Growing New Organs

Rcjohnsen rcjohnsen at aol.com
Sat Feb 26 16:42:33 EST 2000


GROWING NEW ORGANS

Researchers have taken the first steps toward creating semisynthetic, living
organs that can be used as human replacement parts
by David J. Mooney and Antonios G. Mikos 
http://www.sciam.com/1999/0499issue/0499mooney.html

SUBTOPICS: 
A Pinch of Protein
A Dash of Cells
Recipes for the Futur
                               
ILLUSTRATIONS:
Polymer Scaffolding
Vessel Ingrowth via Growth Factors
Vessel Outgrowth via Cell Implants
Plasmids
FURTHER READING


RELATED LINKS 
Illustration: Grant Jerding

   Human body may be more than a sum of parts, but replacing failing parts
should extend and improve life. 
   Every day thousands of people of all ages are admitted to hospitals because
of the malfunction of some vital organ. Because of a dearth of transplantable
organs, many of these people will die. In perhaps the most dramatic example,
the American Heart Association reports only 2,300 of the 40,000 Americans who
needed a new heart in 1997 got one. Lifesaving livers and kidneys likewise are
scarce, as is skin for burn victims and others with wounds that fail to heal.
It can sometimes be easier to repair a damaged automobile than the vehicle's
driver because the former may be rebuilt using spare parts, a luxury that human
beings simply have not enjoyed.
An exciting new strategy, however, is poised to revolutionize the treatment of
patients who need new vital structures: the creation of man-made tissues or
organs, known as neo-organs. In one scenario, a tissue engineer injects or
places a given molecule, such as a growth factor, into a wound or an organ that
requires regeneration. These molecules cause the patient's own cells to migrate
into the wound site, turn into the right type of cell and regenerate the
tissue. In the second, and more ambitious, procedure, the patient receives
cells--either his or her own or those of a donor--that have been harvested
previously and incorporated into three-dimensional scaffolds of biodegradable
polymers, such as those used to make dissolvable sutures. The entire structure
of cells and scaffolding is transplanted into the wound site, where the cells
replicate, reorganize and form new tissue. At the same time, the artificial
polymers break down, leaving only a completely natural final product in the
body--a neo-organ. The creation of neo-organs applies the basic knowledge
gained in biology over the past few decades to the problems of tissue and organ
reconstruction, just as advances in materials science make possible entirely
new types of architectural design.

... grows to fill a space between two bone segments. A dog leg bone with a
missing section is held in place with braces (a). A polymer scaffold primed
with bone growth-promoting proteins (b) fills in the gap. The scaffold is
slowly infiltrated by new bone (c) and ultimately gets completely replaced (d).
The cells (e) have their own blood supply (red and blue vessels). The leg bone
has healed 
   Science-fiction fans are often confronted with the concept of tissue
engineering. Various television programs and movies have pictured individual
organs or whole people (or aliens) growing from a few isolated cells in a vat
of some powerful nutrient. Tissue engineering does not yet rival these
fictional presentations, but a glimpse of the future has already arrived. The
creation of tissue for medical use is already a fact, to a limited extent, in
hospitals across the U.S. These groundbreaking applications involve fabricated
skin, cartilage, bone, ligament and tendon and make musings of "off-the-shelf"
whole organs seem less than far-fetched.
   Indeed, evidence abounds that it is at least theoretically possible to
engineer large, complex organs such as livers, kidneys, breasts, bladders and
intestines, all of which include many different kinds of cells. The proof can
be found in any expectant mother's womb, where a small group of
undifferentiated cells finds the way to develop into a complex individual with
multiple organs and tissues with vastly different properties and functions.
Barring any unforeseen impediments, teasing out the details of the process by
which a liver becomes a liver, or a lung a lung, will eventually allow
researchers to replicate that process.
A Pinch of Protein
   Cells behave in predictable ways when exposed to particular biochemical
factors. In the simpler technique for growing new tissue, the engineer exposes
a wound or damaged organ to factors that act as proponents of healing or
regeneration. This concept is based on two key observations, in bones and in
blood vessels. 
   In 1965 Marshall R. Urist of the University of California at Los Angeles
demonstrated that new, bony tissue would form in animals that received implants
of powdered bone. His observation led to the isolation of the specific proteins
(the bone morphogenetic proteins, or BMPs) responsible for this activity and
the determination of the DNA sequences of the relevant genes. A number of
companies subsequently began to produce large quantities of recombinant human
BMPs; the genes coding for BMPs were inserted into mammalian cell lines that
then produced the proteins.

