A CONVERSATION WITH | MARIE FILBIN
Defying Irreversibility in Spinal Cord Injuries
By CLAUDIA DREIFUS
Published: March 16, 2004
he first thought that Dr. Marie T. Filbin wants to convey is that it is possible to teach at a municipal college and have a great research career.
"People are always putting down the City University because it is not Harvard or Rockefeller," said Dr. Filbin, 48, a professor of biological sciences at Hunter College of the City University of New York. "But Hunter is a great place for a researcher. My students are wonderful."
A scientist who praises an employer is rare enough. But then Marie Filbin is an unusual scientist. Her specialty is practical: she studies why injured nerve cells do not regenerate themselves — a factor crucial to understanding the mysteries of paralysis.
When not at her bench, she delivers lectures — really progress reports on her research — at nursing homes. And when she speaks to paralysis patients, she says, "They put me on the spot."
"I like giving them confidence that people are trying," Dr. Filbin said. "It makes me try harder."
Q. A maxim of biology is that the nerves of mammals, once injured or damaged, can never grow back. That's why some spinal cord injuries have long been thought to be irreversible. Why do you believe that your research might change that?
A. That's what used to be the dogma. But we know a lot more than we ever did about nerve cells and how they work. Since the 1980's, the thinking has been that the problem isn't the nerve cells, but something in their environment that inhibits injured nerves from regenerating their axons — those long armlike parts of nerve cells. Scientists have learned in recent years that the myelin, the insulation that protects nerves when they are intact, may be part of the problem. Damaged myelin actually has chemical inhibitors that can stop regeneration.
At my laboratory, we've been looking at a protein, the myelin-associated glycoprotein — MAG for short — which, under certain circumstances, can either stop or promote axonal growth. This protein may be one of the keys to changing things.
Q. How did you find the protein, and why is it important?
A. In the early 1980's, I was doing postdoctoral research on myelin at Johns Hopkins. I was studying how it works, and why it falls apart in diseases like multiple sclerosis. That led to work in the 1990's here at Hunter College where we made the discovery that MAG inhibited axon regeneration. We found when we grew nerve cells in the presence of this protein, axons would not grow. To that date, there had only been one other protein found with this property. Now, we think there are at least three.
So we looked at how the MAG tells nerves not to grow. We asked how we can overcome this chemical inhibition. And with that question came the second important discovery made in my lab. We found that if you change the chemistry of the nerve cell, elevate a molecule — the cyclic AMP, or the cAMP — nerve cell regrowth is no longer inhibited. They grow quite lovely, in fact.
Q. Is this chemical reaction within the injured cell the only factor preventing regeneration?
A. No. And we are looking at others. The scar that forms on the nerve after an injury is also part of the problem. It is believed that a scar takes some time to form. So if you can get the nerves to grow before the scar forms, this also might be helpful. Most researchers think that an eventual therapy will involve blocking the myelin inhibitors and preventing a scar from forming — or dispersing it, once it has formed.
Q. Does this mean that you're looking for some kind of drug or chemical that could stop the injured myelin from blocking regeneration?
A. Right. And there are a few possibilities that already look promising. There are already drugs on the market that were developed for other purposes, but can cause cAMP to rise. These may eventually be used for spinal cord injury.
Q. Do you use human embryonic stem cells in your research?
A. Not right now. But at some point, I might. I am collaborating with Dr. Thomas Jessell at Columbia University on a related project. Together, we are trying to turn rodent embryonic stem cells into neurons and then transplant them into a rat or mouse. The idea is to see if this can replace lost or damaged neurons. If our rodent trials look promising, of course we'll want to take it to the next level, which means experiments with human embryonic stem cells.
I can't do that. At Hunter College, we get federal funds. The same is true for Dr. Jessell at Columbia. As you know, in August 2001 President Bush issued an order barring the use of federal funding to study new embryonic stem cell lines.
Q. When the president issued that order, he exempted some 60 already existing stem cell lines from the prohibition. Can't you use them?
A. I'm afraid not. The available stem cell lines can never be transplanted into a human because they have been exposed to mice cells. For what we hope to do, we need to make new human embryonic stem cells — and then transplant them into an injured person and see if, under the right conditions, new neurons will grow.
In theory, I could take my research to one of the few private venues in the United States that is doing stem cell investigations. However, I am committed to Hunter and to my students here.
Q. There are American scientists doing human stem cell work. How do they manage it?
A. At Stanford they've put up a whole separate building with private money. The reason for the separate building is to make sure that nothing that gets federal funding commingles with the stem cell research. You can't use the same bench, the same beakers, and the same glassware.
There's nothing like that near to me, though; if there was, I could seek collaboration there and take my work to the next level.
Q. Christopher Reeve has regained some slight function in his finger because of an aggressive physical therapy regime. Have you been surprised by his progress?
A. Yes and no. With spinal cord injury, there can be a certain degree of spontaneous recovery for up to two years after the trauma. There are a few reports of paralyzed people who walked again after six months. We don't know why. But it's probably that the nerves were not severed but only crushed, and the nerves woke up again.
Q. Did you grow up wanting to be a researcher of nerve diseases?
A. The main thing was that I wanted to get out of Northern Ireland, where I grew up. The town of Lurgan. There were a lot of shootings and bombs there during the years I was a teenager. I remember getting caught in a gun battle in the middle of town and being dragged to safety by an I.R.A. man. I just wanted to get out. I went to convent school with wonderful science teachers and they showed me the fun of science. Science was my escape.
I was particularly taken by the idea of the nervous system. I remember being fascinated by the image of the synapse, where two nerve cells come in contact with each other. Is this how one thinks, this place where there is chemical communication between the two cells, I wondered? My best friend at school, Nuala Mooney, was similarly intrigued, and we swore we'd get away by studying science in some safe place elsewhere. And that's what we eventually did. Together, we went to Bath University in England. Nuala today is head of her own lab in Paris.
LASER PHOTOSTIMULATION: An old mystery metamorphosing into a new millennium marvel
The notion that light, in the visible and near infrared ranges, can produce photochemical and photobiological changes that ameliorate pain and/or promote tissue repair was first observed in the late 1960s. At that time the prevailing notion was that lasers were uniquely photodestructive, prompting attempts to develop powerful lasers that my yield military superiority. Thus, the mood was not right and neither were medical minds ready to accept the idea that a tool that can cut, vaporize, and otherwise destroy tissue could be used for beneficial purposes. Skepticism about the therapeutic value of laser therapy continues to linger, but much has changed in the last four decades. The cold war is over, the arms race has waned, and works supporting the medical benefits of laser therapy have been on the rise.
Since this special issue of the journal presents some of the major works and accomplishments in the field, it is perhaps appropriate to use Dr. Toshio Ohshiro's summary of his 27-year experience in the field as a backdrop for a meaningful discussion of our achievements to date. About 20 years ago, Ohshiro's group demonstrated that 830 nm portable GaAlAs diode laser system attenuates pain when used in the contact mode, within dose parameters that include an average power of 60 mW. The works of Dr. Kevin Moore of the U.K., and Professor Osamu Kemmotsu of Japan, support the findings of Ohshiro et al., given that both have shown, in a series of double-blind studies, that low power laser phototherapy ameliorates pain in a wide range of conditions, particularly in patients with post-hepertic neuralgia.
Similar successes have been demonstrated in the treatment of other conditions—hypertrophic scars and keloids, failing skin grafts and flaps, hyper- and hypopigmentation, vitiligo, atopic dermatitis, atrophic skin, psoriasis vulgaris, Buerger's disease, and strawberry hemangioma in infants. The effects of laser therapy on some of these conditions have been studied by Drs. Junichiro Kubota of Japan and L. Schindl of Austria. Kubota's works on skin grafts reveal accelerated angiogenesis, better reperfusion and flap survival following laser therapy. Schindl's works elucidate the effectiveness of laser therapy in relieving the pain associated with Buerger's disease, i.e., thromboangiitis obliterans, showing that this beneficial effect is sustained for at least nine months. This carryover effect is partially explained by the work of Ohshiro et al., which indicates that laser therapy stimulates neoangiogenesis in persons with Buerger's disease.
The mechanisms involved in laser amelioration of pain continue to emerge. When pain is associated with inflammation, the evidence suggests that the mediators of inflammation are influenced by light in such a way that the overall process of inflammation proceeds faster. Works by Dr. Mary Dyson and her group have shown that mast cells are induced to release granules containing heparin and other chemical mediators of inflammation. Her group and others have also shown that certain white blood cells, e.g., macrophages are activated by light and caused to release a host of chemicals—including growth factors—that further promote the process of inflammation. Drs. G. Barberis, Vilma Campana, Fernando Soriano and their associates of Cordoba, Argentina, have demonstrated the involvement of other factors in laser amelioration of pain. They have shown that 632.8 nm light reduces the pain associated with rheumatoid arthritis by modulating the level of prostaglandin E2 (PGE2). A finding supported by some of the early works of Endre Mester and the recent report published by the husband and wife pair of Drs. Constantine and Laura Ailioaie of Romania.
Another mechanism via which laser therapy can modulate pain is presented in this issue of the journal by Nelson and Friedman of the U.S. Using a 1.7 mW 632.5 nm light source, they irradiated the maxillary nerve intraorally for two minutes and showed that somatosensory trigeminal evoked potentials (STEP) decreased 60% immediately after irradiation, and by as much as 72%, 20 minutes later. Given that STEP provides an objective assessment of pain, the sharp decrease in STEP amplitude observed in this study suggests a direct effect of laser treatment on the nerves that carry the sensation of pain. Their finding is supported by the work of Professor Aldo Brugnera of Săo Paulo, Brazil (also in this issue of the journal), which indicates rapid relieve from the sharp intense pain associated with dentinal hypersensitivity when 4 J cm2 energy fluence of either 780 nm or 830 nm wavelength is used to treat this condition over a relatively short period. Other evidence supporting the idea that light affects nerve conduction can be seen in some of the works of Prof. G. David Baxter and his group of Ireland.
Since Endre Mester first published his pioneering work identifying the therapeutic value of laser therapy in wound healing, gaining a better understanding of how light energy promotes tissue repair has been a primary focus of most researchers. Important advances have been made in the areas of skin repair, muscle repair, nerve repair, tendon repair, cartilage repair, bone repair, and gum and dental tissue repair. It is now known that when healing is impaired, the aforementioned tissues respond positively to appropriate doses of light, especially light wavelengths within the 600 to 1000 nm range. The exact fluence necessary to achieve optimal healing continues to be explored for each tissue, but there is a growing consensus that accelerated healing can be accomplished with doses ranging from 1 to 6 J cm2. As detailed in our work presented in this issue of the journal, the amount of light absorbed by each tissue differs, even when wavelength is kept constant. Therefore, the energy fluence needed to optimize healing will differ among tissues.
While it is difficult to acknowledge every important work published to date, the sustained contributions of the Schindls of Austria, Dr. Farouk Al-Watban of Saudi Arabia, Prof. Leonardo Longo of Italy, Prof. Zlatko Simunovic of Switzerland and his associates, Profs. Rachel Lubart and Uri Oron of Israel, Sandra Giavelli of Italy, S.M. Ghamsari of Japan, and Dr. Ray Lanzafame of the U.S. are noteworthy in the area of skin repair, as are the important works emerging from our laboratory and those of Drs. Harry Whelan of the U.S., and Pamela Houghton of Canada. The labs of Drs. Shimon Rochkind and Juanita Anders of Israel and the U.S. respectively, have made significant contributions revealing the positive effects of specific energy fluences of light on nerve tissue regeneration, including regeneration of the spinal cord, a part of the central nervous system once considered inert to healing.
A substantial amount of evidence has emerged supporting the use of low energy lasers to promote fracture repair since Mario Trelles of Spain published his pioneering work, which revealed the positive effects of laser therapy on bone healing. The contributors in this area include Dr. R. Giardino and coworkers of Italy, Dr. J. Chen of China and Dr. Shimon Rochkind and others detailed in his review article published in this volume. Supporting their findings are reports which demonstrate the effectiveness of laser therapy in promoting bone, oral or dental tissue repair with or without surgery. These include the works of Professor Tony Pinhiero and his group in Brazil, one of which appears in this volume. The cellular studies of Dr. Luciana Almeida Lopes in this area are noteworthy. Tendon and cartilage are two other connective tissues that respond positively to laser therapy, as evidenced by several reports on tendon repair from our group, and the works of Giardino and his associates published in a wide range of journals.
Whereas little was known about light-tissue interaction 40 years ago, the works of Professor Lubart, Professor Passarella of Italy, and others have shown that light energy is absorbed by endogenous chromophores in the mitochondria and membranes of cells. Furthermore, Drs. Tiina Karu and Kira Samoilova of Russia, Patrick Abergel of the U.S., and others have demonstrated that the energy absorbed is used to synthesize DNA, RNA, proteins and various enzymes, resulting in cell proliferation, and tissue repair. Given our understanding of the mechanisms involved in laser tissue repair and pain modulation, it is clear that this form of treatment is emerging from its obscurity of the late 60s to a technological breakthrough that is becoming a mainstay clinical armamentarium of the new millennium. More remains to be done. If the high rate of producing the objective evidence supporting the efficacy of light in treating a wide range of conditions is sustained, its widespread acceptance within the first decade of the new millennium is assured.
Chukuka S. Enwemeka, Ph.D., FACSM
Editor-in-Chief
Email: Enwemeka@kumc.edu
Stops Pulled on Nerve Regeneration
"Stepping on the gas" could lead to better spinal cord treatments
By Gabe Romain, Betterhumans Staff
3/2/2004 • Hits: 265 • Comments: 0
A two-pronged strategy of enhancing growth and halting growth inhibitors greatly improves the regeneration of nerve fibers and could lead to new treatments for spinal cord injury and some eye and brain diseases.
By combining the two strategies, researchers at Children's Hospital Boston and Harvard Medical School in Massachusetts have achieved about three times more regeneration of nerve fibers than previously attained.
"When we combined these two therapies—activating the growth program in nerve cells and overcoming the inhibitory signaling—we got very dramatic regeneration," says Children's Hospital researcher Larry Benowitz.
Broken transmission
Nerve fibers, known as axons, are long cellular projections that conduct electrical impulses away from a neuron's body.
Axons are the primary transmission lines of the nervous system, and as bundles, they make up nerves.
Normally, axons can't regenerate because several proteins in myelin—an electrically insulating fatty layer that surrounds the axons—strongly suppresses cell growth.
Two-pronged strategy
Over the past two years, researchers have developed techniques that disable the inhibitory action of myelin proteins, but such techniques did not make nerves regenerate.
Benowitz and colleagues therefore tried a two-pronged approach to stimulating nerve regrowth, reasoning that blocking inhibition alone would be like trying to drive a car only by taking a foot off the brake.
