells are the predominant regulator of
stem cell repopulation in the adult bone marrow," states Dr. Daniel
Nolan, a senior scientist in Dr. Rafii's lab and a co-author of the
One other important concern addressed in this study was whether
forced expansion of the stem cells over a long period of time would
induce cancerous mutations in the stem cells. However, the authors
of this study show that, even after one year, there was no
indication of tumor formation, such as leukemias, when the expanded
stem cells were transplanted back into mice. This suggests that the
endothelial cells provide a milieu that proliferates stem cells
without creating cancer risk.
The current breakthrough represents the culmination of many years
of work by Dr. Rafii and his lab, including their research in
converting adult mouse spermatogonial stem cells to endothelial
cells (Nature, September 2007) and in deriving stable, copious
endothelial cells from human embryonic stem cells (Nature
Biotechnology, Jan. 17, 2010).
The ability to generate many stable endothelial cells from human
embryonic stem cells leads to new research opportunities, according
to Dr Zev Rosenwaks, who is a co-author in this study and director
and physician-in-chief of the Ronald O Perelman and Claudia Cohen
Center for Reproductive Medicine, as well as the director of the
Tri-Institutional Stem Cell Initiative Derivation Unit at Weill
Cornell Medical College.
Dr Rosenwaks says, "Generation of endothelial cells derived from
diseased embryonic stem cells that are being propagated in our
Derivation Unit will open up new avenues of research to molecularly
eavesdrop on the communication between vascular cells and stem
cells. This innovative line of investigation — to determine how
normal and abnormal human vascular cells induce the formation of
organs during development of embryos and how dysfunction of
endothelial cells results in developmental defects — will lay the
foundation for novel platforms for therapeutic organ regeneration."
Dr Rafii sees even more opportunities. "Identification of as yet
unrecognized growth factors produced by human embryonic cell-derived
endothelium and adult endothelial cells that support stem cell
expansion and differentiation will establish a new arena in stem
cell biology. We will be able to selectively activate endothelial
cells not only to induce organ regeneration, but also to inhibit
specifically the production of endothelial cell-derived factors in
order to block the growth of tumors.
"Our findings are the first steps toward such goals and they
highlight the potential of vascular cells for generating sufficient
stem cells for therapeutic organ regeneration, tumor targeting, and
gene therapy applications," concludes Dr. Rafii.