Positive Results from Phase 2 Study in Spinal Cord Injury


Stem Cells, Inc., has released the six-month results from cohort I of an ongoing Phase 2 clinical trial of human neural stem cells for the treatment of chronic cervical spinal cord injuries. The data displayed significant improvements in muscle strength had occurred in five of the six patients treated. Of these five patients, four of them also showed improved performance on functional tasks that assesses dexterity and fine motor skills. Furthermore, these four patients improved in the level of spinal cord injury according to the classification system provided by the International Standards for Neurological Classification of Spinal Cord Injury or ISNCSCI.

Stem Cells, Inc., expects to release their detailed final 12-month results on this first open-cohort later this quarter.

Chief medical officer, Stephen Huhn, presented these data at the American Spinal Injury Association annual meeting in Philadelphia, on Friday, April 15.  Dr. Huhn also believes that the interim results are very encouraging and reason to be quite hopeful.

“The emerging data continue to be very encouraging,” said Dr. Huhn. “We believe that these types of motor changes will improve the independence and quality of life of patients and are the first demonstration that a cellular therapy has the ability to impact recovery in chronic spinal cord injury. We currently have thirteen sites in the United States and Canada that are actively recruiting patients. We have enrolled and randomized 19 of the 40 total patients in the statistically powered, single-blind, randomized controlled, Cohort II. We are projecting to complete enrollment by the end of September so that we can have final results in 2017.”

The present Phase 2 clinical trial is a multi-center enterprise that includes physicians and scientists at 13 different sites in the united States and Canada. Incidentally, these sites are presently actively recruiting patients.

Stem Cells, Inc., has enrolled and randomized 19 of the 40 total patients in this statistically powered, single-blind, randomized controlled, cohort II.

The Phase 2 study, “Study of Human Central Nervous System (CNS) Stem Cell Transplantation in Cervical Spinal Cord Injury,” will determine the safety and efficacy of transplanting the company’s proprietary human neural stem cells (HuCNS-SC cells) into patients with traumatic injury of the cervical region of the spinal cord.

Cohort I is an open label dose-ranging cohort in six AIS-A or AIS-B subjects. For those of you not familiar with the American Spinal Injury Impairment Scale (ASI A-E scale), here is a summary of the classification scheme:

ASI – A = Complete paralysis; No sensory or motor function is preserved in the sacral segments S4-5.
ASI – B = Sensory Incomplete; Sensory but not motor function is preserved below the neurological level and includes the sacral segments S4-5 (light touch or pin prick at S4-5 or deep anal pressure) AND no motor function is preserved more than three levels below the motor level on either side of the body.
ASI – C = Motor Incomplete; Motor function is preserved below the neurological level**, and more than half of key muscle functions below the neurological level of injury (NLI) have a muscle grade less than 3 (Grades 0-2).
ASI – D = Motor Incomplete; Motor function is preserved below
the neurological level**, and at least half (half or more) of key muscle functions below the NLI have a muscle grade > 3.
ASI – E = Normal; If sensation and motor function as tested with the ISNCSCI are graded as normal in all segments, and the patient had prior deficits, then the AIS grade is E. Someone without an initial SCI does not receive an AIS grade.
Cohort II is a randomized, controlled, single-blinded cohort in forty AIS-B subjects. Cohort III, which will only be conducted at the discretion of the sponsor, is an open-label arm that involves six AIS-C subjects.
The primary efficacy outcome will focus on changes in the upper extremity strength as measured in the hands, arms, and shoulders.  This trial will enroll up to 52 subjects.
StemCells, Inc. has demonstrated the safety and efficacy of their HuCNS-SC cell in preclinical studies in laboratory rodents.  Additional Phase I studies yielded positive human safety data.  Furthermore, completed and ongoing clinical studies in which its proprietary HuCNS-SC cells have been transplanted directly into all three components of the central nervous system: the brain, the spinal cord and the retina of the eye, have further demonstrated the safety of HuCNS SC cells in human patients.
StemCells, Inc. clinicians and scientists believe that HuCNS-SC cells may have broad therapeutic application for many diseases and disorders of the CNS. Because the transplanted HuCNS-SC cells have been shown to engraft and survive long-term, there is the possibility of a durable clinical effect following a single transplantation.
The HuCNS-SC platform technology is a highly purified composition of human neural stem cells (tissue-derived or “adult” stem cells). Manufactured under cGMP standards, the Company’s HuCNS-SC cells are purified, expanded in culture, cryopreserved, and then stored as banks of cells, ready to be made into individual patient doses when needed.

Stem Cell-Based Spinal Cord Repair Enables Robust Corticospinal Regeneration


In the March 28th, 2016 issue of the journal Nature Medicine, Mark Tuszynski and his colleagues from the University of California, San Diego, in collaboration with colleagues from Japan and Wisconsin, report that they were able to successfully coax stem cell-derived neurons to regenerate damaged corticospinal tracts in rats. Furthermore, this regeneration produced observable, functional benefits.

What is the “corticospinal tract” you ask? The corticospinal tracts are part of the “pyramidal tracts” that include the corticospinal and corticobulbar tracts. The pyramidal tracts are the main controllers of voluntary movement and connect their nerve fibers eventually to cells that serve voluntary muscles and allow them to contract. We call such nerves “motor nerves,” and the corticospinal nerve tracts are among the most important of the motor nerve tracts.

These neural tracts are collectively called “pyramidal tracts” because they pass through a small area of the brain stem known as the pyramids, which lie on the ventral side of the medulla oblongata. Both pyramidal tracts originate in the forebrain; specifically from the so-called “motor cortex” of the forebrain. The motor cortex lies just in front of the central sulcus of the forebrain. In the motor cortex, lies thousands of “upper motor neurons” that extend their axons down to the brain stem and spinal cord.

Forebrain areas

In the brain stem, the majority of these corticospinal tracts crossover (or decussate) to the other side of the brain stem and travel down the opposite side of the spinal cord. The corticospinal axons extend all the way down the spinal cord, until they make a connection (synapse) with a “lower motor neuron” that extends its axon to the skeletal muscles that it will direct to contract. The corticobulbar tract contains nerves that conduct nerve impulses from cranial nerves and these help the muscles of the face and neck contract, and are involved in facial expressions, swallowing, chewing, and so on.

Corticospinal tracts

Damage to the upper motor neurons as a result of a stroke can rob a person of the ability to move, since the muscles that are attached to the upper motor neurons cannot receive any signals to contract. Likewise, damage to the axonal tracts (also known as nerve fibers) can paralyze a patient and rob them of their ability to move.

The director of this research project, Mark Tuszynski, MD, PhD, professor in the UC San Diego School of Medicine Department of Neurosciences and director of the UC San Diego Translational Neuroscience Institute, said: “The corticospinal projection is the most important motor system in humans. It has not been successfully regenerated before. Many have tried, many have failed – including us, in previous efforts.”

Dr. Tuszynski continued, “The new thing here was that we used neural stem cells for the first time to determine whether they, unlike any other cell type tested, would support regeneration. And to our surprise, they did.”