"Our idea was to step on the gas—to activate the growth state at the same time," says Benowitz. "Knocking out inhibitory molecules alone is not enough, because the nerve cells themselves are still in a sluggish state."
Stimulating growth
To stimulate optic nerve regrowth in rats, the researchers first had to damage their optic nerves, which stimulated immune cells to travel to the site and release growth factors that repaired the damage.
As Benowitz and colleague had found previously, these growth factors activate genes in the retinal nerve cells, causing new axons to grow in the optic nerve.
To enhance this growth, the researchers used a modified virus to deliver into retinal cells a gene designed to turn off the proteins that are programmed to stop regrowth.
Fine tuning
Although the amount of axon regeneration wasn't enough to restore sight, it was about triple that achieved by stimulating growth factors alone, says Benowitz, who, along with his colleagues, will continue studying the optic nerve.
"We have to fine-tune the system, and we have some ideas of how to do it," says Benowitz.
One big challenge for restoring sight is getting nerve fibers from the eye to hook up to the correct centers in the brain so that images aren't scrambled.
"It's a mapping problem," says Benowitz. "We have to retain the proper organization of fiber projections to the brain."
The research is reported in the Journal of Neuroscience (read abstract).
http://www.healthcentral.com/news/NewsFullText.cfm?id=517517
Stem Cell Source Found in Human Brain
The discovery could mark a step toward repairing damage.
THURSDAY, Feb. 19 (HealthDayNews) -- Researchers have discovered a cell-producing region in the human brain that resembles one found in rodent brains -- but with a basic difference that has profound implications for research aimed at repairing damage by growing new cells.
In rodents, a region called the subventricular zone (SVZ), roughly in the middle of the brain, constantly produces cells that migrate to the olfactory bulb, which governs smell. The rodent SVZ produces stem cells, which have the ability to be transformed into many different kinds of cells. Most end up as nerve cells.
Until now, no such cell-producing region has been identified in the human brain. But researchers at the University of California at San Francisco, studying tissue samples taken from patients undergoing brain surgery and post-mortem brain donors, report in the Feb. 19 issue of the journal Nature that they have found one.
It is a ribbon of astrocytes, which make up the great majority of brain cells, the report says. It is also something that has not been seen in any animal brain and that has its own unusual characteristics.
"Unexpectedly, we find no evidence of chains of migrating neuroblasts [young nerve cells] in the SVZ or in the pathway to the olfactory bulb," the researchers write. "Our work identifies SVZ astrocytes as neural stem cells in a niche of unique organization in the human brain."
So what is happening to those cells? One important fact is that something is preventing them from becoming nerve cells, as happens in the rodent brain, says Dr. Pasko Rakic, professor of neuroscience at Yale University School of Medicine and author of an accompanying editorial.
The study shows that the failure of human brains to repair damage is not due to a lack of stem cells, Rakic says. "In humans, there is the same potential pool of cells, but they do not migrate as they do in the mouse," he says. "It is important to see what prevents it."
The human nervous system differs from that of animals in important ways, he notes, using the salamander as an example. "You can cut the spinal cord of salamanders and they grow back," Rakic says. "In humans, this cannot be done."
One obvious difference between humans and salamanders -- and mice -- is that human brains are much more highly organized, he says. The result is a complex structure built to resist changes, such as the introduction of the new cells produced in the SVZ, Rakic says.
But the whole idea of stem cells research is to implant new cells into damaged areas, where they can replace dead nerve cells. Now the challenge is "not just how you implant the cells, but how they make the right connections," he says.
Successful implantation remains a challenge, one that this discovery may help overcome, says Dr. Nader Sanai, a neuroscientist at UCSF and lead author of the journal report.
"The most interesting aspect of the work to me is identification of this structure in the human brain," he says. "In the past it has been observed that stem cells are created in the brain. Now we know where."
But almost as interesting is the indication that the brain suppresses the ability of these stem cells to become functioning nerve cells, Sanai says.
"Understanding the mechanism behind that repressive activity could be a very powerful tool," he says. "We could manipulate that area and other areas in the opposite direction to harness the potential of stem cells."
More information
The basics of stem cells and their possible use in medicine can be found at the National Institutes of Health (stemcells.nih.gov target=_new) or Northwestern University (www.northwestern.edu target=_new).
Antibiotic provides promise in treatment of spinal cord injuries
Treatment prevents later-stage tissue loss contributing to long-term injury
Researchers at Brigham and Women's Hospital (BWH) and Children's Hospital Boston (CHB) have found that a commonly prescribed antibiotic could be used to help prevent paralysis and other long-term functional deficits associated with a partial spinal cord injury (SCI). Researchers in the field have known that a significant proportion of paralysis and long-term functional disorders associated with SCI are triggered by post-trauma tissue loss. Administering the antibiotic, minocycline, to rats within the first hour after a paralyzing injury has been shown to reduce this tissue loss and ultimately enable more hind-leg function
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Minocycline, a well-known neuroprotector that is currently being tested to treat stroke, ALS, Huntington's disease and head trauma, has no observable side effects in the rat model and can be given for up to an hour after SCI, providing a more realistic timeframe for clinical use.
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Since MP can be administered up to 8 hours after an injury it actually allows more flexibility than minocycline.
quote:
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"We believe that if minocycline is demonstrated effective in clinical trials of SCI, it will likely be part of a comprehensive cocktail of medications targeting the acute and chronic injuries of this devastating disease," said Robert M. Friedlander, MD of BWH, HMS associate professor of Neurosurgery and co-lead author of the study. "Because minocycline has already been proven as an effective neuroprotector and is capable of penetrating the blood-brain barrier, we believe that it may become the next-generation therapy for treating SCI."
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Key advance reported in regenerating nerve fibers
Two-pronged approach synergizes growth
BOSTON -- Researchers at Children's Hospital Boston and Harvard Medical School have advanced a decades-old quest to get injured nerves to regenerate. By combining two strategies – activating nerve cells' natural growth state and using gene therapy to mute the effects of growth-inhibiting factors – they achieved about three times more regeneration of nerve fibers than previously attained. The study involved the optic nerve, which connects nerve cells in the retina with visual centers in the brain, but the Children's team has already begun to extend the approach to nerves damaged by spinal cord injury, stroke, and certain neurodegenerative diseases. Results appear in the February 18th Journal of Neuroscience.
Normally, injured nerve fibers, known as axons, can't regenerate. Axons conduct impulses away from the body of the nerve cell, forming connections with other nerve cells or with muscles. One reason axons can't regenerate has been known for about 15 years: Several proteins in the myelin, an insulating sheath wrapped around the axons, strongly suppress growth. Over the past two years, researchers have developed techniques that disable the inhibitory action of myelin proteins, but this approach by itself has produced relatively little axon growth.
The Children's Hospital team, led by Dr. Larry Benowitz, director of Neuroscience Research, reasoned that blocking inhibition alone would be like trying to drive a car only by taking a foot off the brake. "Our idea was to step on the gas – to activate the growth state at the same time," Benowitz said. "Knocking out inhibitory molecules alone is not enough, because the nerve cells themselves are still in a sluggish state."
The researchers injured the optic nerves of rats, then used a two-pronged approach to get the axons to regenerate. To gas up the sluggish nerve cells, Dr. Dietmar Fischer, first author of the study, caused an inflammatory reaction by deliberately injuring the lens of the eye. Though seemingly harmful, this injury actually stimulates immune cells known as macrophages to travel to the site and release growth factors. As Benowitz's lab had found previously, these growth factors activated genes in the retinal nerve cells, causing new axons to grow into the optic nerve.
To try to enhance this growth, the researchers added a gene-therapy technique. Using a modified, non-infectious virus as a carrier, they transferred a gene developed by co-investigator Dr. Zhigang He into retinal nerve cells that effectively removed the "braking" action of the myelin proteins – spurring production of a molecule that sopped these inhibitory proteins up before they could block growth.
"When we combined these two therapies – activating the growth program in nerve cells and overcoming the inhibitory signaling – we got very dramatic regeneration," said Benowitz, who is also an associate professor of neurosurgery at Harvard Medical School and holds a Ph.D. in biology/psychobiology. The amount of axon regeneration wasn't enough to restore sight, but was about triple that achieved by stimulating growth factors alone, he said.
Benowitz's lab will continue working with the optic nerve in hopes of restoring vision. "We have to fine-tune the system, and we have some ideas of how to do it," Benowitz said. "But then we come to another big hurdle." That hurdle is getting the nerve fibers from the eye to hook up to the correct centers in the brain in such a way that visual images do not become scrambled. "It's a mapping problem," Benowitz said. "We have to retain the proper organization of fiber projections to the brain."
Meanwhile, he and his colleagues have begun using a similar two-pronged approach to regrow axons damaged by stroke or spinal-cord injury. They have already found a way to step on the gas – using a small molecule known as inosine to switch damaged nerve cells in the cerebral cortex into a growth state. In 2002, they reported that inosine helped stroke-impaired rats to regrow nerve connections between brain and spinal cord and partially recover motor function.
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The current research was supported by the National Eye Institutes, Boston Life Sciences Inc., the German Research Foundation, and the Paralyzed Veterans of America.
Children's Hospital Boston is home to the world's largest research enterprise based at a pediatric medical center, where its discoveries have benefited both children and adults for more than 130 years. More than 500 scientists, including seven members of the National Academy of Sciences, nine members of the Institute of Medicine and nine members of the Howard Hughes Medical Institute comprise Children's research community. Children's is the primary pediatric teaching affiliate of Harvard Medical School. For more information about the hospital visit: www.childrenshospital.org
Scientists have created an unlimited supply of a type of nerve cell found in the spinal cord – a self-renewing cell line that offers a limitless supply of human nerve cells in the laboratory. Such a supply has long been one goal of neurologists anxious to replace dead or dying cells with healthy ones in a host of neurological diseases
Spinal cords fixed in tests
By Lee Bowman
Scripps Howard News Service
With a genetic tweak, scientists have created an unlimited supply of a type of nerve cells found in the spinal cord and have been able to use the cells to partially repair damaged spinal cords in lab animals.
While application of the discovery to humans is still years away, being able to generate such a limitless supply of the specialized nerve cells has long been a goal toward treating many neurological diseases.
EVANSTON, Ill. --- Scientists at Northwestern
University have designed synthetic molecules that
promote neuron growth, a promising development that
could lead to the reversal of paralysis due to spinal
cord injury.
"We have created new materials that because of their
chemical structure interact with cells of the central
nervous system in ways that may help prevent the
formation of the scar that is often linked to
paralysis after spinal cord injury," said Samuel I.
Stupp, Board of Trustees Professor of Materials
Science and Engineering, Chemistry and Medicine.
19:00 22 January 04 NewScientist.com news service
A liquid that forms a gel-like mass of nanofibres on
contact with water could provide the most promising
vehicle yet for the regeneration of damaged spinal
cords.
Many groups working in regenerative medicine are
trying to develop artificial scaffolds that store or
attract cells and then control their growth and final
identity.
Chemical gradient steers nerve growth in spinal cord
A research team at the University of Chicago has discovered a crucial signaling pathway that controls the growth of nascent nerves within the spinal cord, guiding them toward the brain during development.
The study, published in the Dec. 12, 2003, issue of the journal Science, solves a long-standing scientific mystery. It may also help restore function to people with paralyzing spinal cord injuries.
"This is the first guidance mechanism that regulates growth of nerve cells up and down the spinal cord," said Yimin Zou, Ph.D., assistant professor of neurobiology, pharmacology and physiology at the University of Chicago.
"This is exciting to scientists because these neurons are the primary model system we use to understand assembly of the nervous system," he said. "It's exciting to clinicians because it could help regenerate damaged axons in the central nervous system."
The study focused on commissural neurons, which are found in the spinal cord. These neurons receive sensory signals such as pain, heat or cold from the primary neurons that reach from the hands or feet, for example, to the spinal cord. The commissural neurons relay those signals up the spinal cord to the nerve cells that process the information in the brain.
In a meticulous series of experiments with rats, Zou and colleagues show that a gradient of chemoattractant(s) along the spinal cord, probably formed by one or multiple Wnt proteins, lures growing commissural neurons toward the brain.
The Wnt family of proteins carry signals from cell to cell, regulating the interactions between cells during many development processes. Wnt proteins bind to receptors of the "Frizzled" family on the cell surface.
In the Science paper, Zou and colleague show that the Wnt gradient is detected by a receptor known as Frizzled3, found at the tips of these growing neurons. Commissural axons in Frizzled3-deficient mice (generously provided by Jeremy Nathans of Johns Hopkins Medical School) lost directionality of growth along the spinal cord.
If Wnt proteins could be used to entice damaged commissural neurons to regenerate and restore the connections between nerve cells of the spinal cord and the brain, it could revolutionize treatment of paralyzing spinal cord injuries.
Many researchers are studying ways to use stem cells to regenerate damaged tissues. Even if stem cells can be successfully "trained" to become the type of neurons needed and transplanted into the damaged central nervous system, "they still need to be guided precisely to their targets in order to rebuild the connections," explained Zou. "Understanding how the brain and spinal cord are connected during embryonic development should give us clues about how to repair these connections in adulthood."
But, "this is just half of the battle," Zou cautioned. A spine-injured patient would also have to rebuild the other nerves, which carry messages from the brain to the spinal cord, such as the corticospinal tracts. The cues that steer these brain axons down the spinal cord have not yet been identified.
Scientists have long wondered how something as complex as the human nervous system, with more than 100 billion neurons, each connected to a thousand or more target cells, gets correctly assembled.
In the 1990s, they found the first of many chemical signals that regulate the growth of commissural neurons, helping them locate, recognize and connect with their appropriate partners. Several sets of signals work together to guide these budding nerve cells through each step.
These cues act on the growing tips of axons, long narrow processes sent out by neurons in search of other nerve cells. Axons are tipped with growth cones that can detect extracellular signals, such as Wnt4, and then grow toward or away from the source.
The axon's journey from the cell body of a commissural neuron, found at the back of the spinal cord, up to the brain is a long and complicated one. It relies on the coordinated action of several signaling systems, each controlling one part of the journey then handing off to a different set of cues.
Substances known as Netrin-1 and Sonic hedgehog, for example, tell the axons from commissural neurons to grow from the back of the spinal cord to the front. As these axons cross the midline, they stop responding to Netrin and Sonic hedgehog but begin to respond to a new set of proteins, known as Slits and Semaphorins, that repel them, shifting the axis of growth away from back to front (dorsal-ventral) and toward top to bottom (anterior-posterior).