Vascularization of new tissue


VESSEL INGROWTH VIA FACTORS




... can be accomplished in two ways. Vessels from the surrounding tissue can be
induced to infiltrate the tissue. Such vessel growth is promoted by including
growth factors (blue dots) in the polymer scaffold of the insert (a). These
factors diffuse into the local environment, where they encourage existing blood
vessels to grow into the polymer (b). Ultimately, cells growing in from both
sides knit together to form a continuous vessel (c) . .
   Various clinical trials are under way to test the ability of these bone
growth promoters to regenerate bony tissue. Applications of this approach that
are currently being tested include healing acute bone fractures caused by
accidents and boosting the regeneration of diseased periodontal tissues.
Creative BioMolecules in Hopkinton, Mass., recently completed clinical trials
showing that BMP-7 does indeed help heal severe bone fractures. This trial
followed 122 patients with leg fractures in which the sections failed to rejoin
after nine months. Patients whose healing was encouraged by BMP-7 did as well
as those who received a surgical graft of bone harvested from another part of
their body.
   A critical challenge in engineering neo-organs is feeding every cell.
Tissues more than a few millimeters thick require blood vessels to grow into
them and supply nutrients. Fortunately, investigations by Judah Folkman have
shown that cells already in the body can be coaxed into producing new blood
vessels. Folkman, a cancer researcher at Harvard Medical School's Children's
Hospital, recognized this possibility almost three decades ago in studies
aimed, ironically, at the prevention of cellular growth in the form of
cancerous tumors.

Vessels 


VESSEL OUTGROWTH VIA CELL IMPLANTS





... may also grow from within a polymer scaffold if that scaffold is seeded (d)
with endothelial cells (purple). The cells will proliferate within the polymer
and grow outward toward the natural tissue (e). These new vessels combine with
existing blood vessels (red) to create a continuous vessel (f) 

   Folkman perceived that developing tumors need to grow their own blood
vessels to supply themselves with nutrients. In 1972 he proposed that specific
molecules could be used to inhibit such vessel growth, or angiogenesis, and
perhaps starve tumors. (This avenue of attack against cancer became a major
news story in 1998.) Realizing that other molecules would undoubtedly abet
angiogenesis, he and others have subsequently identified a number of factors in
each category.
   That work is now being exploited by tissue engineers. Many
angiogenesis-stimulating molecules are commercially available in recombinant
form, and animal studies have shown that such molecules promote the growth of
new blood vessels that bypass blockages in, for example, the coronary artery.
Small-scale trials are also under way to test this approach in the treatment of
similar conditions in human subjects.
   Scientists must surmount a few obstacles, however, before drugs that promote
tissue and organ formation become commonplace. To date, only the factors
responsible for bone and blood vessel growth have been characterized. To
regenerate other organs, such as a liver, for example, the specific molecules
for their development must be identified and produced reliably.
   An additional, practical issue is how best to administer the substances that
would shape organ regeneration. Researchers must answer these questions: What
specific concentrations of the molecules are needed for the desired effect? How
long should the cells be exposed? How long will the factors be active in the
body? Certainly multiple factors will be needed for complex organs, but when
exactly in the development of the organ does one factor need to replace
another? Controlled drug-delivery technology such as transdermal patches
developed by the pharmaceutical industry will surely aid efforts to resolve
these concerns.
   In particular, injectable polymers may facilitate the delivery of bioactive
molecules where they are needed, with minimal surgical intervention. Michael J.
Yaszemski of the Mayo Clinic, Alan W. Yasko of the M. D. Anderson Cancer Center
in Houston and one of us (Mikos) are developing new injectable biodegradable
polymers for orthopedic applications. The polymers are moldable, so they can
fill irregularly shaped defects, and they harden in 10 to 15 minutes to provide
the reconstructed skeletal region with mechanical properties similar to those
of the bone they replace. These polymers subsequently degrade in a controlled
fashion, over a period of weeks to months, and newly grown bone fills the site.
   We have also been studying the potential of injectable, biodegradable
hydrogels--gelatinlike, water-filled polymers--for treating dental defects,
such as poor bonding between teeth and the underlying bone, through guided bone
regeneration. The hydrogels incorporate molecules that both modulate cellular
function and induce bone formation; they provide a scaffold on which new bone
can grow, and they minimize the formation of scar tissue within the regenerated
region.
   An intriguing variation of more conventional drug delivery has been
pioneered by Jeffrey F. Bonadio, Steven A. Goldstein and their co-workers at
the University of Michigan. (Bonadio is now at Selective Genetics in San
Diego.) Their approach combines the concepts of gene therapy and tissue
engineering. Instead of administering growth factors directly, they insert
genes that encode those molecules. The genes are part of a plasmid, a circular
piece of DNA constructed for this purpose. The surrounding cells take up the
DNA and treat it as their own. They turn into tiny factories, churning out the
factors coded for by the plasmid. Because the inserted DNA is free-floating,
rather than incorporated into the cells' own DNA, it eventually degrades and
the product ceases to be synthesized. Plasmid inserts have successfully
promoted bone regrowth in animals; the duration of their effects is still being
investigated.