In this experiment, Tuszynski, and his colleagues and collaborators used rats that had suffered spinal cord injuries and had trouble moving their forelimbs. Then they implanted grafted multipotent neural progenitor cells (MNPCs) into those sites within the spinal cord that had suffered injury, where corticospinal axonal tracts had been severed or damaged. The MNPCs had been previously treated to differentiate into spinal cord-specific motor neurons. Fortunately, the MNPCs prodigiously formed lower motor neurons that made good, solid, functional synapses with interneurons and upper motor neurons that improved forelimb movements in the rats. This work put the lie to previous beliefs about corticospinal neurons; namely that they lacked any of the internal mechanisms required to regenerate severed or damaged connections.

Even though several previous studies have demonstrated functional recovery in spinal cord-injured rats through the use of stem cell-based treatments, none of these studies has convincingly demonstrated regeneration of corticospinal axons.

“We humans use corticospinal axons for voluntary movement,” said Tuszynski. “In the absence of regeneration of this system in previous studies, I was doubtful that most therapies taken to humans would improve function. Now that we can regenerate the most important motor system for humans, I think that the potential for translation is more promising.”

This is certainly exciting work, but even though it worked in rats, it may not yet work in humans. The road from pre-clinical studies in animals to clinical trials in humans is a long, tedious, frustrating, and uncertain pathway, pockmarked with the failures of past therapies that worked well in animals but failed to translate into successes in human patients.

“There is more work to do prior to moving to humans,” Tuszynski said. We must establish long-term safety and long-term functional benefit in animals. We must devise methods for transferring this technology to humans in larger animal models. And we must identify the best type of human neural stem cell to bring to the clinic.”

Combination of Mesenchymal Stem Cells and Schwann Cells Used to Treat Spinal Cord Injury


Spinal cord injuries represent an immensely difficult problem for regenerative medicine. The extensive nature of the damage to the spinal cord is difficult to repair, and the transformation that the injury wrecks in the spinal cord makes the spinal cord inhospitable to cellular repair.

Fortunately some headway is being made, and several clinical trials have shown some success with particular stem cells. Neural stem cells can differentiate into new neurons and glial cells and replace dead or damaged cells (see Tsukamoto A., et Al., Stem Cell Res Ther 4,102, 2013 ). Oligodendrocyte progenitor cells (OPCs) derived from embryonic stem cells or other sources can replace the myelin sheath that died off as a result of the injury (Alsanie WF, Niclis JC, Petratos S. Stem Cells Dev. 2013 Sep 15;22(18):2459-76).  Olfactory ensheathing cells can move across the glial scar and facilitate the regrowth of severed axons across the scar (Tabakow P, et al., Cell Transplant. 2014;23(12):1631-55). Mesenchymal stem cells can mitigate the inflammation in the damaged spinal cord, and, maybe, stimulate endogenous stem cell populations to repair the spinal cord (Geffner L.F., et al., Cell Transplant 17,1277, 2008). Therefore, several cell types seem to have some ability to heal the damaged spinal cord.

A new clinical trial from the Zali laboratory at Shahid Beheshti University of Medical Sciences, in Tehran, Iran, has examined the used of two different stem cells to treat spinal cord injury patients. This trial was a small, Phase I trial that only tested the safety of these treatments.

Zali and his colleagues assessed the safety and feasibility of transplanting a combination of bone marrow mesenchymal stem cells (MSCs) and Schwann cells (SCs) into the cerebral spinal fluid (CSF) of patients with chronic spinal cord injury. SCs are cells that insulate peripheral nerves with a myelin sheath. Even though SCs are not found in the central nervous system, they do the same job as oligodendrocytes, and several experiments have shown that when transplanted into the central nervous system, SCs can do the job of oligodendrocytes in the central nervous system.

In this trial, six subjects with complete spinal cord injury according to International Standard of Neurological Classification for Spinal Cord Injury (ISNCSCI) developed by the American Spinal Injury Association were treated with co-transplantation of their own MSCs and SCs by means of a lumbar puncture. The neurological status of these patients was ascertained by the ISNCSCI and by assessment of each patient’s functional status according to the Spinal Cord Independent Measure. Before and after cell transplantation, the spinal cord of each patient was imaged by means of magnetic resonance imaging (MRI). All patients also underwent electromyography, urodynamic study (UDS) and MRI tractograghy before the procedure and after the procedure if patients reported any changes in motor function or any changes in urinary sensation.

In a span of 30 months following the procedure, radiological findings were unchanged for each patients. There were no signs or indications of neoplastic tissue overgrowth in any patient. In one patients, their American Spinal Injury Association class was downgraded from A to B. This same patients had increased bladder compliance, which correlated quite well with the axonal regeneration detected in MRI tractography. None of these patients showed any improvement in motor function.

To summarize, there were no adverse effects detected around 30 months after the transplantations. These results suggest that this stem cell combination is safe as a treatment for spinal cord injury. While improvement of observed in one patients, because the trial was not designed to investigate the efficacy of the treatment, it is difficult to make any hard-and-fast conclusions about the efficacy of this treatment at this time. However, the fact that one patient did improve is at least encouraging.

These data were reported in the journal Spinal Cord (Spinal Cord. 2015 Nov 3. doi: 10.1038/sc.2015.142).

Regenerating Nerve Tissue in Spinal Cord Injuries


Severe injuries to the neck during recreational activities such as horseback riding or playing football can permanently alter someone’s life dramatically. With no options for the repair of spinal cord injuries, many are left with little hope for recovery.

New work by researchers at Rush University Medical Center (RUMC) in Chicago is investigating a new therapy that uses stem cells to treat spinal cord injuries within the first 14 to 30 days of injury. Rush is one of only two centers in the country currently studying this new approach.

“There are currently no therapies that successfully reverse the damage seen in the more than 12,000 individuals who suffer a spinal cord injury each year in the United States alone,” says Richard G. Fessler, MD, PhD, professor of neurological surgery at RUMC. An estimated 1.3 million Americans are living with a spinal cord injury.

“These injuries can be devastating, causing both emotional and physical distress, but there is now hope. This is a new era where we are now able to test whether a dose of stem cells delivered directly to the injured site can have an impact on motor or sensory function,” Fessler continued. “If we could generate even modest improvements in motor or sensory function, it would result in significant improvements in quality of life.”

Dr. Fessler is the principal investigator at RUMC of a clinical trial that involves progenitor cells that are likely to develop into a certain cell types. Specifically, this study is studying nerve cells known as oligodendrocyte progenitor cells, which potentially can make poorly functioning nerves function better. A San Francisco Bay-area biotechnology company known as Asterias Biotherapeutics, developed the cells and is sponsoring the trial.

This clinical trial is designed to assess the safety and efficacy of increasing doses of AST-OPC1 to treat individuals with a cervical spinal cord injury that resulted in tetraplegia, the partial or total paralysis of arms, legs and torso. As of mid-August, one individual has been enrolled in the study at Rush and there are high hopes that others will be enrolled as well in the near future.