At that point, Wnt proteins take over, drawing the axons up toward the brain. Without the Wnt/Frizzled signaling, the axons wander aimlessly, "knotting and stalling," noted the authors.
These findings will also allow scientists to explore "how growth cones undergo remodeling during navigation so that they constantly adjust the direction of their growth," Zou added. "This should help explain how the complicated connections in our nervous system are established and potentially lead to ways to remobilize the guidance programs to repair the damaged circuits in adulthood."
Pioneers bring hope of cure for paralysis
Scottish lasers’ key role in experiment
By Noel Young
http://ww1.sundayherald.com/37910
Hopes of a breakthrough in treating patients with paralysing back injuries have been raised by a groundbreaking experiment which for the first time fused together a mammal’s broken spinal cord.
The technique in which Scottish-designed lasers were used to repair the spinal cords of paralysed rats – restoring the animals’ mobility – is likely to be tested on humans next year.
Professor Juanita Anders, the lead researcher on the project at the Uniformed Services University in Maryland, said that some of the advances in the field of light therapy are “almost too incredible to believe.”
Dr Jackson Streeter, whose firm has now licensed the technology developed by Anders, said: “If we could have lasered Christopher Reeve in the days immediately after his injuries, he might have been walking today”.
At the moment the university researchers are concentrating on acute injuries – those which have just happened – as opposed to chronic conditions. They are confident however that the more complex treatment of old injuries – such as those sustained by Reeve in 1995 – is also within their grasp.
The US university where the laser work has been carried out is funded by the US Defence department. It has 900 students, most of whom wear uniform.
The university is on the same heavily-guarded campus as Bethesda Naval Hospital where President George W Bush undergoes regular medical examinations.
The laser breakthrough work was carried out by Anders, her associate Dr Kimberly Byrnes and six other team members. Using lasers from Thor International, of Kilmartin, Argyll, the team was able to restore complete mobility to 10 white laboratory rats which had previously had their spinal cords cut.
A group of ten rats, which also had their cords cut but were not given the light treatment made no recovery.
“The 10 animals chosen received daily doses of light for about 50 minutes a day for two weeks, “ said Anders. “Nine weeks later when they were tested, they had recovered their mobility.”
Dr Byrnes will make a presentation on the work, which earned her a PhD, at a Society for Neuroscience conference in New Orleans today. The research will also be detailed in a book issued to delegates at the conference, the world’s largest and most influential gathering of its kind with 28,000 experts expected to attend.
The experiment has its roots in former US President Ronald Reagan’s controversial Star Wars programme. “We were called to a conference and asked to put up bio projects involving the use of lasers,’’ said Anders. ‘‘My project won funding of $60,000 a year and we were on our way.”
The toughest challenge facing the scientists was how to establish that lasers were actually penetrating the flesh and reaching the broken spinal cord.
That was where the lasers produced by the tiny Thor company came into their own. Thor employs just three people at Kilmartin, designing and building the instruments, led by Peter Gaskin. Three more staff at its adminstrative and marketing HQ in Amersham, Buckinghamshire, make up the workforce.
Managing director James Carroll said: “I don’t know how the Americans heard about us but we have since had lots of flattering e-mails, praising the stability of our equipment.’’
Anders said: “The power output from other lasers tended to be inconsistent and we had to be precise. The Thor machines were the right ones for the job.”
She added that they still did not fully understand the mechanism, in much the same way that the mechanism of acupuncture is not understood.
“We believe the light somehow alters the behaviour of the cells, inhibiting the immune system and allowing the neurons that make up the spinal cord to regroup,’’ she said.
The professor added that she had faced massive cynicism over the years regarding her research. She said: “They would ask me: ‘Why are you wasting your time on this? You will never get light to penetrate flesh.’’’
Her immediate boss was among the unconvinced, “until I showed him the changes in the cells of the rats after our experiments. Now he is completely on side,” she said.
The lab team are now working on ways to enable the light to reach the spinal cord in a human. “So far the results are encouraging,” said Anders.
The first tests are likely to be carried out by PhotoThera, a firm in San Diego, California, and will initially concentrate on the treatment of strokes.
“We believe the technique developed by Dr Anders will penetrate skull bone and tissue and enable us to reach the site of a stroke with a laser and minimise brain damage,” said the company’s Dr Streeter. Tests on spinal injuries are also a priority but may be delayed because the project’s neurosurgeon who was working with Streeter is in Iraq and it is not known when he might return.
“We are tentatively aiming for the beginning of 2005 to commence our research,” said Streeter.
The title: ” Fetal cell transplantation for spinal cord injury ”
The short title: “ Cell transplantation for spinal injury”
Samuil S. Rabinovich1, Victor I. Seledtsov2, Olga V. Poveschenko2, Vladimir V. Senuykov2, Vadim Ya. Taraban2, Vladimir I. Yarochno, Nicolay G. Kolosov1, Vladimir Kozlov2, Sergey A. Savchenko1.
1 Novosibirsk State Medical Academy, 2 Institute of Clinical Immunology, 3 34 Municipal Clinical Hospital, Novosibirsk, Russia.
The corresponding author:
Victor Seledtsov M.D., D.Sc.
Institute of Clinical Immunology
Russian Academy of Medical Sciences
14 Yadrintsevskaya str.
630099, Novosibirsk, Russia
e-mail: vs@online.nsk.su
Tel/Fax: +7 (3832) 28-26-73
Fax: +7 (3832) 22-70-28
Abstract.
The cells from fetal nervous and hemopoietic tissues (gestational age 16-22 weeks) were subarachnoidally implanted into 15 patients of 18-to-52 year old with severe consequences of the traumatic spinal cord injure (SCI) at cervical or thoracic spine level. The times after SCI were from 1 month to 6 years. The number of fetal cell transplantations (FCT) into a patient varied from 1 to 4. The time intervals between FCT were 14-to-30 days. In 11 of 15 cases FCT was performed immediately after resection of the connective tissue cyst(s) that has been formed within the site of SCI. Before FCT the neurological state of each of the patients was characterized by complete motor and sensory function disorder and was consistent with a grade A of SCI according to Frankel classification. With FCT therapy, the neurological state of 6 patients became clinically consistent with C grade of SCI. These patients received an ability to walk with the aid of crutches or a special hinged apparatus. The less, although noticeable, clinical improvements were noted in other 5 FCT-treated patients. Their state became consistent with grade B of SCI and was characterized by appearance of contracting activity in some muscles and incomplete restoration of sensitivity. Lastly, 4 patients subjected to FCT therapy did not demonstrate any clinical improvements. No serious complications of FCT were noted. The results presented herein point out a clinical relevance of the FCT-based approach to treating severe consequences of SCI.
The human adult nervous tissue is well known to have a very limited reparative potential. Therefore, the treatment of the patients with severe injures of the central nervous system (CNS) frequently do not give desired results. Over last decades a considerable progress was reached in the understanding of the mechanisms regulating cell reparative activity in CNS. The accumulating evidence clearly outlines two feasible approaches to improve reconstruction of injured CNS. The first is based on creation in CNS of conditions which are favorable to nerve fiber growth, whereas the second, on replacement of destroyed neurons by new functionally active neural cells. The transplantation of fetal-derived immature cells appears to unite both these approaches. In fact, it has been established that when grafted into adult CNS the fetal-derived cells are able both to elaborate the factors favoring axonal growth from recipient neurons and to provide the generation in CNS of new functionally active donor neurons [reviewed in 1-3]. In this paper we present the results of applying fetal (neural plus hemopoietic) cell transplantation (FCT) therapy in 15 spinal cord injury (SCI) patients.
MATERIALS AND METHODS
The study was performed in the exact accordance with the protocol that has been approved by the Scientific Council and Ethics Committee at the Institute of Clinical Immunology. Informed consent was obtained from each subject who have been enrolled in the study.
The fetal brain neural and hemopoietic liver tissues were isolated from human fetus (gestational age 16-22 weeks) after spontaneous or prostaglandin-induced abortion, and then prepared in the form of cell suspension, as described early [4]. The cells were further cryopreserved in the standard way in RPMI 1640 medium containing 50 % fetal bovine serum and 10 % dimethyl sulfoxide [5], and stored in liquid nitrogen until use. On the day of transplantation, the cell suspensions were thawed at 370 C, washed extensively, and assayed for cell viability by a trypan blue exclusion method in the routine way. The cell suspension designed for transplantation was composed of equal numbers of cells obtained from three distinct donors. The overall number of viable cells in such suspension was 25 x 107, whereas a ratio of cells from neural tissue to those from liver one was 10.
When it was reasonable, the FCT was performed immediately after operative resection of connective tissue cyst that has been formed within the site of SCI. In the several cases, along with the above described cell suspension, a fetal spinal cord fragment together with olfactory ensheathing cells (2 x 105) was implanted into spinal cord lesion. The olfactory ensheathing cells have been previously reported [reviewed in 1] to be effective stimulators of nerve fiber growth and myelination.
Before FCT therapy, all 15 patients with SCI enrolled in the study had complete motor and sensory function disorder. The patient characteristics ( age, time after SCI, lesion level, number of performed FCT) are shown in Table 1. The neurological status of the patients was examined in terms of modified Frankel definitions. For the Frankel score, a five scale subdivision was used: A = complete motor and sensory function disorder; B = motor complete, sensory incomplete function disorder; C = motor and sensory incomplete function disorder; D = useful motor function with or without auxiliary means; E = no motor or sensory function disorder [6]. The dynamic examination of motor functions in the patients was conducted with usage of ASIA (American Spinal Injury Association) motor scale [6].
RESULTS
As shown in Table 1, clinical improvements was noted in 11 of FCT-treated 15 patients. Six patients changed their neurological state from A to C grade of SCI. These patients received an ability to walk with the aid of crutches or orthoses. The less, although noticeable, clinical improvements were noted else in FCT-treated 5 patients. Their state became consistent to grade B of SCI and was characterized by appearance of contracting activity in some muscles and incomplete restoration of sensitivity. Lastly, 4 FCT-treated patients did not demonstrate any clinical improvements. Three cases with most improvements are described in detail below.
Case 1. 52-year-old male patient was admitted to the Emergency City Hospital 6 hours after a vehicular accident. On admission the patient was in grave condition: he was stuporous, although followed instructions and correctly answered simple questions; his puls rate was 110 b min –1, arterial blood pressure 90/60; his respiration was self-dependent with the of 22 min –1 ; the functions of his granial nerves were not compromised; there was teraplegia and apparent muscle hypotonia; his tendon and periosteal bone reflexes were flaccid in both upper and lower extremities (D=S), his abdominal reflexes were absent; there was profound disorder of all kinds of sensitivity on both sides of his body lower Th1 level, but also vesical disfunction; the palpation of _3-to-C6 vertebras was painful. X-ray examination failed to reveal any traumatic bone changes in cervical spine. Magnetic resonance imaging (MRI) showed the rounded area (0.7 x 0.5 cm) of hypointensity with clear-cut contours at C3-C4 junction (see Figure 1, A) and the centro-dextral spindle-shaped area of 5.8 x 0.3 cm, giving signal of increased intensity at T2 regime along the whole length from C2 to C5 level. A diagnosis was made out: a cord contusion at the C3-C5 level and a syndrome of complete SCI.
The standard therapy aimed for brain dehydration was started immediately after the cervical spine fixation. The cell suspension (10 ml) composed of cells obtained from fetal neural and hemopoietic tissues was grafted subarachnoidally (via lumbar puncture) into patients 4 days after trauma (day 0). On 28 days after FCT the first signs of spinal function restoration were noted: the level of sensory disturbance lowered down to Th4 and some motor activity of the left foot appeared. A clinic examination of the patient performed on 40 day after FCT revealed the following: a complete restoration of pain and touch sensitivity; kinesthesia restoration (S>D); the appearance of some sighs for vesical reflex; active leg motion with the strength of 3 and 2 points (according to the ASIA scale) at the left and the right side, respectively. The muscle strength in arms was 2 points at the left side and absent at the right one. Tendom and periosteal reflexes were active. MRI scan showed the intramedullar heterogeneous formation with indistinct contours (the cyst might presumably be embedded in neural cell graft) at C7, as well as the normal thickness and structure of the spinal cord bellow the level of SCI (Th1-Th3) (see Figure 1, B). On 96 day after FCT, the complete restoration of all kinds sensitivity and functions of pelvis organs was noted. The strength in arms and legs was found to reach for 4 and 5 points, respectively. The patient began walking with crutches.
Case 2. 24-year-old male patient with SCI at _7 was admitted to the neurosurgical department 6 days after an incidence. In the view of bursting fracture of C7 vertebra and spinal cord compression the operation was made: a partial resection of C7 vertebral body and ventral spondylodesis at _6-Th1 with a titan-nickel (Ti-Ni) implant. Seven weeks later patient was again admitted to the neurosurgery department for further his management. The overall health status of the patient on admission was of average severity. His consciousness was clear, cardiovascular and respiratory systems were without any pathological changes. Neurological parameters according to the ASIA scale was described as follows: inferior flaccid paraplegia, superior paraparesis (strength of arms at distal and proximal parts was 3 and 5 points, respectively), conductive type of disorders of all kinds of sensitivity lower Th1 level, tendon and periosteal reflexes active on upper extremities and flaccid on lower ones (D=S). Urination was via cystostoma. The patient condition was complicated by 26 _ 18 _ 5 cm necrotic pressure ulcer over the sacrum. MRI examination revealed deformation and size diminution of C6-_7 vertebral bodies. At C6-_7 level Ti-Ni implant was visualised. The spinal cord was thin at the injure level, but its continuity was not broken. The intramedullary cyst of 1.01 _ 0.2 cm was clearly visualized at _7 segment (Figure 2, A). At Th1-3 level the spinal cord heterogeneity with the hyperintensive areas of up to 0.12 cm were detected in T2 regime.
The patient was subjected to three FCT with 14 day intervals. At the first and third FCT, the suspension of viable cells in a volume of 10 ml was injected subarachnoidally via a lumbar puncture. The intermediate second FCT was performed immidiately after surgery: laminectomy at C7, extraction of intramedullary cyst and implantation into the made cavity of a fetal spinal cord fragment enriched with olfactory ensheathing cells.
One and a half month after the last FCT, the apparent clinical improvements in the patient were noted: complete restoration of sensitivity and vesical reflex , the presence of strength of 5 points in arms from both sides and some motions in foots and in knee joints, and complete healing of pressure ulcer. On MRI scan in the field of a former cyst the areas of various densities with indistinct contours were seen (see Figure 2, B). Those areas might be visible manifestation of reparative activity of the grafted cells. Ten month after the FCT therapy the patient entirely controlled the functions of pelvis organs. He was able to stand, leaning on crutches, and to walk, using a special hinged apparatus.