Plasmids, circlets of DNA (yellow)

... find their way from a polymer scaffold to a nearby cell in the body, where
they serve as the blueprints for making desirable proteins. Adding the proteins
themselves would be less effective because the proteins tend to degrade much
faster than the plasmids do. Researchers attempting to use growth promoters in
tissue engineering may thus find it more reliable to insert plasmids than the
proteins they encode..

   One of us (Mooney), along with Lonnie D. Shea and our other aforementioned
Michigan colleagues, recently demonstrated with animals that three-dimensional
biodegradable polymers spiked with plasmids will release that DNA over extended
periods and simultaneously serve as a scaffold for new tissue formation. The
DNA finds its way into adjacent cells as they migrate into the polymer
scaffold. The cells then express the desired proteins. This technique makes it
possible to control tissue formation more precisely; physicians might one day
be able to manage the dose and time course of molecule production by the cells
that take up the DNA and deliver multiple genes at various times to promote
tissue formation in stages.
A Dash of Cells
   Promoting tissue and organ development via growth factors is obviously a
considerable step forward. But it pales in comparison to the ultimate goal of
the tissue engineer: the creation from scratch of whole neo-organs. Science
fiction's conception of prefabricated "spare parts" is slowly taking shape in
the efforts to transplant cells directly to the body that will then develop
into the proper bodily component. The best way to sprout organs and tissues is
still to rely on the body's own biochemical wisdom; the appropriate cells are
transferred, in a three-dimensional matrix, to the desired site, and growth
unfolds within the person or organism rather than in an external, artificial
environment. This approach, pioneered by Ioannis V. Yannas, Eugene Bell and
Robert S. Langer of the Massachusetts Institute of Technology, Joseph P.
Vacanti of Harvard Medical School and others in the 1970s and 1980s, is now
actually in use in some patients, notably those with skin wounds or cartilage
damage.
   The usual procedure entails the multiplication of isolated cells in culture.
These cells are then used to seed a matrix, typically one consisting of
synthetic polymers or collagen, the protein that forms the natural support
scaffolding of most tissues. In addition to merely delivering the cells, the
matrix both creates and maintains a space for the formation of the tissue and
guides its structural development. Once the developmental rules for a given
organ or tissue are fully known, any of those entities could theoretically be
grown from a small sample of starter cells. (A sufficient understanding of the
developmental pathways should eventually allow the transfer of this procedure
from the body to the laboratory, making true off-the-shelf organs possible. A
surgeon could implant these immediately in an emergency situation--an appealing
notion, because failing organs can quickly lead to death--instead of waiting
weeks or months to grow a new organ in the laboratory or to use growth factors
to induce the patient's own body to grow the tissues.)
   In the case of skin, the future is here. The U.S. Food and Drug
Administration has already approved a living skin product--and others are now
in the regulatory pipeline. The need for skin is acute: every year 600,000
Americans suffer from diabetic ulcers, which are particularly difficult to
heal; another 600,000 have skin removed to treat skin cancer; and between
10,000 and 15,000 undergo skin grafts to treat severe burns.
   The next tissue to be widely used in humans will most likely be cartilage
for orthopedic, craniofacial and urological applications. Currently available
cartilage is insufficient for the half a million operations annually in the
U.S. that repair damaged joints and for the additional 28,000 face and head
reconstructive surgeries. Cartilage, which has low nutrient needs, does not
require growth of new blood vessels--an advantage for its straightforward
development as an engineered tissue.
   Genzyme Tissue Repair in Cambridge, Mass., has received FDA approval to
engineer tissues derived from a patient's own cells for the repair of traumatic
knee-cartilage damage. Its procedure involves growing the patient's cells in
the lab, harvested from the same knee under repair when possible, and then
implanting those cells into the injury. Depending on the patient and the extent
of the defect, full regeneration takes between 12 and 18 months. In animal
studies, Charles A. Vacanti of the University of Massachusetts Medical School
in Worcester, his brother, Joseph Vacanti, Langer and their colleagues have
shown that new cartilage can be grown in the shapes of ears, noses and other
recognizable forms.
   The relative ease of growing cartilage has led Anthony J. Atala of Harvard
Medical School's Children's Hospital to develop a novel approach for treating
urological disorders such as incontinence. Reprogenesis in Cambridge, Mass.,
which supports Atala's research, is testing whether cartilage cells can be
removed from patients, multiplied in the laboratory and used to add bulk to the
urethra or ureters to alleviate urinary incontinence in adults and bladder
reflux in children. These conditions are often caused by a lack of muscle tone
that allows urine to flow forward unexpectedly or, in the childhood syndrome,
to back up. Currently patients with severe incontinence or bladder reflux may
undergo various procedures, including complex surgery. Adults sometimes receive
collagen that provides the same bulk as the cartilage implant, but collagen
eventually degrades. The new approach involves minimally invasive surgery to
deliver the cells and grow the new tissue.
   Walter D. Holder, Jr., and Craig R. Halberstadt of Carolinas Medical Center
in Charlotte, N.C., and one of us (Mooney) have begun to apply such general
tissue-engineering concepts to a major women's health issue. We are attempting
to use tissue from the legs or buttocks to grow new breast tissue, to replace
that removed in mastectomies or lumpectomies. We propose to take a biopsy of
the patient's tissue, isolate cells from this biopsy and multiply these cells
outside the body. The woman's own cells would then be returned to her in a
biodegradable polymer matrix. Back in the body, cell growth and the