Three escalating doses of AST-OPC1 will be examined in patients with subacute, neurologically complete injury to the cervical spinal cord (the spinal cord in the neck, specifically, the spinal nerves known as C5 to C7). These individuals essentially have lost all sensation and movement below their injury site and have severe paralysis of the upper and lower limbs.

In order for this therapy to work, the spinal cord must be continuous not severed. Patients must be able to begin treatment within 25 days of their injury.

Fessler and his group will administer AST-OPC1 between 14 to 30 days after sustaining the injury. Following the treatment, patients will receive frequent neurological exams and imaging in order to assess the efficacy of the treatment. Furthermore, patients will be followed for 15 years thereafter.

“If this treatment proves to be safe and effective, in the future, it also might be used for peripheral nerve injury or other conditions that affect the spinal cord, such as multiple sclerosis or ALS,” Fessler says.

The study is recruiting male and female patients ages 18 to 65 who have recently experienced a cervical spinal cord injury at the neck that resulted in partial or total paralysis of arms, legs and torso. All participants must be able to provide consent and commit to a long-term follow-up study.

Stem Cells Inc Spinal Cord Injury Trial Shows Sustained Improvements in Sensory Function


A cellular therapeutic company known as Stem Cells, Incorporated has been carrying out a Phase I/II clinical trial that was specifically designed to assess both safety and preliminary efficacy of their proprietary HuCNS-SC cells as a treatment for chronic spinal cord injury. Recently, Dr. Armin Curt, the principal investigator of this clinical trial, presented a summary of the safety and preliminary efficacy data from this Phase I/II study at the 4th Joint International Spinal Cord Society (ISCoS) and American Spinal Injury Association (ASIA) meeting which was held in Montreal, Canada.

Spinal cord injury patients are classified by a system that was developed by the American Spinal Injury Association (ASIA) and uses grades A through E on the American Spinal Injury Association Impairment Scale (AIS) to indicate the severity of the spinal cord injury. AIS Grade A injuries consist of a loss of all spinal cord function (sensation and movement) below the level of injury is lost. This is known as a complete injury. All the other AIS grades are considered incomplete. Patients with Grade B injuries have some sensation below the level of injury, but there is no movement below the injury.. In patients with AIS Grade C injuries, there is both sensation and movement, but most of the muscles below the injury cannot function against resistance and that includes gravity. Those with AIS Grade D spinal cord injuries have some sensation and movement, but more than half of the muscles below the injury can function against resistance. Finally those with AIS Grade E injuries have both normal sensation and movement, but there may be other signs of injury, for example, pain.

For this trial, Stem Cell Inc enrolled 12 subjects who had suffered from a severe spinal cord injury at the thoracic or chest level (T2-T11); seven AIS A and 5 AIS B patients.. In order to qualify for this study, all patients had to be classified as either AIS A or B and a minimum of 3 months from injury.

The trial involved internationally prominent medical centers for spinal cord injury and rehabilitation, and associated principal investigators; Dr. Armin Curt at the University of Zurich and Balgrist University Hospital, Dr. Steve Casha at the University of Calgary, and Dr. Michael Fehlings at the University of Toronto.

All subjects in this trial received HuCNS-SC cells by means of direct transplantation into the spinal cord and they were also treated, temporarily, with immunosuppressive drugs to prevent the immune system from rejecting the implanted cells. Patients were regularly evaluated for safety of the treatment protocol, and to determine if patients showed any change in neurological function. To determine this, patients were given a standard battery of movement and sensory tests before the surgery and at routine intervals after the procedure. Thus all patients were simultaneously enrolled in a safety evaluation and separate evaluation that tested the efficacy of the procedure as well.

In the safety analyses of these subjects, all the data demonstrated that the surgical transplantation technique and cell dose were safe and well tolerated by all patients. HuCNS-SC cells were injected directly into the spinal cord both above and below the level of injury and none of the patients in sequential examinations over the course of twelve months showed any abnormal changes in spinal cord function associated with the transplantation technique. Additionally, there were no adverse events that could be attributed to the HuCNS-SC cells.

Analyses of the functional data after twelve-months revealed sustained improvements in sensory function that emerged consistently around three months after transplantation and persisted until the end of the study. These gains in sensory function involved multiple sensory pathways and were observed more frequently in the patients with less severe spinal cord injuries. Three of the seven AIS A patients and four of the five AIS B patients showed signs of positive sensory gains. Two patients in the study progressed from AIS A, to the lesser degree of injury grade, AIS B.

“It has been a privilege to be a part of the first study to test the potential of neural stem cell transplantation in thoracic spinal cord injury,” said Dr. Armin Curt, Professor and Chairman of the Spinal Cord Injury Center at Balgrist University Hospital, University of Zurich. “The gains we have detected indicate that areas of sensory function have returned in more than half the patients. Such gains are unlikely to have occurred spontaneously given the average time from injury. This patient population represents a form of spinal cord injury that has historically defied responses to experimental therapies, and the measurable gains we have found strongly argue for a biological result of the transplanted cells. These gains are exciting evidence that we are on the right track for developing this approach for spinal cord injury. This early outcome in thoracic injury firmly supports testing in cervical spinal cord injury.”

Stephen Huhn, M.D., FACS, FAAP, Vice President, Clinical Research and CMO at StemCells, Inc., said, “This research program has the potential to revolutionize the therapeutic paradigm for spinal cord injury patients. The clinical gains observed in this first study are a great beginning. We found evidence of sensory gains in multiple segments of the injured thoracic spinal cord across multiple patients. Our primary focus in this study for spinal cord injury was to evaluate safety and also to look for even small signs of an effect that went beyond the possibility of spontaneous recovery. We are obviously very pleased that the pattern of sensory gains observed in this study are both durable and meaningful, and indicate that the transplantation has impacted the function of damaged neural pathways in the cord. The Company’s development program has now advanced to a Phase II controlled study in cervical spinal cord injury where the corollary of sensory improvements in thoracic spinal cord injury could well be improved motor function in the upper extremities of patients with cervical spinal cord injuries.”

Prenatal Stem Cell Treatment Improves Mobility in Lambs With Spina Bifida


UC Davis fetal surgeon Dr. Diana Farmer has been at the forefront of treating spina bifida in infants while they are still in their mother’s womb. Now, Dr. Farmer and her colleagues have used a large animal model system to study the use of stem cells to improve the clinical outcomes of children who undergo these types of in utero procedures.

Spina bifida is a congenital birth defect that results from abnormal development of the spinal cord. During development, the spinal cord, which beings as a tube (the neural tube), is open at both ends, and these ends eventually close. However, if the posterior opening to the neural tube does not close properly, then the developing spinal cord will have severe structural defects. These structural defects adversely affect the nerves that issue from the spinal cord and spinal bifida can cause lifelong cognitive, urological, musculoskeletal and motor disabilities.

Dr. Farmer’s chief collaborator was another UC Davis science named Aijun Wang, who serves as the co-director of the UC Davis Surgical Bioengineering Laboratory.