Case 3. 25 year-old male patient had received bursting fracture of C7 vertebra with spinal cord compression, as a consequence of a road traffic accidence. Some days later he underwent liminectomy Th6-Th7 and 4 months later he was again admitted into the neurosurgical department for further his management. On admission the patients was in condition of middle severity. His consciousness was clear, cardiovascular and respiratory systems were without any pathological changes. Neurological status had the following characteristics: inferior flaccid paraplegia, conductive disorders of all kinds of sensitivity lower Th5 level. Urination was via cystostoma. MRI scan showed deformation and size diminution of Th6 vertebral body. The spinal cord was thin at the injure level, but its continuity was not broken. At Th6 level, a 3.0 _ 0.5 cm intramedullary cyst was clearly seen. The spinal cord heterogeneity with the hyperintensive areas of up to 0.7 cm at were also visualized in T2 regime.
The patient was subjected to two FCT with 30 days interval. The first FCT was preceded by the surgery: repeated laminectomy at Th5-Th7 level, extraction of intramedullary cyst and implantation into the formed cavity of a fetal spinal cord fragment enriched with olfactory ensheathing cells.
Four months later the last FCT the patient demonstrated complete restoration of all kind of sensitivity, overall control of the functions of pelvis organs and evident improvements in motor sphere: he was able to stand, leaning on crutches.
DISCUSSION
By present there is already a valid experimental basis for applying FCT-based therapy in treating patients with CNS injures [reviewed in 1-3]. It seems quite reasonable that realization of the reparative potential of fetal-derived immature cells in such patients may greatly improve outcome of their disease. Importantly, novel techniques of propagation of immature multi- and unipotent cells in vitro, which are being now actively developed, allow to solve not only technical, but also ethical problems confronting progress in transplantology [reviewed in 7,8] and, thereby, may promote widespread adoption of FCT-based advances in clinical practice. An effective CNS repair appears to require the presence in injured sites of not only neural cells potentially able to provide axonal growth, but also the other cells capable of creating the microenvironment favorable to both growth and myelination of nerve fibers. In our own investigation we transplanted into the SCI patients not only the cells from fetal nervous tissues, but also the fetal liver cells belonging mainly to erythroid and myeloid differentiation lineages. In fact, evidence is accumulating that 1) fetal hematopoietic tissues contain the stem cells capable of differentiating into cells forming nervous tissue [7, 9]; 2) hemopoietic cells can elaborate the mediators able to support the growth and viability of distinct cells including the cells constituting nervous tissue [reviewed in 7]; 3) these cells possess a potent natural suppressor activity [10] that may be directed against developing cell transplantat-induced immune processess; and 4) they may contribute to neovascularization of ischemized tissues [11, 12]. Lastly, there are publications indicating a capacity of immature hemopoietic cells not only to inhibit but even to reverse the development of scar connective tissue [13,14] that is known to represent a insuperable obstacle to axonal growth. Thus, it is reasonable to believe that the fetal hemopoietic tissue-derived cells may be capable of markedly contributing the repair of adult CNS.
In our view, FCT into a SCI patient is most optimal before the formation in the injured site of the fibrous connective cyst that may be a major obstacle for restoration of spinal nerve communications. It is reasonable that in the cases with the formed cysts, FCT should be performed immediately after resection of the cyst and restoration, as far as possible, of canals for nerve fiber growth.
The clinical success of FCT is believed to be significantly defined by survival of the cells implanted in the body. In fact, the fetal neural cells have been previously demonstrated to be able to survive and function in major histocompatibility complex (MHC)- incompatible adult CNS for relatively long period of time [reviewed 1-3]. It should be had in mind, however, that immune privileges of CNS cannot yet guarantee immune unresponsiveness of the host to the allogeneic cells grafted into CNS. Therefore, we transplanted into a recipient only those cells whose alloantigens failed to induce the activation of recipient’s T lymphocytes in vitro assays which have been performed before FCT. Moreover, to our opinion, the transplantation of cells from more than one donor may markedly lessen a risk for prompt immune-mediated rejection of all donor cells. Actually, such cell transplantation might lead to the situation when host’s immune responses might be directed against only a part of the most antigenically incompatible cells, while another part of the cells might survive and be involved in CNS repair. Consistent with this proposition, we, indeed, found that 4 of 12 patients with severe cranial traumas, who were subjected to more than one FCT in the above-described way, developed both cell and humoral immune responses directed against the part of, but not all, transplanted cells (data not presented).
The cases reported herein indicate that the FCT therapy of SCI patients may result in apparent clinical improvements. Only 4 of 15 FCT-treated patients with time after SCI of 2, 3, 4, and 6 years did not show any improvements in their neurological status. Nevertheless, a time boundary, behind which FCT therapy become inefficient, was not yet determined. In fact, this boundary may be quite blurred and dependent on the individual features of patients. In this connection, it should be paid attention to the appreciable clinical improvements in 4 FCT-treated patients with times after SCI from 2.5 to 3 years.
In all 15 cases described in this paper, FCT therapy led to noticeable improvements in patient’s psychological status, which were noted by both the patients, themselves, and their nearest relations.
FCT into CNS appears to be safe and well tolerated. Meningisms, but also raises of body’s temperature up to 38.5 C, were noted in the part of patients during 24-to-48 h after FCT. Those occurrences disappeared, by themselves, not requiring any additional medicament interventions. A long-term (3 year ) follow-up of 18 patients (mainly with brain injures) subjected to FCT therapy did not reveal any serious complications which might be related to grafted fetal cells (manuscript in preparation) .
CONCLUSION
The results presented herein suggest that FCT-based therapy may be successfully applied in treating neurological sequences of SCI. Although much greater clinical experience is needed to determine place and clinical relevance of this therapy in overall complex treatment of the patients with CNS injures.
REFERENCES
1. Reier PJ, Anderson DK, Thompson FJ et al. J Neurotrauma 9 (Suppl. 1), S223-S248 (1992).
2. Famcett JW. Spinal Cord 36, 811-817 (1998).
3. Brundin P, Karlsson J, Emgard M et al. Cell Transplantation 9, 179-195 (2000).
4. Seledtsov VI, Avdeev IV, Morenkov AV et al. Immunobiology 192, 205-217 (1995).
5. Adams RLP. Cell culture for biochemists. Elevier/North-Holland Biomedical Press, 1980: 130.
6. Capaul M, Zollinger H, Satz N et al. Paraplegia 32, 583-587 (1994).
7. Sukhikh GT. Bull Exp Biol Med 126 Suppl 1, 3-13 (1998) (in Russian)
8. Gray JA, Grigoryan G, Virley D. et al. Cell Transplantation 9, 153-168 (2000).
9. Eglitis MA and Mezey E. Proc Natl Acad Sci USA 94, 4080-4085 (1997).
10. Seledtsov VI, Seledtsova GV, Samarin DM et al. Immunobiology 198, 361-374 (1998).
11. Murohara T, Ikeda H, Duan J et al. J Clin Invest 105, 1527-1536 (2000).
12. Fuch S, Baffour R, Zhou M. et al. J Am Coll Cardiol 37, 1726-1732 (2001).
13. Moiseev AY, Samarin DM, Kustov SM et al. Bull Exp Biol Med 126 Supp 1, 129-130 (1998) (in Russian).
14. Kolosov NG, Poveshchenko OV, Efremov AV et al. Bull Exp Biol Med 126 Supp 1, 128-129 (1998) (in Russian).
Table 1. Characteristics of SCI patients and results of their FCT therapy in the term of Frankel definitions.
No. Patient, age (years) Level of SCI Time after SCI Number of FCT Neurological status in terms of Frankel definitions
________________________
before and after FCT therapy
1 K, 52 _6-7 1 month 2 _ _
2 Z, 24* _6 3 months 3 _ _
3 G, 28* Th11 3 months 2 _ _
4 _, 25* Th6 4 months 3 _ _
5. V, 18 Th4 4 months 2 _ _
6. _, 34* Th10 1.5 years 2 _ _
7. _, 34* _6 1.5 years 2 _ _
8. _, 48 Th3 2 years 2 _ _
9. Sh, 28* _6 2.5 years 3 _ _
10. S, 32 * Th9 2.5 years 3 _ _
11. D, 32* _6 3 years 1 _ _
12. S, 18* _7 3 years 2 _ _
13. _, 21 Th4 3 years 2 _ _
14. Kh, 46* _4-6 4 years 4 _ _.
15. _, 38* Th6 6 years 1 _ _
*the case with operative cyst resection
A finding about how genetically engineered stem cells heal damaged nerve fibers months after an injury has provided further evidence that stem cells release therapeutic molecules that can help treat spinal cord damage.
Researchers from Johns Hopkins University School of Medicine in Baltimore, Maryland report that nerves of injured rats regenerated as much as six months after treatment with modified stem cells from mice.
"This is the first demonstration of regeneration in chronically denervated nerves," says Ahmet Hoke of Johns Hopkins.
Spinal cord injury
Spinal cord injury occurs when a traumatic event results in damage to cells within the spinal cord or severs the nerve tracts that relay signals up and down the spinal cord.
When injured in accidents, or by destructive illnesses such diabetes and AIDS, these nerves typically do not regenerate very well.
"The more time a nerve has been disconnected from its target, and the greater the length of nerve regeneration needed, the worse the chances are," says Hoke.
Because stem cells have the potential to develop into any type of cell in the human body, scientists are hoping to use them to repair or replace damaged nerve cells.
Cell mimicry
To this end the researchers attached a freshly cut nerve to one that had been cut six months before and allowed to deteriorate.
They then transplanted stem cells from the nervous systems of mice into the area of nerve repair.
The researchers hoped that stem cells would mimic Schwann cells—a type of supporting cell that helps separate and insulate nerve cells.
"Schwann cells normally secrete growth factors and other proteins that enhance regeneration," says Hoke.
Using stem cells genetically engineered to make a growth factor called GDNF, the researchers observed the regeneration of nerve fibers within six months.
Double benefit
Biochemical analyses of the area surrounding the regenerating nerves suggested that the stem cells were not only providing nutritive molecules but also helping to clear the area of proteins that prevent regeneration.
"Our study gives insight into possible mechanisms of regeneration in chronically denervated nerves, but there's much to do," says Hoke. "We'd like to improve the recovery of nerve function so it's higher than 25%. We also need to identify exactly what combination of growth factors and other proteins and enzymes are missing in chronically denervated nerves."
The findings add to evidence that stem cells support nerve cell regeneration by producing supportive molecules.
This June, researchers reported that stem cells support nerve cell survival in paralyzed rats by releasing TGF-alpha, which promotes nerve cell survival, and BDNF, which strengthens nerve cell connections.
The study by Hoke and colleagues was reported in San Francisco at the annual meeting of the American Neurological Association.
SCIENTISTS USE A VARIETY OF CELLS AND MOLECULES TO COAX REGROWTH OF DAMAGED NERVES.
ORLANDO, Sunday, Nov. 3 - Step by step, scientists are making new advances in regenerating damaged nerves and nerve fibers - advances that may one day help people who have conditions or diseases in which nerve cells have become impaired or have ceased functioning entirely, including spinal cord injuries, blindness, stroke and multiple sclerosis.
One group of scientists, for example, has discovered that transplants of the ensheathing cells from the nerves that humans use to smell can help spinal cord damaged rats regain some of their ability to walk. Another group, also working with rats, has developed a synthetic peptide that promotes new nerve fiber growth in damaged spinal cords. In addition, scientists have found that transplants of a particular cell line can limit loss of vision in rats with retinal degenerative disease. The new studies were reported today during the 32nd annual meeting of the Society for Neuroscience.
Hans Keirstead, PhD, and his colleagues at the University of California-Irvine have found that transplanted human olfactory ensheathing cells (OECs) can help rats with injured spinal cords recover some ability to walk.
"This is the first time human OECs prepared in this high purity manner have been used to investigate their ability to treat the injured spinal cord," says Keirstead. "Because our study used human cells, it has direct significance for clinical use."
OECs are brain cells that enable our sense of smell. When injured, these cells are able to replace themselves - a very rare trait among brain cells. Usually, it's only young cells that are able to regenerate.
"OECs are thus considered a good candidate for the treatment of an injured brain or spinal cord," says Keirstead. In the past, scientists have investigated whether rat OECs are effective in treating injured spinal cords, but with mixed results. Some of the treated animals showed excellent recovery; others showed no recovery at all.
Keirstead and his colleagues wanted to determine what, if any, behavioral and cellular effect human OECs would have on injured spinal cords. They set up an experiment in which they transplanted human OECs into one group of spinal cord injured rats and did not treat another group of similarly-injured rats.
"We first observed that the injured animals transplanted with these human olfactory ensheathing cells were able to regain some walking ability," says Keirstead. "We also observed some regrowth of neurons in the injured spinal cords of the animals that had received the transplants - but not in the spinal cords of non-transplanted animals."
The scientists found that scar formation, which commonly occurs around injury sites and which prevents the growth of neurons, was present in both the transplanted and non-transplanted animals. But in the transplanted animals, some growth of neurons occurred despite the scarring. "This suggests that human OECs transplanted into an area of spinal cord injury provide some support that enables regrowth in an environment that would otherwise prevent it," says Keirstead.
The scientists also found that the human OECs not only survived the period through which the animals were kept alive, but that the cells migrated from the transplant site into its surrounding. Such an ability to migrate through the adult nervous system is a very rare trait, and suggests that the cells may be able to cause regrowth beyond the injury site.
Keirstead is now trying to better understand how human OECs permit the regrowth of neurons. "We're also evaluating the effects of these cells following transplantation into chronically injured spinal cords," says Keirstead. "This would allow us to determine if these human cells are good candidates for treating injuries that have taken place months and maybe even years earlier."
At Yale University, Stephen Strittmatter, MD, PhD, and his colleagues have developed a synthetic peptide that promotes new nerve fiber growth in the damaged spinal cords of laboratory rats, allowing them to walk better. The peptide, NEP1-40, promotes the growth by suppressing the effects of the "Nogo" gene, which has been identified as the gene that prevents the brain and spinal cord from rewiring themselves after an injury.
"We have developed a way to block Nogo action with a peptide that binds to the Nogo receptor, thus preventing it from doing its normal job," says Strittmatter. This finding could one day help people who have experienced a brain or spinal cord injury, a stroke, or who have a degenerative disease such as multiple sclerosis.
To examine whether NEP1-40 would block Nogo and promote nerve regrowth, Strittmatter and his colleagues administered the peptide to spinal cord injured rats for four weeks through a catheter inserted into the animals' spinal cords. "We found that a number of nerve fibers did grow back in the spinal cord and that the rats were able to walk better than without the treatment," says Strittmatter.