deterioration of the matrix would lead to the formation of completely new,
natural tissue. This process would create only a soft-tissue mass, not the
complex system of numerous cell types that makes up a true breast.
Nevertheless, it could provide an alternative to current breast prostheses or
implants. 
   Optimism for the growth of large neo-organs of one or more cell types has
been fueled by success in several animal models of human diseases. Mikos
recently demonstrated that new bone tissue can be grown by transplanting cells
taken from bone marrow and growing them on biodegradable polymers.
Transplantation of cells to skeletal defects makes it possible for cells to
produce factors locally, thus offering a new means of delivery for
growth-promoting drugs.
Recipes for the Future
In any system, size imposes new demands. As previously noted, tissues of any
substantial size need a blood supply. To address that requirement, engineers
may need to transplant the right cell types together with drugs that spur
angiogenesis. Molecules that promote blood vessel growth could be included in
the polymers used as transplant scaffolds. Alternatively, we and others have
proposed that it may be possible to create a blood vessel network within an
engineered organ prior to transplantation by incorporating cells that will
become blood vessels within the scaffold matrix. Such engineered blood vessels
would then need only to connect to surrounding vessels for the engineered
tissue to develop a blood supply.
   In collaboration with Peter J. Polverini of Michigan, Mooney has shown that
transplanted blood vessel cells will indeed form such connections and that the
new vessels are a blend of both implanted and host cells. But this technique
might not work when transplanting engineered tissue into a site where blood
vessels have been damaged by cancer therapy or trauma. In such situations, it
may be necessary to propagate the tissue first at another site in the body
where blood vessels can more readily grow into the new structure. Mikos
collaborates with Michael J. Miller of the M. D. Anderson Cancer Center to
fabricate vascularized bone for reconstructive surgery using this approach. A
jawbone, for instance, could be grown connected to a well-vascularized hipbone
for an oral cancer patient who has received radiation treatments around the
mouth that damaged the blood supply to the jawbone.
   On another front, engineered tissues typically use biomaterials, such as
collagen, that are available from nature or that can be adapted from other
biomedical uses. We and others, however, are developing new biodegradable,
polymeric materials specific to this task. These materials may accurately
determine the size and shape of an engineered tissue, precisely control the
function of cells in contact with the material and degrade at rates that
optimize tissue formation.
   Structural tissues, such as skin, bone and cartilage, will most likely
continue to dominate the first wave of success stories, thanks to their
relative simplicity. The holy grail of tissue engineering, of course, remains
complete internal organs. The liver, for example, performs many chemical
reactions critical to life, and more than 30,000 people die every year because
of liver failure. It has been recognized since at least the time of the ancient
Greek legend of Prometheus that the liver has the unique potential to
regenerate partially after injury, and tissue engineers are now trying to
exploit this property of liver cells.
   A number of investigators, including Joseph Vacanti and Achilles A.
Demetriou of Cedars-Sinai Medical Center in Los Angeles, have demonstrated that
new liverlike tissues can be created in animals from transplanted liver cells.
We have developed new biomaterials for growing liverlike tissues and shown that
delivering drugs to transplanted liver cells can increase their growth. The new
tissues grown in all these studies can replace single chemical functions of the
liver in animals, but the entire function of the organ has not yet been
replicated.
   H. David Humes of Michigan and Atala are using kidney cells to make
neo-organs that possess the filtering capability of the kidney. In addition,
recent animal studies by Joseph Vacanti's group have demonstrated that
intestine can be grown--within the abdominal cavity--and then spliced into
existing intestinal tissue. Human versions of these neointestines could be a
boon to patients suffering from short-bowel syndrome, a condition caused by
birth defects or trauma. This syndrome affects overall physical development
because of digestion problems and subsequent insufficient nutrient intake. The
only available treatment is an intestinal transplant, although few patients
actually get one, again because of the extreme shortage of donated organs.
Recently Atala has also demonstrated in animals that a complete bladder can be
formed with this approach and used to replace the native bladder.
   Even the heart is a target for regrowth. A group of scientists headed by
Michael V. Sefton at the University of Toronto recently began an ambitious
project to grow new hearts for the multitude of people who die from heart
failure every year. It will very likely take scientists 10 to 20 years to learn
how to grow an entire heart, but tissues such as heart valves and blood vessels
may be available sooner. Indeed, several companies, including Advanced Tissue
Sciences in La Jolla, Calif., and Organogenesis in Canton, Mass., are
attempting to develop commercial processes for growing these tissues.
   Prediction, especially in medicine, is fraught with peril. A safe way to
prophesy the future of tissue engineering, however, may be to weigh how
surprised workers in the field would be after being told of a particular
hypothetical advance. Tell us that completely functional skin constructs will
be available for most medical uses within five years, and we would consider
that reasonable. Inform us that fully functional, implantable livers will be
here in five years, and we would be quite incredulous. But tell us that this
same liver will be here in, say, 30 years, and we might nod our heads in
sanguine acceptance--it sounds possible. Ten millennia ago the development of
agriculture freed humanity from a reliance on whatever sustenance nature was
kind enough to provide. The development of tissue engineering should provide an
analogous freedom from the limitations of the human body.