“Prenatal surgery revolutionized spina bifida treatment by improving brain development, but it didn’t benefit motor function as much as we hoped,” said Farmer, who serves as chair of the UC Davis Department of Surgery and is the senior author of this study, which was published online in the journal Stem Cells Translational Medicine.

“We now think that when it’s augmented with stem cells, fetal surgery could actually be a cure,” said Wang.

Years ago, Farmer and her colleagues showed in an extensive clinical trial called the Management of Myelomeningocele Study (MOMS) that babies who were diagnosed with spina bifida and were eligible for in utero surgery had better outcomes that babies who underwent surgery after they were born. Babies with spina bifida who were operated on in utero had a better chance of walking, and not needing a shunt to deal with the pressure problems in the brain that some children with spina bifida experience (see N. Scott Adzick, et al., New England Journal of Medicine 2011;364(11):993-1004). Even with this study, the majority of the babies who were treated with in utero surgery were still unable to walk. To improve a baby’s chances of walking, Farmer and her collaborators turned to stem cell treatments.

Farmer and Wang combined fetal surgery with a the transplantation of stem cells from human placentas to improve neurological capabilities of babies born with spina bifida. In children, spina bifida can range from barely noticeable to rather severe. Myelomeningocele is the most common and, unfortunately, the most disabling form of spina bifida. In babies with myelomeningocele, the spinal emerges through the back and usually pulls brain tissue into the spinal column, which causes cerebrospinal fluid to fill the interior of the brain. Therefore, such patients require permanent shunts in their brains in order to drain the extra cerebrospinal fluid.

Myelomeningocele
Myelomeningocele

In this study, lambs with myelomeningocele were operated on in utero in order to return exposed spinal cord tissue into the vertebral column. Then human placenta-derived mesenchymal stromal cells (PMSCs), which have demonstrated neuroprotective qualities (see Yun HM, et al., Cell Death Dis. 2013;4:e958), were embedded in hydrogel and applied to the site of the lesion. A scaffold was placed on top to hold the hydrogel in place, and the surgical opening was closed.

Six of the animals that received the stem cell treatment were able to walk without noticeable disability within a few hours following birth. However, the six control animals that received only the hydrogel and scaffold were unable to stand.

“We have taken a very important step in expanding what MOMS started,” said Wang. “Next we need to confirm the safety of the approach and determine optimal dosing.”

Farmer and Wang will continue their efforts with funding from the California Institute for Regenerative Medicine. With additional evaluation and FDA approval, the new therapy could be tested in human clinical trials.

“Fetal surgery provided hope that most children with spina bifida would be able to live without shunts,” Farmer said. “Now, we need to complete that process and find out if they can also live without wheelchairs.”

NG2-Expressing Neural Lineage Cells Derived from Embryonic Stem Cells Penetrate Glial Scar and Promote Axonal Outgrowth After Spinal Cord Injury


After a spinal cord injury, resident stem cells in the spinal cord contribute to the production of a glial scar that is rich in chondroitin sulfate proteoglycan (CSPG). The glial scar is a formidable barrier to axonal regeneration in the injured spinal cord, since CSPG actively repels growing axonal growth cones. Even though the glial scar seals off the spinal cord from further damage from inflammation, the long-term effects of the glial scar are to prevent regeneration of spinal nerves, which have the ability to regenerate in culture.

The major components of the site of injury include myelin debris, the scar-forming astrocytes, activated resident microglia and infiltrating blood-borne immune cells, chondroitin sulfate proteoglycans (CSPGs) and other growth-inhibitory matrix components. All of them are potential targets for therapeutic intervention. Many of the interventions can be optimized by considering the beneficial aspects of the scar tissue and fine-tuning the optimal time window for their application. Each target and the strategies directed at its modulation are shown.
The major components of the site of injury include myelin debris, the scar-forming astrocytes, activated resident microglia and infiltrating blood-borne immune cells, chondroitin sulfate proteoglycans (CSPGs) and other growth-inhibitory matrix components. All of them are potential targets for therapeutic intervention. Many of the interventions can be optimized by considering the beneficial aspects of the scar tissue and fine-tuning the optimal time window for their application. Each target and the strategies directed at its modulation are shown.

New work by Sudhakar Vadivelu, in the laboratory of John McDonald at the International Center for Spinal Cord Injury, Hugo W. Moser Research Institute at the Kennedy Krieger Institute, Baltimore, Maryland has discovered new ways to breach the glial scar. Vadivelu and colleagues used a cell culture system that tested the ability of particular cells to help growing axonal growth cones penetrate glial scar material. This culture system showed that embryonic stem cell-derived neural lineage cells (ESNLCs) with prominent expression of nerve glial antigen 2 (NG2) survived, and passed through an increasingly inhibitory gradient of CSPG. These cells also expressed matrix metalloproteinase 9 (MMP-9) at the appropriate stage of their development, which helped poke holes in the CSPG. The outgrowth of axons from ESNLCs followed the NG2-expressing cells because the migrating cells chiseled pathways through the CSPG for the outgrowth of new axons.

To confirm these results in a living animal, Vadivelu and others transplanted embryonic stem cell-derived ESNLCs directly into the cavities of a contused spinal cord of laboratory animals 9 days after injury. One week later, implanted ESNLCs survived and expressed NG2 and MMP-9. The axons of these neurons had grown through long distances (>10 mm), although they preferred to grow across white rather than gray matter.

These data are consistent with CSPG within the injury scar acting as an important impediment to neuronal regeneration, but that NG2+ progenitors derived from ESNLCs can alter the microenvironment within the injured spinal cord to allow axons to grow through such a barrier. This beneficial action seems to be due, in part, at least, to the developmentally-regulated expression of MMP-9. Vadivelu and others conclude from these data that it might be possible to induce axonal regeneration in the human spinal cord by transplanting ESNLCs or other cells that express NG2.

Long-term Tumorgenicity of Induced Pluripotent Stem Cells


A paper from the Okano laboratory has shown that implantation of neural stem cells made from induced pluripotent stem cells can still form tumors ever after a long period of time.

This paper is an important contribution to the safety issues surrounding induced pluripotent stem cells (iPSCs). As noted in previous posts, iPSCs are made from adult cells by means of genetic engineering and cell culture techniques. In short, by introducing four different genes into adult cells and then culturing them in a special culture medium, a fraction of these cells will de-differentiate into cells that resemble embryonic stem cells in many ways, but are not exactly like them.

The Okano laboratory made iPSCs using viruses that integrate into the genome of the host cell, which is not the safest option. However, because in the four-gene cocktail that is normally used to reprogram these cells (Oct-4, Klf-4, Sox2, and c-Myc), the c-Myc gene is often thought to be the main cause of tumor formation. Okano and his collaborators made their iPSCs without the c-Myc gene, but only used the three-gene cocktail of Oct-4, Klf-4, and Sox2. Such a cocktail is much less efficient that the four-gene cocktail, but it supposed to make iPSCs that are altogether safer.