No drug has been developed to promote axon recovery in humans, so it's difficult to predict how well this peptide will work in people, Strittmatter cautions. Nor have any toxicology studies been conducted on NEP1-40. "Before moving to human trials, researchers must first determine whether this synthetic peptide can promote nerve fiber growth in animals weeks and months after injury and whether the compound is effective and safe for human use," he says.
Because damaged nerve fibers in the brain and spinal cord remain at the site of an injury, scientists have some reason to believe that NEP1-40 might promote growth in older injuries. "If we had some way to block these nerve regeneration inhibitors, the damaged fibers might grow back again," says Strittmatter. "Our findings thus suggest that this peptide might have significant therapeutic potential."
Experiments recently conducted by the laboratory of Ray Lund, PhD, of the University of Utah Health Science Center may lead to new therapies for two serious eye conditions that can cause blindness: age-related macular degeneration and retinitis pigmentosa.
More than 10 million Americans currently have age-related macular degeneration, a condition that affects central vision and that is the leading cause of blindness in people over the age of 55. Retinitis pigmentosa, a group of inherited diseases that affect the retina and that result in a progressive loss of vision, afflicts about 100,000 Americans.
Working in collaboration with the laboratory of Glen Prusky, PhD, at the University of Lethbridge in Alberta, Canada, Lund's team has found that transplantations of a human pigment epithelial cell line can limit the deterioration of vision in rats with retinal degenerative disease.
"You can't ask a rat how well it can see, so we had to devise a method of assessing their visual performance," Lund says. His lab did this by developing a visual water task, a combination of a typical alternate choice box and the Morris water maze. "The test is very accurate discriminator of visual performance," says Lund.
For their experiments, Lund and his colleagues used RCS rats, the most widely studied animal model of retinal degeneration. Human cell line cells were transplanted into the eyes of three-week-old rats. When the animals were six months of age or older, their vision was assessed using the visual water task. The cell-grafted rats as a group performed more than twice as well on the test as the sham-injected rats, but only half as well as sighted control rats.
"Our study also showed that cell lines can be used as a potential source of cells for transplantation, which obviates many of the logistical and ethical problems associated with using fresh fetal cells," Lund says.
Lund and his researchers intend to try similar experiments in larger animal models in preparation for human clinical trials.
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A single mutation in a single gene is enough to slowly rob people of their ability to walk, scientists from the University of Michigan and the University of Pennsylvania report today.
And while the inherited defect itself is rare, its discovery may help researchers unravel the mysteries of much more common paralyzing conditions, from spinal cord injury to Lou Gehrig's disease.
In a paper published online today in the American Journal of Human Genetics, and scheduled for the journal's October issue, the U-M and Penn team describes a new gene, called NIPA1, for a form of hereditary spastic paraplegia, or HSP.
HSP is the name given to a group of disorders affecting about 20,000 Americans. HSP gradually disables its victims as long nerve cells in the spinal cord degenerate and muscles weaken and become spastic. It is often misdiagnosed as other nerve disorders, including multiple sclerosis, cerebral palsy, and amyotrophic lateral sclerosis (also called Lou Gehrig's disease).
There is no cure for HSP, which is also sometimes called familial spastic paraparesis or Strumpell-Lorain disease. Treatment is limited to physical therapy and exercise to help retain as much muscle function as possible, drug treatment to tame spastic muscle movements, and medication to treat patients' bladder and bowel control problems, and depression.
"There are direct overlaps between this group of diseases and the nerve degeneration processes in other disorders," says senior author John Fink, M.D., professor of neurology at the University of Michigan Medical School, who has devoted his career to studying and treating HSP and related conditions. "Not only do we hope this discovery will aid HSP patients, but we also believe it will aid in understanding and perhaps stopping the fundamental processes involved in other types of spinal cord degeneration."
The newly described gene mutation on chromosome 15 causes a form of HSP that starts in adulthood, and is passed on in families in a pattern called dominant inheritance. The paper's co-lead authors are U-M research investigator Shirley Rainier, Ph.D., and Penn postdoctoral research fellow Jing-Hua Chai, M.D., who works in the laboratory of co-author Robert D. Nicholls, D.Phil.
The discovery of the mutation was made with the help of two families affected by the disease: one, of Iraqi origin living in Michigan, and the other of Irish descent, living in Arkansas. Affected and unaffected members of both families allowed their DNA to be studied and compared with DNA samples from people without HSP.
The single-nucleotide mutation alters the structure of a protein that the researchers think resides in nerve cell membranes, a feature that may expedite the development of tests and therapies. In addition to further research on the protein's exact function, this gene discovery could be used to develop a genetic test to help give HSP patients and their families a firm diagnosis and genetic counseling. A provisional patent application has been filed jointly by the U-M and Penn.
The U-M team has previously found other genes involved in HSP, including a mutation responsible for a form of HSP that develops in early childhood and may resemble familial cerebral palsy.
That discovery, of a gene on chromosome 14 that encodes a protein called atlastin, has already yielded a diagnostic test that can give patients a firm diagnosis to tell them if they have or might some day develop that form of HSP. It can also help couples whose family or medical history suggests they might be at risk of passing a dominant form of HSP on to their children; in dominant HSP families, there is a 50 percent risk that each child will develop the condition.
Developed by Athena Diagnostics, which licensed U-M's patent on the atlastin gene, the test also screens for a form of HSP found by French scientists, based on a gene for a protein called spastin. Fink has no direct financial interest in Athena Diagnostics.
Fink and Rainier have been looking at chromosome 15 for nearly eight years, after discovering that this region contained a mutant HSP gene. Rainier had narrowed the search to a very small region of chromosome 15, and carefully analyzed genes in this region as potential candidates. Meanwhile, Nicholls and his lab had been searching for genes in the same stretch of DNA.
Sharing of information and materials between the two laboratories resulted in the discovery that NIPA1 mutations cause this particularly severe form of HSP. In their paper, the researchers describe the gene and its encoded protein, as well as clinical analysis of affected subjects. The participation of 105 elderly men and women from a U-M Institute of Gerontology registry enabled this study, allowing the investigators to determine the range of normal variations of the new gene.
The affected members of the Iraqi family living in Michigan and the family from Arkansas are among the nearly 400 HSP patients that Fink and his U-M colleagues see each year. The U-M Neurogenetic Disorders Clinic is the largest clinical and research program for HSP and related disorders in the nation, and one of few that offer comprehensive evaluation, including genetic counseling.
The two families are not related, so the researchers believe that the same mutation occurred spontaneously in both, and likely causes the disease in other families and even in patients who have no family history of the disease.
Although more study is needed to understand exactly how the single-nucleotide mutation leads to a dysfunctional protein and thereby to the disabling symptoms of HSP, Fink and his colleagues feel that they have a good chance of unraveling the mystery. They're working to develop a mouse model of this form of HSP, which will enable further studies of potential drug- and cell-based therapies.
Until this research yields new treatment options, the U-M and a few other centers offer treatment for HSP's effects, and genetic testing and counseling. Support groups and a new Spastic Paraplegia Foundation bring patients and researchers together locally and nationally. Fink serves as the SPF's medical advisor and last year hosted the first International Symposium for HSP.
###
Contact: Kara Gavin
kegavin@umich.edu
734-764-2220
University of Michigan Health System
New hope for the paralysed as scientists re-grow spinal cords (Filed: London Telegraph 13/07/2003)
Thirty years of research culminate in a 'Eureka!' moment for a French biologist working to regenerate severed nerve endings, reports Kim Willsher in Paris.
French scientists have discovered a way to regenerate damaged spinal cords in a breakthrough that might eventually allow paralysed people to walk again.
They have succeeded in regrowing the broken spinal cords of mice after identifying and eliminating two proteins that create scar tissue. This had prevented nerve endings from repairing themselves.
When the scientists, from the French National Health and Medical research institute, manipulated the genes of the mice so that they did not produce these proteins, they discovered that the spinal cord regrew within weeks, enabling the animals to run around again.
Until now the regeneration of nerve cells - or neurones - in the brain and the spinal column has been considered impossible. Previous attempts to restore locomotion by connecting the two extremities of a severed spine have always failed. The research team is now working on ways of using the discovery to develop a treatment for humans.
Prof Alain Privat, 59, who led the team of biologists from the Languedoc University of Science in Montpellier, in the south of France, told The Telegraph yesterday: "It's a very important breakthrough because for the past 100 years researchers working on the regeneration of the central nervous system have been unable to crack this problem. Nobody envisaged being able to repair the system because for a very long time they thought it was something inherent to the fibres themselves that meant they couldn't grow back and restore movement.
"Today, after six years of research, we are sure that we can make them work again if we can stop these two proteins from being produced."
The team is now working on adapting the process so that it can be applied first to mice and monkeys whose genes have not been modified, before carrying out tests on human patients. The key will be to find ways to inhibit the two proteins that stop the cord from repairing itself.
"We've proved that it is possible, but it's going to take at least five years to find out if the technique can be applied to humans. We can produce mutant mice but not mutant people, so we have to find other ways of tackling these proteins," said Prof Privat.
"I cannot be 100 per cent sure it will work on humans but I think the chances are very reasonable. It should work but until we do it we won't be sure."
The next stage involves developing a "genetic treatment" that could be applied to a damaged spine to halt the scarring process and stimulate the regrowth of the nerves. The researchers are also looking at whether the regeneration of the neurones can be jump-started in older injuries, enabling paralysed people such as Christopher Reeve, the actor, to benefit.
Prof Privat believes that the technique could also be used to treat brain injuries and degenerative conditions such as Parkinson's disease. "The neurons in the brain are very similar to those in the spinal cord," he said.
The professor, who is the director of the national health and medical research institute at Languedoc University, has worked with a team of three researchers to try to crack the secret of repairing spinal cord neurones.
The team identified two key proteins, Vimentine and Glial Fibrillary Acidic Protein (GFAP). These proteins, they discovered, were produced in large quantities after a spinal injury and formed a scar, described as an impenetrable wire fence, around the damaged nerve endings, preventing messages being transmitted to the brain.
Under his guidance, his researchers took 100 ordinary laboratory mice and divided them into four groups. The first was left alone as a control set, the second and third were genetically altered to eliminate one of the proteins, and the last group was "double mutants", lacking the genes that produced both proteins. The mice were all tested for their ability to run on a treadmill before their spines were cut.
Prof Privat, 59, said: "After two or three weeks there was a small group of animals that was performing better than the others. After four weeks they were moving their back legs and most of their motor function appeared to have been restored.
"We discovered that the control group and those whose genes had been only partially manipulated remained permanently paralysed. However, the mice in which both the proteins had been knocked out had recovered the use of their legs.
"When we examined these mice we found that there were no scars on the spinal cord fibres, the nervous system circuitry had been reconstructed and that after four weeks there was almost no remnants of paralysis in their hind legs.
"Thanks to this genetic modification the scarring that seems to be the cause of the paralysis didn't happen and the nerve fibres were able to grow back.
"He recalled the moment he and his team realised that they had achieved the breakthrough. "It took just a few seconds for us to realise that we had finally done it."It was a real 'Eureka!' moment and I jumped into my office where I had a very old bottle of my favourite Scotch, Glenlivet. I said to the others, "This is the time to crack it open'," Prof Privat added.
"It is absolutely ground-breaking. I started working on this subject around 30 years ago and I had finally done it."
One of the first problems he faces, however, is finding the estimated Ł3.5 million ( *5 million) needed to continue the research. Until now, the Montpellier team, hit by French government budget cuts, has relied on cash from private associations dedicated to those paralysed by spinal injuries. He said: "We need a relatively small sum of money. We have these encouraging results but barely the means to go on."
The results of the team's research will be published by the National Academy of Sciences, based in Washington, this month.
The Czech doctors have tried to perform an unique surgery
The doctors from the Prague Teaching Hospital Motol carried out an unique surgery. They have transplanted stem cells into spinal cord of a young man who was totally paralyzed after severe cervical spinal cord injury. These stem cells were obtained from patient's own bone marrow and placed in his bloodstream. It is expected that the cells will help recover and form a new nerve tissue at the injury site.
In terms of spinal cord injury treatment, this is one of first surgical procedures being ever done in the world and the first of its kind in the Czech Republic. Stem cells are a modern "hit" in medicine. Last July, the doctors from General Teaching Hospital in Prague treated, in like manner, a patient with very damaged heart tissue after severe heart attack.
"We originally intended to try the treatment in the next coming fall", says Prof. Eva Sykova, a directress of Experimental Medicine Department and project leader.
"But, as we were asked for help by patient's family, we just tried to help." Prof. Sykova added that this kind of treatment is, so far, appropriate for acute injuries – within two or three weeks since injury.
Several professional medical teams in collaboration with Cell Therapy Center and Tissue Replacements at Charles University and Experimental Medicine Department of Academy of Sciences participated in this unique surgery.
Up to the present time, the doctors have been almost powerless in the battle against spinal cord injuries. Therefore, the treatment using stem cells represents a hope. In the bone marrow can be found a quantity of cells which, if transfered into different tissue, can form quite identical cells – the cells which are contained in the mentioned tissue.
Other cells produce growth factors needed for cell regeneration in damaged tissue. Patient is a donor and there is no reason to be anxious about rejection of these cells.
As this is first procedure of its kind in the nation (and one of the first in the world), the researchers and clinicians were obliged to quickly write a clinical study, submit it to ethical commission for approval and elaborate stages of therapy.
Everything took place within 13 days. "We were able to start using stem cells thanks to the fact that a medical team from Motol was ideally prepared," emphasized Prof. Sykova.
The procedure consisted in several steps: in the intensive care unit in Motol, the clinicians working at 2. Children Hospital removed bone marrow from a patient and team from Department of Hematology and Blood Transfusion isolated cells from it.
Other specialists put them into patient's body. After this, the patient was examined by neurologists to find out if the treatment is successful.
"He is doing well," said Eva Sykova. "However, we will be able to observe any effect of this treatment earliest after several weeks, but rather in several months or a year
Curis Compound Stimulates Replication of Brain Progenitor Cells; Findings Suggest Strategy for Regeneration of Tissues in Damaged Brains
CAMBRIDGE, Mass., Sept. 11, 2003--Curis, Inc. (NASDAQ: CRIS) today announced that the current issue of the scientific journal Neuron contains a report demonstrating that the Hedgehog signaling pathway is required for the formation and maintenance of brain progenitor cells. These progenitor cells are the precursors of the nerve cells and support cells that populate the brain. This report also shows that orally administered small molecule agonists developed by Curis can activate the Hedgehog pathway and induce new progenitor cell formation in the brain and, as result, may stimulate normal regenerative processes and promote brain recovery from injury.