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Further Reading
TISSUE ENGINEERING AND THE HUMAN BODY SHOP: DESIGNING "BIOARTIFICIAL ORGANS."
Carol Ezzell in Journal of NIH Research, Vol. 7, No. 7, pages 49-53; July 1995.
PRINCIPLES OF TISSUE ENGINEERING. Edited by R. P. Lanza, R. Langer and W. L.
Chick. R. G. Landes, 1997.
FRONTIERS IN TISSUE ENGINEERING. Edited by Charles W. Patrick, Jr., Antonios G.
Mikos and Larry V. McIntire. Pergamon Press, 1998.

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Related Links:
Doctors herald grow-your-own organs BBC News
Engineered neo-organs for bladder replacement-- study by Frank Oberpenning, Jun
Meng, James J. Yoo and Anthony Atala, Boston, Mass (presented by Dr.
Oberpenning).
The Pittsburgh Tissue Engineering Initiative: a resource for biomedical
technology of tissue engineering.
TissueInformatics Inc.: specializing in digital information and imaging for
tissue engineering. 

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The Authors:
DAVID J. MOONEY and ANTONIOS G. MIKOS have collaborated for eight years. Mooney
has been on the faculty at the University of Michigan since 1994, where he is
associate professor of biologic and materials sciences and of chemical
engineering. He studies how cells respond to external biochemical and
mechanical signals and designs and synthesizes polymer scaffolds used in tissue
engineering. Mikos is associate professor of bioengineering and of chemical
engineering at Rice University. Mikos's research focuses on the synthesis,
processing and evaluation of new biomaterials for tissue engineering, including
those useful for scaffolds, and on nonviral vectors for gene therapy.

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