These iPSCs were differentiated into neural stem cells that grew as tiny spheres of cells, and these “neurospheres” were transplanted into the spinal cords of mice that had suffered a spinal cord injury. The implanted cells differentiated into neurons and glial cells and restored some neural function to these mice. However, the mice were observed for a long period of time after the implantations to assess the long-term safety of these implanted cells.

After 105 days, the implanted mice began to show deterioration of their neural function and their spinal cords showed tumors. It is clear that the Oct-4 gene that was used in the reprogramming procedure was the reason for the tumor transformation.

Graphical Abstract 20141213

This experiment, once again, calls into question the safety of any method for iPSC generation that leaves the transfected genes in the reprogrammed cells. I reported in a previous post that skin cells made from iPSCs that had their transgenes left in them were good at causing tumors and not as good as forming skin cells, but iPSCs without their reprogramming transgenes were safer and more effective tools for regenerative medicine.  This experiment also shows that c-Myc is not the only concern with iPSCs.  Any of the transgenes used for reprogramming can cause problems, and they must be removed if iPSCs are going to produce safe, differentiated cells.  Finally, this experiment pretty much shows that the use of retrovirus tools to introduce genes into cells for the sake of reprogramming is a bad idea if those cells are going to be used for regenerative medicine.  Non-integrating tools are much safer and preferable in these cases.

The Okano paper appeared in Stem Cell Reports.

Compact Spinal Implants to Help Spinal Cord Injured Patients Walk


An interdisciplinary research team at the Ecole Polytechnique Fédérale de Lausanne (EPFL) in Switzerland led by Dr. Grégoire Courtine and Dr. Stéphanie P. Lacour has recently lifted the curtain on their flexible spinal implant called the electronic dura (or e-Dura). According to Courtine and Lacour, this implant greatly improves spinal injury rehabilitation in spinal cord injured rats.

In a paper published by this team earlier in the journal Science this month, the EPFL team showed that, because of its flexibility, this next-generation e-Dura implant lasts longer (up to two months) and causes much less damage than traditional implants.

These latest results are an extension of earlier research in 2012 in which Courtine and Lacour published breath-taking results that showed spinal cord-injured rats that should have been paralyzed had regained the ability to walk, run, and even climb stairs.

Spinal cord injury results in loss of control over the part of the body below the point of injury. Courtine and his coworkers were able to reactivate the spinal cord in rats with a specific combination of drugs plus electrical stimulation to simulate the excitatory input from the brain. The drugs they used, monoamine agonists, bind to receptors and activate them in the same way that such neurotransmitters would in healthy subjects. When the spinal cord was exposed to these drugs plus mild electrical stimulation, the activated nerve cells in the spinal column produced movement in the paralyzed animals.

Spinal Cord Implant to help the Cripples walk

This movement, however, was largely involuntary, since the brain was not able to communicate with the area below the injured spinal cord. However, over time, as the animals trained and repeatedly walked in their harnesses (which kept them safe from falling); they became more confident in their ability to walk again. In fact, the EPFL team noticed a fourfold growth of new nerves in the spinal cord. This new nerve growth eventually restored communication between the brain and the injured area of the spine.

Courtine, whose eyes sparkle as he passionately talks about his research, is eager to take these findings to the clinic to see if they can help human beings who have suffered a catastrophic spinal cord injury. In preparation for clinical trials, however, they came up against another problem and that was the need for a long-term spinal implant that could deliver the chemical and electrical stimulation needed to initiate spinal cord healing. Courtine hopes that his e-Dura can satisfy this need.

e-Dura

The e-Dura meets two important criteria for spinal implants: durability and biocompatibility. In order to reduce the number of surgical procedures an injured patient must undergo, an implantable device needs to last a long time. It also has to be biocompatible and flexible. Early generation implants caused inflammation and the formation of scar tissue, which usually offset any positive results the implant provided.

When tested on laboratory animals, Courtine’s laboratory applied the e-Dura implant beneath the protective dura mater, directly on the spinal cord. Thankfully, the implant did not cause any adverse effects and lasted long enough for the paralyzed animals to complete their rehabilitation. Functionally, the implant also performed very well.

The e-Dura unit contains very small microfluidic channels that are embedded on a flexible silicon substrate. The device delivers precise amounts of drugs directly to the nerve cells in the spinal cord. Cracked gold conducting wires and electrodes that are made of a composite material that consists of silicon and platinum send electrical signals to the injured spinal cord. Electronic circuitry in the implant also provides the opportunity for the EPFL team to monitor the electrical messages sent back and forth to the brain as the new nerves are activated.

Indeed, this research is wonderfully exciting, but it is unclear how well it will work in humans. First of all, humans will probably need a different cocktail of drugs or a distinct electrical stimulation pattern to stimulate the spinal cord to heal itself. As with much clinical research at the beginning stages, there many unanswered questions to date. Advanced clinical trials will hopefully uncover some of these idiosyncrasies that characterize the injured human spinal cord and such answers are an integral part of providing a protocol that applies uses technology to human patients.

While exoskeleton technology also continues to approach consumer markets, it would be better to return to people their natural ability to walk. However you slice it, spinal cord injury patients may have more options in the coming years.

Grafted Stem Cells Display Robust Growth in Spinal Cord Injury Model


University of San Diego neuroscientists have used an animal model of spinal cord injury to test the ability of engrafted stem cells to regenerate damaged nerves. Mark Tuszynski and his team built on earlier work with implanted neural stem cells and embryonic stem cell-derived neural stem cells in rodents that had suffered spinal cord injuries.

In this study, Tuszynski and others used induced pluripotent stem cells that were made from a 86-year-old male. This shows that skin cells, even from human patients who are rather elderly, have the ability to be reprogrammed into embryonic stem cell-like cells. These cells were differentiated into neural stem cells and then implanted into the spinal cords of spinal cord-injured rodents.

The injured spinal cord is a very hostile place for implanted cells. Inflammation in the spinal cord summons white blood cells to devour cell debris. White blood cells are rather messy eaters and they release enzymes and toxic molecules that can kill off nearby cells. Also, regenerating cells run into a barrier made by support cells called glial cells that inhibit regenerating neurons from regenerating. Thus, the injured spinal cord is quite the toxic waste dump.

To get over this, Tuszynski and his coworkers treated their induced pluripotent stem cell-derived neural stem cells with growth factors. In fact, when the cells were implanted into the animal spinal cords, they were embedded in a matrix that contained growth factors. After three months, Tuszynski and his colleagues observed extensive axonal growth projecting from grafted neurons that reached long distances in both directions along the spinal cord from the brain to the tail end of the spinal cord. These sprouted axons appeared to make connections with the existing rat neurons. Importantly, these axons extended from the site of the injury, which is astounding given that the injured area of the spinal cord has characteristics that are inimical to neuronal and axon growth.

Even though Tuszynski and others showed that neural stem cells made from embryonic stem cells can populate the damaged spinal cord, using induced pluripotent stem cell-derived neural stem cells has an inherent advantage since these cells are less likely to be rejected by the patient’s immune system. Furthermore, the induced pluripotent stem cell-derived neural stem cells showed dramatic growth in the damaged spinal cord, but the implanted animals did not regain the use of their forelimbs. The implanted human cells were fairly young when the implanted animals were tested. Therefore, they might need to mature before they could restore function to the implanted animals.