The report, entitled "Sonic Hedgehog is Required for Progenitor Cell Maintenance in Telencephalic Stem Cell Niches," was published online yesterday in the journal Neuron. The research was performed by laboratories at the New York University Medical Center, Harvard University, Goteborg University, and Curis, Inc. The authors conclude that use of Hedgehog agonists may constitute a new therapeutic approach to the repair of brain damage and that "this strategy may allow the body's own developmental pathways and resident progenitor cells to be utilized for regenerative therapies."
Dr. Lee Rubin, Curis' Chief Scientific Officer, said, "The ability of Curis' Hedgehog agonists to promote the development of new nerve cells represents an important advance in our understanding of the potential mechanisms by which these drug candidates exert their reparative/regenerative effects. We have already observed that stimulation of the Hedgehog pathway by agonists can minimize tissue damage in response to injury, such as stroke. These new data and our own internal research suggest a broader, perhaps more fundamental, role in promoting tissue regeneration by stimulating the formation of new nerve cells within the brain itself."
Daniel Passeri, Curis' President and Chief Executive Officer stated, "This new report adds to the increasingly large body of evidence that validates the Hedgehog pathway as a significant target for the development of new drug candidates for neurological disorders. Recently, we have seen that Curis' Hedgehog agonist drug development candidates have shown efficacy in models of Parkinson's disease, diabetic neuropathy, and stroke. The potential of being able to use these Hedgehog agonists to induce the brain to form new nerve cells opens the door to many new therapeutic opportunities."
About Curis, Inc.
Curis, Inc. is a therapeutic drug development company. The Company's technology focus is on regulatory pathways that control repair and regeneration. Curis' product development involves the use of proteins or small molecules to modulate these pathways. Curis has successfully used this technology and product development approach to produce several promising drug product candidates in the fields of kidney disease, neurological disorders, cancer, and alopecia (hair loss). For more information, please visit the Curis web site at www.curis.com.
This press release contains forward-looking statements within the meaning of the Private Securities Litigation Reform Act of 1995. Statements concerning Curis' or management's intentions, plans, expectations or predictions of future events are forward-looking statements. Such statements may contain the words "believes", "expects", "anticipates", "plans", "estimates" or similar expressions. Forward looking statements involve known and unknown risks, uncertainties and other factors that may cause the actual results to be materially different from those indicated by such forward-looking statements. Actual results can be affected by, among other things, uncertainties relating to product development, clinical trials, regulatory actions or delays, the ability to obtain or maintain patent or other proprietary intellectual property protection, changes in or an inability to execute Curis' realigned business strategy and other risk factors identified in Curis' most recent Annual Report on Form 10-K and subsequent reports filed with the Securities and Exchange Commission. Curis disclaims any intention or obligation to update any of the forward-looking statements after the date of this press release.
Contacts
Curis, Inc. Christopher U. Missling, Ph.D., 617-503-6587 Chief Financial Officer Marc F. Charette, Ph.D., 617-503-6629 Vice President, Corporate Communications
J Neurotrauma. 2003 Jan;20(1):1-16. Related Articles, Links
Delayed transplantation of olfactory ensheathing glia promotes sparing/regeneration of supraspinal axons in the contused adult rat spinal cord.
Plant GW, Christensen CL, Oudega M, Bunge MB.
The Chambers Family Electron Microscopy Laboratory, The Miami Project To Cure Paralysis, Miami, Florida, USA. gplant@anhb.uwa.edu.au
The aim of this study was to determine the preferred time and environment for transplantation of olfactory ensheathing glia (OEG) into the moderately contused adult rat thoracic spinal cord. Purified OEG were suspended in culture medium with or without fibrinogen and injected into the contused cord segment at 30 min or 7 days after injury. Control animals received a contusion injury only or injection of only medium 7 days after contusion. The effects on axonal sparing/regeneration and functional recovery were evaluated 8 weeks after injury. The grafts largely filled the lesion site, reducing cavitation, and appeared continuous with the spinal nervous tissue. Whereas in 7d/medium only animals, 54% of spinal tissue within a 2.5-mm-long segment of cord centered at the injury site was spared, significantly more tissue was spared in 0 d/OEG-medium (73%), 0 d/OEG-fibrin (66%), 7 d/OEG-medium (70%), and 7 d/OEG-fibrin (68%) grafted animals. Compared with controls, the grafted animals exhibited more serotonergic axons within the transplant, the surrounding white matter, and the spinal cord up to at least 20 mm caudal to the graft. Retrograde tracing revealed that all but the 0 d/OEG-fibrin graft promoted sparing/regeneration of supraspinal axons compared with controls. Overall, the 7 d/OEG-medium group resulted in the best response, with twice as many labeled neurons in the brain compared with 7 d/medium only controls. Of the labeled neurons, 68% were located in the reticular formation, and 4% in the red, 4% in the raphe, and 5% in the vestibular nuclei. Hindlimb performance was modestly but significantly improved in the 7 d/OEG-medium group. Our results demonstrate that transplantation of OEG into the moderately contused adult rat thoracic spinal cord promotes sparing/regeneration of supraspinal axons and that 7 d transplantation is more effective than acute transplantation of OEG. Our results have relevant implications for future surgical repair strategies of the contused spinal cord.
August 29, 2003-- This past summer, the Tulane University Center for Gene Therapy received two grants totaling over $3 million to develop various stem cell research projects.
This funding, from the National Institute of Health, comes in the face of an expanding body of knowledge on both stem cells and their evident potential to repair damaged tissues.
Ultimately, if laboratory protocols are refined, it appears stem cell therapy could become an effective treatment for a vast array of terminal conditions such as chronic heart disease, Alzheimer's, Parkinson's and liver, kidney and lung diseases.
While stem cells were found over 100 years ago, research on tissue reparation is relatively new. Investigation into this area has only occurred in the past 20 years. Therefore, scientists still do not have a clear understanding of which stem cells relate to which disease, making centers such as Tulane's necessary.
Tulane's Center for Gene Therapy was established in July 2000. It was initially run by Dr. Darwin Prockop and a group of 14 others.
Two major projects are now being conducted under the recent grants given by NIH. The first is an initiative that prepares stem cells for multiple research purposes. Prockop and a team of Tulane University graduate students are setting up a stem cell distribution center where they will send out standardized stem cells to various national and international affiliate scientists. The distribution center will offer adult human and rat stem cells.
The second project is being conducted in affiliation with William Chilean, a co-principal investigator at LSU Health Sciences Center. This project works to better characterize heart disease by developing models for the condition using stem cells from the patients' own bone marrow. As a part of this project, the researchers will also induce heart attacks in rats by tying off their coronary arteries and then inject stem cells into the rats' systems to examine the effect. These experimental trials will provide insight to see if stem cell therapy could be valuable to patients with heart conditions.
Prockoff was hesitant to make any presumptions about where the research might lead. He did acknowledge that stem cells have a remarkable capacity to rebuild tissues by targeting damaged cells, and this capacity could be cultivated to treat patients with damaged tissues whose bodies simply cannot repair the damages quickly enough.
Therapy for a patient would consist of taking a sample of a patient's bone marrow and using it to make more of the patient's own stem cells. "The form of stem cell therapy we are interested in is merely accelerating the body's natural healing process," Prockop said.
Prockop's group is currently planning a clinical trial to treat patients with spinal cord injuries. It is scheduled to take place in June of 2004.
As for the long-term future goals of the center, Prockop said, "We wish to cure the diseases."
Treatment of chronic spinal cord injury in the rat with grafts of autologous pre-ligated peripheral nerve tissue and neurotrophic factors
A growing body of evidence suggests that Schwann cells (SCs) and neurotrophic factors can facilitate regeneration of axons in acute models of spinal cord injury. However, little is known about whether they are effective in promoting axonal regeneration in chronic spinal cord injury. We aimed to evaluate the effects of grafting pre-ligated saphaneous nerves together with a cocktail of neurotrophic factors on the regeneration of corticospinal neurons and functional recovery (BBB score) in a contusion model of chronic spinal cord injury in rats. The spinal cord was injured with a New York University type weight drop apparatus (10 gm x 12.5 mm) in 76 female adult (8 week) rats. At 3 weeks after injury, the resulting cavity was injected with either vehicle, minced pre-injured nerve or minced pre-injured nerve plus neurotrophic factors (BDNF, NT3 and GDNF). Behavioral tests (BBB score) and anterograde tracing was performed before sacrifice two or six months after graft. There was a significant difference between BBB scores of the groups, with rats receiving pre-ligated nerve plus neurotrophic factors having the highest scores. Tracing results provided evidence of significant regeneration of corticospinal tracts in rats treated with the pre-ligated peripheral nerves and neurotrophic factors. We conclude that a combination of pre-ligated nerve tissue and neurotrophic factors may be a promising approach for the treatment of chronic spinal cord injury.
Localization of the precursor for brain-derived neurotrophic factor in the peripheral and central nervous system
The precursors for neurotrophins are proteolytically cleaved to form biologically active mature molecules which activate their receptors p75NTR and trks. A recent study showed that the precursor for NGF can bind to p75NTR with a high affinity and induces apoptosis of neurons in vitro. However, the functions of precursors for neurotrophins in vivo are not known. To examine the functions of pro-neurotrophins in vivo, it is essential to know where they are expressed in the nervous system. Our previous studies showed that BDNF is widely distributed in the nervous system and anterogradely transported by many groups of neurons. In the present study, we have raised and characterized rabbit polyclonal antibodies against a peptide coding for the precursor region of the BDNF gene. The antibody specifically recognizes the precursor for BDNF but not other proneurotrophins by Western blot. With the affinity purified precursor antibody, we have mapped the distribution and localization of the precursor for BDNF. Preliminary results showed that, like mature BDNF, pro-BDNF is localized to nerve terminals of sensory neurons in the superficial layers of dorsal horn and peripheral tissues. The nerve terminals immunoreactive for pro-BDNF were also widely distributed in the brain, with the pattern similar to mature BDNF. These results suggest that pro-BDNF may be released from nerve terminals in the spinal cord and brain. The function of pro-BDNF in vivo remains to be determined.
Neurotrophins is required for the enhanced regeneration of ascending neurons after a conditioning nerve lesion
A conditioning sciatic nerve lesion can enhance regeneration of the injured spinal cord but the mechanisms are not known. A peripheral nerve lesion could trigger up-regulation of neurotrophins in neurons and glia in the DRG. We hypothesize that endogenous neurotrophins play important roles in the enhanced axonal regeneration. We tested this hypothesis in vivo with antisera to neurotrophins. The left sciatic nerves in adult Sprague-Dawley rats were ligated and cut under anaesthesia with Halothane. One week later the dorsal column was crushed and a piece of gel foam soaked with the antisera to NGF, BDNF, NT3 or normal sheep serum (NSS) was put on the crush site. The animals were i.p. injected with the respective antiserum or NSS twice a week (10 µl/g). The regeneration of ascending sensory neurons were assessed by anterograde and retrograde tracing techniques one week after the cord injury. In NSS treated rats, regenerating fibres were found in and rostral to the cavity. However, antisera to neurotrophins prevented the growth of sensory axons into the cavity. The retrograde labelling data showed that 45.44 ± 9.58 neurons were labelled in the ipsilateral DRG of rats treated with NSS. The number of labelled neurons in the ipsilateral DRG was significantly reduced in the rats treated with antisera to neurotrophins (to NT3: 6 ± 3.23; to BDNF: 5 ± 2.98; to NGF: 5.5± 2.71). These results suggest that endogenous neurotrophins play an important role in the enhanced nerve regeneration after a conditioning lesion.
Anterograde and retrograde transport of the precursors for neurotrophins in the rat sciatic nerve
Neurotrophins are growth factors that promote proliferation, differentiation and survival of cells in the developing nervous system. They are synthesized as precursors that are proteolytically cleaved to mature biologically active neurotrophins. And the regulation of the proteolytic process may mediate the proapoptotic or prosurvival effects of neurotrophins. Our studies suggest that endogenous brain-derived neurotrophic factor(BDNF) is transported both anterogradely and retrogradely, whereas endogenous neurotrophin-3 display exclusive retrograde transport. To address the active transport of precursors of neurotrophins, we used a double ligation(DL) procedure that distinguishes between anterograde and retrograde flow to visualize the axonal transport of endogenous precursor of BDNF(pro-BDNF) and NT-3(pro-NT-3).Adult Sprague-Dawley rat was done sciatic nerve double ligation on middle thigh level under anaesthesia with helathone. Rat was allowed to survive for 18h before perfusion with 4% paraformaldehyde under an overdose of anaesthesia with pentobarbital. Sciatic nerves were cryostat sectioned and pro-BDNF and pro-NT-3 immunohistochemistry were performed. Pro-BDNF immunoreactivity was accumulated on both the proximal and distal to the DL in the intact rat sciatic nerve, whereas pro-NT-3 immunoreactivity was predominantly accumulation in the proximal side. These studies demonstrate that pro-BDNF, like BDNF, is anterogradely and retrogradely transported in the periphery nerves but that pro-NT-3 is anterogradely transported from the neuronal cell bodies to the periphery terminal. Our results suggest that neurotrophin precursor may play a role in the periphery nerve response to the injury.
Neuroregeneration Laboratory
This group focuses on neurotrophic factors and their roles in survival and regeneration of normal and injured nerves. Neurotrophic factors are essential for the generation and survival of nerve cells during development. They can be potential drugs for the treatment of neurodegenerative diseases such as Alzheimer’s and Parkinson’s diseases or nerve injuries including spinal cord injury and stroke. We are interested in their roles in neuropathic pain, cell survival and nerve regeneration after nerve injury. We use a variety of techniques including biochemical, immunological, molecular and histological approaches in our research projects.
Investigators
Xin-Fu Zhou MSc., PhD
Li Li MSc., PhD
Visiting Research Fellows / Scholars
Shi-Qing Feng, MBBS,PhD, Associate Professor, Orthopaedics, Tianjin Medical University Hospital. Tianjin, China
Hui Wang, MBBS, MSc, Lecturer, Department of Anatomy and Neurobiology, Xiangha Medical
College, Central South University of China, Hunan, P. R. China.
* Treatment of chronic spinal cord injury in the rat with grafts of autologous pre-ligated peripheral nerve tissue and neurotrophic factors
* Localization of the precursor for brain-derived neurotrophic factor in the peripheral and central nervous system
* Neurotrophins is required for the enhanced regeneration of ascending neurons after a conditioning nerve lesion
* Anterograde and retrograde transport of the precursors for neurotrophins in the rat sciatic nerve
Collaborative Research
R.A. Rush, Department of Human Physiology, Flinders University.
Bing-Ren Huang, Chinese Academy of Medical Sciences and Beijing Union Medical College, Bejing, China.