“There are several important considerations that future studies will address,” Tuszynski said. “These include whether the extensive number of human axons make correct or incorrect connections; whether the new connections contain the appropriate chemical neurotransmitters to form functional connections; whether connections once formed are permanent or transient; and exactly how long it takes human cells to become mature. These considerations will determine how viable a candidate these cells might before use in humans.”

Tuszynski and his group hope to identify the most promising neural stem cell type for repairing spinal cord injuries. Tuszynski emphasized their commitment to a careful, methodical approach:

“Ultimately, we can only translate our animal studies into reliable human treatments by testing different neural stem cell types, carefully analyzing the results, and improving the procedure. We are encouraged, but we continue to work hard to rationally to identify the optimal cell type and procedural methods that can be safely and effectively used for human clinical trials.”

Nose Stem Cells Help Bulgarian Man Walk With Braces


Darek Fidyka, a 38-year-old Bulgarian man, was severely injured by a stab wound in 2010 and consequently lost the ability to walk.

Now, a new procedure using stem cells from his nose has given him the ability to walk with the help of braces.

Olfactory ensheathing cells or OECs (also known as olfactory ensheathing glial or OEGs) are found in the olfactory system, inside the skull and in the covering of cells that lines the roof of the nose. OECs share similarities to other glial cells like Schwann cells, astrocytes, and oligodendrocytes. OECs can aid the extension of neural projections known as axons from the nasal tissue to the olfactory glomeruli. OECs can do this because they secrete several interesting neurotrophic factors and cell adhesion molecules and migrate along with the regenerating axons. Because of these properties, OECs can escort axonal extension through glial scars that are made in a spinal cord after a spinal cord injury. These scars inhibit the outgrowth of new axons but OECs can allow regenerating axons to bridge these glial scars.

An advantage of OECs is that they can coexist with astrocytes, the cells that contribute to the formation of the glial scar, and even seem to prevent the out-of-hand response astrocytes have in response to injury in which they synthesize a host of molecules that inhibit axon regeneration called “inhibitory proteoglycans.”

The pioneering technique used in this procedure, according to Geoffrey Raisman, a professor at University College London’s (UCL) institute of neurology, used OECs to construct a kind of bridge between two stumps of the damaged spinal column.

“We believe… this procedure is the breakthrough which, as it is further developed, will result in a historic change in the currently hopeless outlook for people disabled by spinal cord injury,” said Riesman, who led this research project.

Raisman, who is a spinal injury specialist at UCL, collaborated with neurosurgeons at Wroclaw University Hospital in Poland to remove one of Fidyka’s olfactory bulbs, which give people their sense of smell, and transplant his olfactory ensheathing cells (OECs) in combination with. olfactory nerve fibroblasts (ONFs) into the damaged spinal cord areas. Following 19 months of treatment, Fidyka recovered some voluntary movement and some sensation in his legs.

The Nicholls Spinal Injury Foundation, a British-based charity which part-funded the research, said in statement that Fidyka was continuing to improve more than predicted, and was now able to drive and live more independently.

OECs have been used before to treat spinal cord injury patients. I refer you to chapter 27 in my book, The Stem Cell Epistles, to learn more about these. The novel technique in this paper is the additional use of nasal fibroblasts and the construction of a bridge between the two damaged remnants of the spinal cord.

The reason OECs were recruited to treat spinal cord injuries is that when axons that carry information about smells are damaged, the neuron simply regenerates its atonal extension, which grows into the olfactory bulbs. OECs facilitate this process by re-opening the surface of the olfactory bulbs in order for the new axons to enter them. Thus Raisman and others have the notion that transplanted OECs in the damaged spinal cord could equally facilitate the regeneration of severed nerve fibers.

Raisman also added that the technique used in this case, that is bridging the spinal cord with nerve grafts from the patient, had been used in animal studies for years, but was never used in a human patient in combination with OECs.

“The OECs and the ONFs appeared to work together, but the mechanism between their interaction is still unclear,” he said in a statement about the work.

Several spinal cord injury experts who were not directly involved in this work said its results offered some new hope. However, they were also quick to add that more work needed to be done to precisely determine what had led to this success. More patients must be successfully treated with this procedure before its potential can be properly assessed.

“While this study is only in one patient, it provides hope of a possible treatment for restoration of some function in individuals with complete spinal cord injury,” said John Sladek, a professor of neurology and pediatrics at the University of Colorado School of Medicine in the United States.

Raisman and his team now plan to repeat the treatment technique in between three and five spinal cord injury patients over the next three to five years. “This Nose will enable a gradual optimization of the procedures,” he told Reuters.

Transplanted Mesenchymal Stem Cells Prevent Bladder Scarring After Spinal Cord Injury


A collaborative research effort between laboratories from Canada and South Korea have shown that a cultured mesenchymal stem cell line called B10 can differentiate into smooth muscle cells and improve bladder function after a spinal cord injury.

Spinal cord injury can affect the lower portion of the urinary tract. Overactive bladder, urinary retention, and increased bladder thickness and fibrosis (bladder scarring) can result from spinal cord injuries. Human mesenchymal stem cells (MSCs) can differentiate under certain conditions into smooth muscle. For this reason, MSCs have therapeutic potential for patients who have suffered from spinal cord injuries.

Seung U. Kim and his colleagues from Gachon University Gil Hospital in Inchon, South Korea have made an immortalized human mesenchymal stem cell line by transfecting primary cell cultures of fetal human bone marrow mesenchymal stem cells with a retroviral vector that contains the v-myc oncogene. This particular cells line, which they called HM3.B10 (or B10 for short), grows well in culture and can also differentiates into several different cell types.

In this present study, which was published in the journal Cell Transplantation, Kim and his colleagues and collaborators injected B10 hMSCs directly into the bladder wall of mice that had suffered a spinal cord injury but were not treated showed no such improvement.

“Human MSCs can secrete growth factors,” said study co-author Seung U. Kim of the Division of Neurology at the University of British Columbia Hospital, Vancouver, Canada. “In a previous study, we showed that B 10 cells secrete various growth factors including hepatocyte growth factor (HGF) and that HGF inhibits collagen deposits in bladder outlet obstructions in rats more than hMSCs alone. In this study, the SCI control group that did not receive B10 cells showed degenerated spinal neurons and did not recover. The B10-injected group appeared to have regenerated bladder smooth muscle cells.”

Four weeks after the initial spinal cord injury, the mice in the B10-treated group received injections of B10 cells transplanted directly into the bladder wall. Kim and his team used magnetic resonance imaging (MRI) to track the transplanted B10 cells. The injected B10 cells had been previously labeled with fluorescent magnetic particles, which made them visible in an MRI.

“HGF plays an essential role in tissue regeneration and angiogenesis and acts as a potent antifibrotic agent,” explained Kim.

These experiments also indicated that local stem cell injections rather than systemic, intravenous infusion was the preferred method of administration, since systemic injection caused the hMSCs get stuck largely in the blood vessels of the lungs instead of the bladder.

The ability of the mice to void their bladders was assessed four weeks after the B10 transplantations. MRI analyses clearly showed strong signals in the bladder as a result of the labeled cells that had been previously transplanted. Post-mortem analyses of the bladders of the transplanted group showed even more pronounced differences, since the B10-injected animals had improved smooth muscle cells and reduced scarring.

These results suggest that MSC-based cell transplantation may be a novel therapeutic strategy for bladder dysfunction in patients with SCI.

“This study provides potential evidence that an human [sic] stable immortalized MSC line could be useful in the treatment of spinal cord injury-related problems such as bladder dysfunction.” said Dr. David Eve, associate editor of Cell Transplantation and Instructor at the Center of Excellence for Aging & Brain Repair at the University of South Florida. “Further studies to elucidate the mechanisms of action and the long-term effects of the cells, as well as confirm the optimal route of administration, will help to illuminate what the true benefit of these cells could be.”

Human Stem Cell-Derived Neurons Grow New Axons In Spinal Cord Injured Rats


A stem cell-based treatment for spinal cord injury took one more baby step forward when scientists from the laboratory of Mark Tuszynski at the at the University of California, San Diego used cells derived from an elderly man’s skin to regrow neural connections in rats with damaged spinal cords.

Tuszynski and others published their results in the Aug. 7 online issue of the journal Neuron. In that paper, Tuszynski and his co-worker report that human stem cells triggered the growth of numerous axons in the damaged spinal cord. Axons are those fibers that extend from the main part or body a neuron (nerve cell) that serve to send electrical impulses away from the body to other cells. Some of these new axons even grew into the animals’ brains.

Axon picture

Dr. Mark Tuszynski is a professor of neurosciences at the University of California, San Diego. “This degree of growth in axons has not been appreciated before,” he said. However, Tuszynski also cautioned that there is still much to be learned about how these newly established nerve fibers behave in laboratory animals. He likened the potential for stem-cell-induced axon growth to nuclear fusion. If it’s contained, you get energy; if it’s not contained, you get an explosion. “Too much axon growth into the wrong places would be a bad thing,” Tuszynski added.

Stem cell researchers have examined the potential for stem cells to restore functioning nerve connections in people with spinal cord injuries. Embryonic stem cells have been used to make new neurons and to also make “oligodendrocyte progenitor cells” or OPCs, which make the insulating myelin sheath that enwraps the axons of spinal nerves. However, several other types of stem cells can make OPCs and new neurons and these stem cells do not come from embryos (for more, see chapter 27 of my book, The Stem Cell Epistles).

In this study, Tuszynski and his team used induced pluripotent stem cells or iPSCs, which are derived from mature adult cells by means of genetic engineering and cell culture techniques. They used cells from a healthy 86-year-old man and genetically reprogrammed so that they were reprogrammed into iPSCs. These iPSCs were then differentiated into neurons that were implanted into a special scaffold embedded with proteins called growth factors, and then grafted into the spinal cords of laboratory rats with spinal cord injuries.

Over the course of several months, these animals showed new, mature neurons and extensive growth in the cells’ axons. These fibers grew through the injury-related scar tissue in the animals’ spinal cords and connected with resident rat neurons.

This is an enormous advance, because the wounded spinal cord creates a “Glial scar” that contains a host of molecules that repel growing axons. Even though this glial scar prevents the immune system from leaking into the spinal cord and destroying it, this same scar prevents the regeneration of damaged neurons and their severed axons.

Glial scar axon repulsion

Dr. David Langer, director of neurosurgery at Lenox Hill Hospital in New York City said: “One of the big obstacles [in this type of research] is this area of scarring in the spinal cord. Getting neurons to traverse it is a real challenge,” said Langer, who was not involved in the research. “The beauty of this study,” he said, “is that they got the neurons to survive and traverse the scar.”

Langer also cautioned, much like Tuszynski, that this experimental success is just a preliminary step. There are, in his words, “huge questions” as to whether or not these axons can make appropriate connections and actually restore function to spine-damaged lab animals. “It’s not just a matter of having the cables,” Langer said. “The wiring has to work.”

And even if this stem cell approach does pan out in animals, Langer added, it would all have to be translated to humans. “We have a long way to go until we’re there,” he said. “It’s not that people shouldn’t have hope. But it should be a realistic hope.”

A few biotech companies have already launched early-stage clinical trials using embryonic (Geron) or fetal stem cells (StemCells Inc) to treat patients with spinal cord injuries. But Tuszynski said his team’s findings offer a cautionary note about moving to human trials too quickly. “We still have a lot to learn,” he said. “We want to be very sure these axons don’t make inappropriate connections. And we need to see if the new connections formed by these axons are stable.”

Ideally, Tuszynski added, if stem cells were to be used in treating spinal cord injuries, they’d be generated as they were in this study — by creating them from a patient’s own cells. That way, he explained, patients would not need immune-suppressing drugs afterward.

Gene Inhibitor Plus Fish Fibrin Restore Nerve Function Lost After a Spinal Cord Injury


Scientists at UC Irvine’s Reeve-Irvine Research Center have discovered that injections of salmon fibrin injections into the injured spinal cord plus injections of a gene inhibitor into the brain restored voluntary motor function impaired by spinal cord injury.

Gail Lewandowski and Oswald Steward, director of the Reeve-Irvine Research Center at UCI, examined rodents that had received spinal cord injuries.  They were able to heal the damage by developmentally turning back the clock in a molecular pathway that is critical to the formation of the corticospinal nerve tract, and by providing a scaffold for the growing neurons so that the axons of these growing neurons could grow and make the necessary connections with other cells.  Their research was published in the July 23 issue of The Journal of Neuroscience.

The work of Steward and Lewandowski is an extension of previous research at UC Irvine from 2010.  Steward and his colleagues discovered that the axon of neurons grow quite well once an enzyme called PTEN is removed from the cells.  PTEN is short for “phosphatase and tensin homolog,” and it removes phosphate groups from specific proteins and lipids.  In doing so, PTEN signals to cells to stop dividing and it can also direct cells to undergo programmed cell death (a kind of self-destruct program).  PTEN also prevents damaged tissues from regenerating sometimes, because it is a protein that puts the brakes of cell division.  Mutations in PTEN are common in certain cancers, but the down-regulation of PTEN is required for severed axons to re-form, extend, migrate to their original site, and form new connections with their target cells.

PTEN function

 

After two years, team from U.C. Irvine discovered that injections of salmon fibrin into the damaged spinal cord or rats filled cavities at the injury site and provided the axons with a scaffolding upon which they could grow, reconnect and facilitate recovery. Fibrin produced by the blood system when the blood vessels are breached and it is a fibrous, insoluble protein produced by the blood clotting process.  Surgeons even use it as a kind of surgical glue.

“This is a major next step in our effort to identify treatments that restore functional losses suffered by those with spinal cord injury,” said Steward, professor of anatomy & neurobiology and director of the Reeve-Irvine Research Center. “Paralysis and loss of function from spinal cord injury has been considered irreversible, but our discovery points the way toward a potential therapy to induce regeneration of nerve connections.”