Xue-Gang Luo, Department of Anatomy, Hunan Medical University, Changsha, China.
Tinghua Wang Neuroscience Institute, Kuming Medical College, Kuming, P.R. China
Publications
Wang H. and X-F Zhou. (2002) Injection of brain-derived neurotrophic factor in the rostral ventrolateral medulla increases arterial blood pressure in rats. Neuroscience, 112:967-975
Zhao SP, Zhou XF. (2002) Co-expression of trkA and p75 neurotrophin receptor in extracranial olfactory neuroblastoma cells. Neuropathol Appl Neurobiol. 28(4):301-7
Li L, Xian CJ, Zhong JH, Zhou XF. (2002) Effect of lumbar 5 ventral root transection on pain behaviors: a novel rat model for neuropathic pain without axotomy of primary sensory neurons. Exp Neurol. 175(1):23-34
Townley SL, Grimbaldeston MA, Ferguson I, Rush RA, Zhang SH, Zhou XF, Conner JM, Finlay-Jones JJ, Hart PH. (2002) Nerve growth factor, neuropeptides, and mast cells in ultraviolet-B-induced systemic suppression of contact hypersensitivity responses in mice. J Invest Dermatol. 118(3):396-401
Li Li, and Zhou X-F: (2001) Pericellular Griffonia simplicifolia I isolectin B4-binding ring structures in the dorsal root ganglia following peripheral nerve injury in rats. J Comp Neurol. 439(3):259-74.
Xian, C.J., Zhou, X.F., (2001) Sensitive and non-radioactive in situ detection of neurotrophin mRNAs in the nervous system. Methods Mol Biol. 169,91-98.
Luo X-G., Rush R.A. and Zhou X-F. (2001) Ultrastructural localization of brain-derived neurotrophic factor in rat primary sensory neurons. Neurosci. Res., 39: 377-384.
Xian CJ, L Li, Y-S Deng, S-P Zhao, X-F Zhou. (2001) Lack of effects of transforming growth factor-alpha gene knockout on peripheral nerve regeneration may result from compensatory mechanisms. Experimental Neurology 172(1):182-8.
Lu J, X-F Zhou, RA Rush. (2001) Small Primary Sensory Neurons Innervating Epidermis and Viscera Display Differential Phenotype in the Adult Rat. Neurosci. Res. 41:355-63.
Chie E, Liu D, Zhou XF, Rush RA (2001) Quantification of neurotrophin MRNA by RT-PCR. Methods Mol Biol 169:81-90
Zhang SH, Zhou XF, Rush RA (2001) Extraction and quantification of the neurotrophins. Methods Mol Biol 169:31-41
Deng Y-S, Zhong J-H and Zhou X-F: (2000) BDNF is involved in sympathetic sprouting in the dorsal root ganglia following peripheral nerve injury in rats. Neurotoxicity Research 1: 311-322.
Huang B-R, Gu J-J, Ming H, Lai D-B and Zhou X-F: (2000) Differential actions of neurotrophins on apoptosis mediated by the low affinity neurotrophin receptor p75NTR in immortalized neuronal cell lines. Neurochemistry International 36:55-65.
Zhou X-F, Deng Y-S, Xian CJ and Zhong J-H: (2000) Neurotrophins from dorsal root ganglia trigger allodynia after spinal nerve injury in rats. European Journal of Neuroscience 12: 100-105.
Deng Y-S, J-H Zhong and Zhou X-F: (2000) Effects of endogenous neurotrophins on sympathetic sprouting in the dorsal root ganglia and allodynia following spinal nerve injury. Experimental Neurology 164: 344-350.
Li L, Deng Y-S, and Zhou X-F: (2000) Down-regulation of TrkA in primary sensory neurons in response to peripheral nerve injury in the rat. Neuroscience Research 38: 183-191
Zhang J-Y, Lo X-G, Xian C, Liu Z-H and Zhou X-F: (2000) Endogenous BDNF is required for myelination and regeneration of injured sciatic nerve in rodents. European Journal of Neuroscience 4171-4180.
Xian C and Zhou X-F: (2000) Roles of transforming growth factor alpha and related molecules in the nervous system (Invited review). Molecular Neurobiology 20:157-183.
Ferguson IA, Lu JJ, Zhou X-F and Rush RA: The low affinity neurotrophin receptor, p75: a multifunctional molecule with a role in nerve regeneration? In Degeneration and Regeneration in the Nervous System. PP221-237, Ed NR Saunders and KM Dziegelewska. Harwood Academic Publishers, Australia (2000)
Scientists used the chemical in laboratory tests
Scientists have found a chemical which can influences the direction of nerve cell growth - and say it could help repair damaged spinal cords.
One of the reasons that severe spinal cord injury causes permanent damage is that the nerve cells involved do not regrow in the same way as normal cells.
Scientists are hopeful that there must be naturally-produced chemicals which encourage these "axons" to grow out and bridge the gaps caused by injury or disease.
However, these chemicals have remained elusive.
Now a team of scientists from Johns Hopkins University in Baltimore, US, has found, by chance, a chemical which appears to be able to influence nerve growth.
When they put cells next to a source of the substance, called semaphorin-7a, there was more cell growth in the direction of the source than in other directions.
Mice bred to lack semaphorin-7a had brains which did not develop properly.
Future hopes
The scientists believe that if they can harness this chemical, and others like it which have the opposite effect, it might one day be possible to encourage spinal injuries to heal more efficiently.
Professor Alex Kolodkin, one of the researchers, said the find was impressive but that much work remained to be done.
"I've been studying semaphorins for about a decade and didn't expect to find any that stimulated axon growth, certainly not to extent we saw in the lab and in mice.
"Now we need to figure out how semaphorins balance their repulsive and attractive effects."
Specific aim
He said that it might be problematic to encourage growth in one type of cells - nerve cells - without getting unwanted growth elsewhere.
His colleague Dr Jeroen Pasterkamp, who co-authored the research, published in the journal Nature, said: "Our next steps are to find otu exactly how semaphorin-7a's message is passed along inside the nerve, which will hopefully reveal a useful, specific target for promoting axon growth following nerve injury or degeneration."
Dr Patrick Mehlen, from the University of Lyon in France, said: "A deeper understanding of the intracellular signalling pathways that are activated by semaphorins will provide insight into a variety of physiological processes, from vascular and neural development to immunity."
Bathed in nutrients, but in the absence of any particular cue, axons will extend from cortex nerve cells in all directions (top image). Toss in a growth stimulator like semaphorin-7a, and axon growth is heavier nearest the cue (bottom image). Credit: Johns Hopkins Medical Institutions, Nature.
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Full size image available through contact
For decades, scientists have hunted for signals that guide nerve cells' tentacle-like axons, hoping to understand how these cell tips reach out to distant targets. It's knowledge that might one day help researchers learn how to rebuild nerves lost to spinal cord injuries or diseases like Huntington's.
Now, a Johns Hopkins team studying a family of proteins best known for repelling axons and inhibiting their growth reports finding one member that unexpectedly promotes axon growth instead. In their experiments, rat nerves in the lab grew more and longer axons on the side nearest a source of this protein, called semaphorin-7a. Moreover, in mice without semaphorin-7a, axons of some odor-sensing nerve cells never reached their targets, the scientists report in the July 24 issue of Nature.
"I've been studying semaphorins for about a decade and didn't expect to find any that stimulated axon growth, certainly not to the extent we saw in the lab and in mice," says Alex Kolodkin, Ph.D., professor of neuroscience in The Johns Hopkins University School of Medicine's Institute for Basic Biomedical Sciences. "Now we need to figure out how semaphorins balance their repulsive and attractive effects."
Part of the answer to this paradox, Kolodkin says, is that semaphorin-7a interacts with different proteins than its relatives. In experiments with rat nerve cells involved in sensing odors, first author and postdoctoral fellow Jeroen Pasterkamp, Ph.D., found that semaphorin-7a spurs axon growth by hooking onto proteins called integrins, which are found on nerves and many other cell types.
Among their many roles, integrins (pronounced IN-teh-grins) help control cells' interactions with their surroundings by capturing chemical signals and conveying the messages to cells' internal machinery. Even though this is the first report to link semaphorins and integrins, both protein families are rapidly being recognized as major contributors to neurological function and disease, says Kolodkin.
"Because integrins are important throughout the body, targeting them to stimulate axon growth or re-growth in a particular area of the brain or spinal cord presents many problems," notes Pasterkamp. "Our next steps are to find out exactly how semaphorin-7a's message is passed along inside the nerve, which will hopefully reveal a useful, specific target for promoting axon growth following nerve injury or degeneration."
As the researchers learn more of the specifics about how semaphorin-7a differs from its relatives, they also hope to redraw their picture of how semaphorins as a family affect nerve development throughout life, they say.
###
The research was funded by the Kirsch Foundation, the National Institute of Neurological Disease and Stroke (part of the National Institutes of Health), the Netherlands Organization for Scientific Research and the Human Frontier Science Program.
Authors on the paper are Pasterkamp and Kolodkin of Johns Hopkins, and Jacques Peschon and Melanie Spriggs of Amgen Corporation, Seattle, Wash.
Note to Editors: An image that shows the growth-stimulating effects of semaphorin-7a is available at http://www.hopkinsmedicine.org/press/2003/July/030723.htm
(CAPTION FOR IMAGE) Bathed in nutrients, but in the absence of any particular cue, axons will extend from cortex nerve cells in all directions (top image). Toss in a growth stimulator like semaphorin-7a, and axon growth is heavier nearest the cue (bottom image). Credit: Johns Hopkins Medical Institutions, Nature.
On the Web:
http://www.nature.com/nature
Johns Hopkins Medical Institutions' news releases are available on an EMBARGOED basis on EurekAlert at http://www.eurekalert.org, and from the Office of Communications and Public Affairs' direct e-mail news release service. To enroll, call 410-955-4288 or send e-mail to bsimpkins@jhmi.edu.
On a POST-EMBARGOED basis find them at http://www.hopkinsmedicine.org
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Nature 424, 711 (14 August 2003); doi:10.1038/424711a
Chinese fusion method promises fresh route to human stem cells
CARINA DENNIS
Biologists in China have reprogrammed human cells by fusing them with rabbit eggs emptied of their genetic material. And they have extracted stem cells, which have the potential to form a wide array of different cell types, from the resulting embryos.
C. DENNIS; SHANGHAI SECOND MEDICAL UNIV.
A team led by Huizhen Sheng (left) has devised a technique for reprogramming adult human cells by fusing them with empty rabbit eggs (above).
The researchers, led by Huizhen Sheng of Shanghai Second Medical University, think that these 'derived' stem cells could provide scientists in the field with an alternative to stem-cell lines extracted from human embryos. But some researchers who have seen the work point out that the derived cells don't seem to have the same ability as human embryonic stem cells to grow indefinitely in culture.
The work will be published online this week in Cell Research, a peer-reviewed journal supported by the Chinese Academy of Sciences. The paper will appear in print later this month (Y. Chen et al. Cell Res. 13, 251–263; 2003).
Sheng's work has already created a buzz after rumours of it circulated in the scientific community and were reported in The Wall Street Journal in March 2002 (see Nature 419, 334–336; 2002). The publication is likely to reignite debate over the ethics of cross-species reprogramming. But cell biologists say that having the data available for public discussion will help researchers and regulators to decide what kinds of cross-species work should be pursued.
Reprogramming adult cells to assume an embryonic state could offer a way to grow new cells and tissues to replace those lost to ageing and disease. And using an individual's own cells, in a process often called therapeutic cloning, may avoid problems of the immune system rejecting the cell therapy.
To reprogramme an adult cell, it can either be fused with or have its nucleus injected into an egg that has had its own nucleus removed. The reconstituted cell is then tricked into dividing as if it were an embryo, with all memory of its previous life as a liver, skin or kidney cell erased. After 5–7 days, embryonic stem cells — which can seemingly grow indefinitely — can be extracted from the growing ball of cells. By changing the growth conditions, these cells can then be coaxed to develop into many different cell types.
Until now, scientists have only been able to generate animal stem-cell lines from the reprogrammed nuclei of cells. "This is the first paper to convincingly show that you can get human reprogramming," says Robin Lovell-Badge, a cell biologist at the National Institute for Medical Research in London.
Sheng claims to have successfully reprogrammed cells from the foreskin tissues of males aged 5, 42 and 52 years and from the facial skin of a 60-year old woman. "It just goes to show that age doesn't matter," says Lovell-Badge.
But Sheng has had a tough time convincing some experts. "I first submitted the paper more than two years ago," she says. It is understood that the paper was considered and rejected by other journals before this week's publication in China.
"I was frustrated that it took so long to get the paper published," Sheng says, "and it still may take a while for people to accept the work. But the scientific community has the right to question the details of the work and we have a responsibility to respond to them."
Doug Melton a cell biologist at Harvard University, for one, has concerns about the nature of the derived embryonic stem-cell lines. "I'm convinced that the cells do have the capacity to differentiate into different cell types, but it's unclear how long the cells can grow in culture," he says.
"It would be very surprising if the cell lines were stable," Melton adds, noting that many interspecies hybrids are unstable, because of incompatibilities between the nucleus and mitochondria (the energy-producing compartments of cells) from different species.
And Rudolf Jaenisch, of the Whitehead Institute in Cambridge, Massachusetts, is not convinced that the derived cells meet the usual criteria for embryonic stem cells. "An important criterion is indefinite growth," he says. "This is not shown."
But reservations aside, Melton is pleased that the work has finally seen the light of day. "I'm glad to see it published as it will encourage others to try it," he says.
At this stage, Sheng has no plans to use stem cells created by her method to treat humans. "It is a research tool. But there is the possibility that if it were proved to be safe enough for clinical use, it could provide a solution to human egg shortages for reprogramming in the future," says Sheng.
While the scientific community debates her work, Sheng is keen to test the embryonic stem cells she has generated in an animal model. "It will be important to see whether they will be tolerated by the immune system and whether they can correct an animal model of human disease," she says.
Offtopic leszek, de a szintevesztes javito szemuveghez szolnek hozza. Volt szerencsem errol beszelgetni az egyik fejlesztovel. Mivel nagyon marginalisan de temamba vag a dolog nagyon felkleltette az en erdeklodesemet is. Nem ez az orszag tehet rola, hogy ez a szemuveg nem kerult be a gyakorlatba. A ceg amelyik fejlesztette rentgeteg penzt es idot es zsenialitast ultetett bele, de sajnos erdeklodes hianyaban a gyartast nem tudtak megkezdeni. Szamomra is rejtely, hzogy miert nem erdekelte ez a szemeszeket, kozlekedes es munkabiztonsagi szakembereket. Hangsulyozom nem a magyar egeszsegugy tehet rola, hogy ez igy van.
Bathed in nutrients, but in the absence of any particular cue, axons will extend from cortex nerve cells in all directions (top image). Toss in a growth stimulator like semaphorin-7a, and axon growth is heavier nearest the cue (bottom image). Credit: Johns Hopkins Medical Institutions, Nature.
For decades, scientists have hunted for signals that guide nerve cells' tentacle-like axons, hoping to understand how these cell tips reach out to distant targets. It's knowledge that might one day help researchers learn how to rebuild nerves lost to spinal cord injuries or diseases like Huntington's.
Now, a Johns Hopkins team studying a family of proteins best known for repelling axons and inhibiting their growth reports finding one member that unexpectedly promotes axon growth instead. In their experiments, rat nerves in the lab grew more and longer axons on the side nearest a source of this protein, called semaphorin-7a. Moreover, in mice without semaphorin-7a, axons of some odor-sensing nerve cells never reached their targets, the scientists report in the July 24 issue of Nature.
"I've been studying semaphorins for about a decade and didn't expect to find any that stimulated axon growth, certainly not to the extent we saw in the lab and in mice," says Alex Kolodkin, Ph.D., professor of neuroscience in The Johns Hopkins University School of Medicine's Institute for Basic Biomedical Sciences. "Now we need to figure out how semaphorins balance their repulsive and attractive effects."
Part of the answer to this paradox, Kolodkin says, is that semaphorin-7a interacts with different proteins than its relatives. In experiments with rat nerve cells involved in sensing odors, first author and postdoctoral fellow Jeroen Pasterkamp, Ph.D., found that semaphorin-7a spurs axon growth by hooking onto proteins called integrins, which are found on nerves and many other cell types.
Among their many roles, integrins (pronounced IN-teh-grins) help control cells' interactions with their surroundings by capturing chemical signals and conveying the messages to cells' internal machinery. Even though this is the first report to link semaphorins and integrins, both protein families are rapidly being recognized as major contributors to neurological function and disease, says Kolodkin.
"Because integrins are important throughout the body, targeting them to stimulate axon growth or re-growth in a particular area of the brain or spinal cord presents many problems," notes Pasterkamp. "Our next steps are to find out exactly how semaphorin-7a's message is passed along inside the nerve, which will hopefully reveal a useful, specific target for promoting axon growth following nerve injury or degeneration."
As the researchers learn more of the specifics about how semaphorin-7a differs from its relatives, they also hope to redraw their picture of how semaphorins as a family affect nerve development throughout life, they say.
###
The research was funded by the Kirsch Foundation, the National Institute of Neurological Disease and Stroke (part of the National Institutes of Health), the Netherlands Organization for Scientific Research and the Human Frontier Science Program.
Authors on the paper are Pasterkamp and Kolodkin of Johns Hopkins, and Jacques Peschon and Melanie Spriggs of Amgen Corporation, Seattle, Wash.
Note to Editors: An image that shows the growth-stimulating effects of semaphorin-7a is available at http://www.hopkinsmedicine.org/press/2003/July/030723.htm
(CAPTION FOR IMAGE) Bathed in nutrients, but in the absence of any particular cue, axons will extend from cortex nerve cells in all directions (top image). Toss in a growth stimulator like semaphorin-7a, and axon growth is heavier nearest the cue (bottom image). Credit: Johns Hopkins Medical Institutions, Nature.
On the Web:
http://www.nature.com/nature
Johns Hopkins Medical Institutions' news releases are available on an EMBARGOED basis on EurekAlert at http://www.eurekalert.org, and from the Office of Communications and Public Affairs' direct e-mail news release service. To enroll, call 410-955-4288 or send e-mail to bsimpkins@jhmi.edu.
On a POST-EMBARGOED basis find them at http://www.hopkinsmedicine.org
Rat Healing Raises Hope for Spinal Cure
Corinne Amoo
Reuters
July 11, 2003
Rat Healing Raises Hope for Spinal Cure
By Corinne Amoo
LONDON (Reuters) - Scientists said on Monday they were closer to finding a cure for people paralyzed by spinal cord damage, following the successful healing of an adult rat.
Pilot trials for human patients will begin within three years, research team leader Dr. Geoffrey Raisman told Reuters. "This is ground-breaking evidence because it will help people with spinal cord injury who have previously had no hope of recovery," he said.
The trials will involve transplanting cells from other areas of the body into the spinal cord so they can provide a bridge over the spinal cord blockage and provide it with a channel to repair itself.
Currently there is no cure for patients who suffer spinal and brain cord injuries, which depending on the severity of injury, leaves them paralyzed, without control over their bowels, bladder, blood pressure and with no sexual function.
Scientists healed the injured spinal cord of a rat by growing a bridge of olfactory nerve cells across scar tissue, which acted as a guide so the severed nerve fibres were able to find their way to the right targets in the rat's brain.
If the human pilot trials are successful, patients will be able to make important movements like reaching for and picking things up. The restoration of other functions like bladder control require further research, Raisman said.
The pilot trials will involve transplanting cells from the patients' nasal lining into the spinal cord.
"Until we carry out the tests in humans we do not know if it will work, but the organization of the rat's spinal cord and its control mechanisms that we have looked at are essentially similar to those found in humans," said Raisman.
The trials will take at least three years to be carried out because the research team needs to be sure that the cells used for repair are present and in the right numbers, he said.
Details of Raisman's findings are published in the June Journal of the Royal Society of Medicine.
Research to Be Presented Suggesting Reversal of Lou Gehrig's Disease May Now Be Possible
Annual Meeting: American Spinal Injury Association,
April 2-6, 2003 - Hotel Inter-Continental Miami, Florida
MIAMI, April 2 /PRNewswire/ -- Reversal of Lou Gehrig's disease may now be possible using thrombopoietin and thyroid hormone to cause regeneration of endogenous stem cells of the central nervous system. Evidence suggests spinal cord injury can be reversed through regeneration by stem cells. This research will be presented by George R. Schwartz, M.D., a senior researcher at Neuroregeneron Co., this week at the annual meeting of the American Spinal Injury Association held in Miami, Florida.
The Food and Drug Administration (FDA) biologics division approved a unique clinical trial in May 2002, authorizing the use of thrombopoietin for Lou Gehrig's disease (amyotrophic lateral sclerosis) in a 40-year-old mother of three small children whose clinical condition was deteriorating rapidly. She was more than 90% paralyzed, with minimal speech capability, tremendous difficulty swallowing, and rapidly failing respiratory function.
Top
The new treatment was approved for a clinical trial after it was demonstrated that platelet growth factors could be tremendously increased through use of thrombopoietin. Platelet growth factors act as stimulants for the growth and development of glial cells which act as repair cells for dying nerve cells. In addition, platelet growth factors can stimulate immature cells to differentiate into cells which act as neurons. Thyroid hormone was added to the trial after experimental evidence demonstrated that thyroid hormone acted as a signaling substance helpful for repair cells to function. Platelets were raised in cycles to more than 10 times the normal level resulting is blood serum rich in platelet growth factors. At day 42 of this clinical trial, this patient showed remarkably improved head and neck control and strength. At day 45, she exhibited improvement in tongue strength and motion with improved swallowing functions. As a result, a feeding tube was not necessary.
At day 60, increased leg muscle strength was clearly evident. Along with this motion, the patient was able to turn her arms and hands which had been paralyzed for more than a year. At day 110, she began to move her hands. Muscle strength throughout her body increased and her pelvic muscles could support more weight.
Top
The patient showed clear reversal of a previously deteriorating condition, and return of functions. Her downhill course stopped. The nerve cell regeneration and reversal of paralysis in this patient with Lou Gehrig's disease suggests that spinal cord injury and paralysis can also be treated with re-growth of the nerve cells of the spinal cord. Further trials are urgently needed since the average length of life in ALS patients is 3-5 years after diagnosis. There is also some indication that regeneration in cases of spinal cord injury would be more effective soon after the injury.
The FDA has been extremely supportive of this clinical trial and has urged that other trials be conducted as soon as possible. "If this proves out, it is a very exciting result indicating a new treatment and approach to Lou Gehrig's disease and spinal cord injury," said a senior neurologist with the FDA in Rockville, Maryland.
Top
However, despite the encouraging results and excitement generated by this clinical trial, the Genentech company has decided not to release the drug thrombopoietin for any further trials in neurologic disease or injury. "We will not proceed with any further trials at this time," said Mary Stutts, director of corporate relations at Genentech. The medication was manufactured in substantial quantity in the late 1990s and the current stock of clinical grade thrombopoietin will expire in the year 2003. "Remanufacture is not planned at this time," confirmed Heather Mccauley, spokeswoman at Genentech. She offered no other explanation for the decision not to conduct any further clinical testing.
Top
A director of the Lou Gehrig's clinic at the Massachusetts General Hospital and a professor at the Harvard Medical School, has prepared a trial for ten people with this disease. "I am totally puzzled," commented the doctor, who was rebuffed when he approached Genentech with his proposal. "This defies all common sense and scientific responsibility. We have no other treatments for these conditions," he explained.
Dr. Schwartz, who has been following his patient closely with the FDA approved trial, is also puzzled. "Are they blind to the implications of this drug for use in neurologic diseases or injury?" he remarked. Monica Collier, one of the researchers who has been following this patient's ground-breaking clinical course, expressed amazement at the lack of compassion shown by the spokespeople at Genentech. "I cannot understand their approach," she said. "It would seem to be in their interest to try to develop this medication, and the patients just cannot wait."
Top
Dr. Schwartz added, "I know the people at Genentech would be happier if there was a large amount of animal experimentation before the clinical trial. However, the reality is that animal models are not suitable to test for regeneration of nervous system cells at this time. We have a treatment which is ground-breaking and which is working in our patient. Let us go forward with further testing. Re-manufacture will take years. Meanwhile all the medication for clinical testing is literally going to waste. Patients are suffering and family and spouses are watching tragic deterioration in their loved ones."
Contact: George R. Schwartz, M.D. Senior Researcher, Neuroregeneron Company (a Division of Schwartz Pharma LLC) Tel: 505-610-8243 http://www.healingresearch.org.
This release was issued through eReleases(TM). For more information, visit http://www.ereleases.com.
New hope for the paralysed as scientists re-grow spinal cords (Filed: London Telegraph 13/07/2003)
Thirty years of research culminate in a 'Eureka!' moment for a French biologist working to regenerate severed nerve endings, reports Kim Willsher in Paris.
French scientists have discovered a way to regenerate damaged spinal cords in a breakthrough that might eventually allow paralysed people to walk again.
They have succeeded in regrowing the broken spinal cords of mice after identifying and eliminating two proteins that create scar tissue. This had prevented nerve endings from repairing themselves.
When the scientists, from the French National Health and Medical research institute, manipulated the genes of the mice so that they did not produce these proteins, they discovered that the spinal cord regrew within weeks, enabling the animals to run around again.
Until now the regeneration of nerve cells - or neurones - in the brain and the spinal column has been considered impossible. Previous attempts to restore locomotion by connecting the two extremities of a severed spine have always failed. The research team is now working on ways of using the discovery to develop a treatment for humans.
Prof Alain Privat, 59, who led the team of biologists from the Languedoc University of Science in Montpellier, in the south of France, told The Telegraph yesterday: "It's a very important breakthrough because for the past 100 years researchers working on the regeneration of the central nervous system have been unable to crack this problem. Nobody envisaged being able to repair the system because for a very long time they thought it was something inherent to the fibres themselves that meant they couldn't grow back and restore movement.
"Today, after six years of research, we are sure that we can make them work again if we can stop these two proteins from being produced."
The team is now working on adapting the process so that it can be applied first to mice and monkeys whose genes have not been modified, before carrying out tests on human patients. The key will be to find ways to inhibit the two proteins that stop the cord from repairing itself.
"We've proved that it is possible, but it's going to take at least five years to find out if the technique can be applied to humans. We can produce mutant mice but not mutant people, so we have to find other ways of tackling these proteins," said Prof Privat.
"I cannot be 100 per cent sure it will work on humans but I think the chances are very reasonable. It should work but until we do it we won't be sure."
The next stage involves developing a "genetic treatment" that could be applied to a damaged spine to halt the scarring process and stimulate the regrowth of the nerves. The researchers are also looking at whether the regeneration of the neurones can be jump-started in older injuries, enabling paralysed people such as Christopher Reeve, the actor, to benefit.
Prof Privat believes that the technique could also be used to treat brain injuries and degenerative conditions such as Parkinson's disease. "The neurons in the brain are very similar to those in the spinal cord," he said.
The professor, who is the director of the national health and medical research institute at Languedoc University, has worked with a team of three researchers to try to crack the secret of repairing spinal cord neurones.
The team identified two key proteins, Vimentine and Glial Fibrillary Acidic Protein (GFAP). These proteins, they discovered, were produced in large quantities after a spinal injury and formed a scar, described as an impenetrable wire fence, around the damaged nerve endings, preventing messages being transmitted to the brain.
Under his guidance, his researchers took 100 ordinary laboratory mice and divided them into four groups. The first was left alone as a control set, the second and third were genetically altered to eliminate one of the proteins, and the last group was "double mutants", lacking the genes that produced both proteins. The mice were all tested for their ability to run on a treadmill before their spines were cut.
Prof Privat, 59, said: "After two or three weeks there was a small group of animals that was performing better than the others. After four weeks they were moving their back legs and most of their motor function appeared to have been restored.
"We discovered that the control group and those whose genes had been only partially manipulated remained permanently paralysed. However, the mice in which both the proteins had been knocked out had recovered the use of their legs.
"When we examined these mice we found that there were no scars on the spinal cord fibres, the nervous system circuitry had been reconstructed and that after four weeks there was almost no remnants of paralysis in their hind legs.
"Thanks to this genetic modification the scarring that seems to be the cause of the paralysis didn't happen and the nerve fibres were able to grow back.
"He recalled the moment he and his team realised that they had achieved the breakthrough. "It took just a few seconds for us to realise that we had finally done it."It was a real 'Eureka!' moment and I jumped into my office where I had a very old bottle of my favourite Scotch, Glenlivet. I said to the others, "This is the time to crack it open'," Prof Privat added.
"It is absolutely ground-breaking. I started working on this subject around 30 years ago and I had finally done it."
One of the first problems he faces, however, is finding the estimated Ł3.5 million ( *5 million) needed to continue the research. Until now, the Montpellier team, hit by French government budget cuts, has relied on cash from private associations dedicated to those paralysed by spinal injuries. He said: "We need a relatively small sum of money. We have these encouraging results but barely the means to go on."
The results of the team's research will be published by the National Academy of Sciences, based in Washington, this month.
[This message was edited by James Kelly on 07-14-03 at 12:45 PM.]