In their study, Steward and Lewandowski subjected rats to spinal cord injuries, and then assessed their defects.  Because these were upper back injuries, the rats all showed impaired forelimb (hand) movement.  Steward and Lewandowski then treated these animals with a combination of salmon fibrin at the site of injury and a modified virus that made a molecule that inhibited PTEN.  These viruses were genetically engineered adenovirus-associated viruses encoded a small RNA that inhibited translation of the PTEN gene (AAVshPTEN).  This greatly decreased the levels of PTEN protein in the neurons.  Other rodents received control treatments of only AAVshPTEN and no salmon fibrin.

The results were remarkable.  Those rats that received the PTEN inhibitor alone showed no improvement in their forelimb function, but those animals who were given AAVshPTEN plus the salmon fibrin recovered forelimb use (at least reaching and grasping).

“The data suggest that the combination of PTEN deletion and salmon fibrin injection into the lesion can significantly enhance motor skills by enabling regenerative growth of corticospinal tract axons,” Steward said.

Corticospinal Nerve tract

Statistics compiled by the Christopher & Dana Reeve Foundation suggests that approximately 2 percent of Americans have some form of paralysis that is the result of a spinal cord injury.  Spinal cord injuries break connections between nerves and muscles or nerves and other nerves.  Even injuries the size of a grape can cause complete loss of function below the level of the injury.  Injuries to the neck can cause paralysis of the arms and legs, an absence of sensation below the shoulders, bladder and bowel incontinence, sexual dysfunction, and secondary health risks such as susceptibility to urinary tract infections, pressure sores and blood clots due to an inability to move one’s legs.

Steward said the next objective is to learn how long after injury this combination treatment can be effectively administered. “It would be a huge step if it could be delivered in the chronic period weeks and months after an injury, but we need to determine this before we can engage in clinical trials,” he said.

Stem Cells Inc. Reports Additional Spinal Cord Injury Patients Transplanted with Neural Stem Cell Line Show Functional Improvements


StemCells, Inc. has developed a proprietary stem cell line called HuCNS-SC.  This stem cell line is a neural stem cell line, and neural stem cells can readily form neurons (the conducting cells of the nervous system), or glial cells (the support cells of the nervous system). In order to determine if these cells can regenerate spinal nerves in patients who have suffered a spinal cord injury, StemCells Inc. has commissioned a clinical trial to test their cells in human spinal cord injured patients.

Early indications showed that the HuCNS-SC cells were safe, but some patients have shows improvements in sensation.  Now StemCells Inc has issued an announcement that these initially reported improvements in only a few patients have also been confirmed in other patients.

According to Armin Curt, M.D., Professor and Chairman of the Spinal Cord Injury Center at Balgrist University Hospital, University of Zurich, and the principal investigator of their Phase I/II trial, the initial improvements that were observed in the first two patients treated with their HuCNS-SC neural stem cells have now been observed in two additional patients who have also been treated with these stem cells. These results come from an interim analysis of recent clinical data.

In a presentation to the Annual Meeting of the American Spinal Injury Association in San Antonio, Texas, Dr. Curt showed data on AIS B subjects who were transplanted with HuCNS-SC neural stem cells in the Phase I/II chronic spinal cord injury trial. This trial is different from the AIS A patients who have no mobility or sensory perception below the point of injury, since AIS B subjects are less severely injured, and are paralyzed but retain sensory perception below the point of injury. Two of the three AIS B patients who are participating in the study showed significant gains in sensory perception. The third patient remained stable.  These interim results confirm the favorable safety profile of these stem cells and the surgical implant procedure used to transplant them into the spinal cords of spinal cord injury patients.

Also included in Dr. Curt’s presentation was data from a total of five new subjects with a minimum six-month follow-up. In total, Stem Cells Inc. has now reported clinical updates on a total of eight of the twelve patients enrolled in its Phase I/II clinical trial that is testing this Company’s proprietary HuCNS-SC (purified human neural stem cells) platform technology for treating chronic thoracic spinal cord injury.

“Thoracic spinal cord injury was chosen as the indication in this first trial primarily to demonstrate safety. This patient population represents a form of spinal cord injury that has historically defied responses to experimental therapies and is associated with a very high hurdle to demonstrate any measurable clinical change. Because of the severity associated with thoracic injury, gains in multiple sensory modalities and segments are unexpected, and changes in motor function are even more unlikely,” said Dr. Curt. “In contrast, the cervical cord, which controls more motor function, may represent a patient population in which motor responses to transplant may be more readily anticipated.”

“We are seeing multi-segmental gains and a return of function in the cord in multiple patients. This indicates something that was not working in the spinal cord, now appears to be working following transplantation. This is even more significant because of the time that has elapsed from the date of injury, which ranges from 4 months to 24 months across the subjects with sensory gains,” said Stephen Huhn, M.D., FACS, FAAP, vice president, CNS clinical research at StemCells, Inc. “These results are exciting with respect to the expansion of this trial into patients with cervical injury because even a gain of one to two segments in cervical spinal cord injury patients can allow for additional function in the upper extremities.”

New Analysis of Stem Cell Treatments for Spinal Cord Injury in Laboratory Animals


A host of preclinical studies have examined the ability of stem cells to improve the condition of laboratory animals that have suffered a spinal cord injury. While these studies vary in their size, design, and quality, there has been little cumulative analysis of the data generated by these studies.

Fortunately, there is a powerful analytical tool that can examine data from many studies and this type of analysis is called a “meta-analysis.” Meta-analyses use sophisticated statistical packages to systematically reassess a compilation of the data contained within these papers. Meta-analyses are exhausting, but potentially very useful. Such a meta-analysis is also very important because it provides researchers with an indication of what problems must be worked out before these treatments advance to human clinical trials and what aspects of the treatment work better than others.

A recent meta-analysis of stem cell therapy on animal models of spinal cord injury has been published by Ana Antonic, MSc, David Howells, Ph.D., and colleagues from the Florey Institute and the University of Melbourne, Australia, along with Malcolm MacLeod and colleagues from the University of Edinburgh, UK in the open access journal PLOS Biology.

The goal of regenerative spinal cord treatments is to use stem cells to replace dead cells within damaged areas of the spinal cord. Such treatments would be given to spinal cord injury patients in the hope of improving the ability to move and to feel below the site of the injury. Many experiments that utilize animal models of spinal cord injury have used stem cells to treat laboratory animals that have suffered spinal cord injury, but, unfortunately, these studies are limited in scale by size (as a result of financial considerations), practical and ethical considerations. Such limitations hamper each individual study’s statistical power to detect the true effects of the stem cell implantation. Also, these studies use different types of stem cells in their treatment scenarios, inject those cells differently induce spinal cord injuries differently, and test their animals for functional recovery differently.

To assess these studies, this new paper examined 156 published studies, all of which tested the effects of stem cell treatments on about 6,000 spinal cord-injured animals.

Overall, they found that stem cell treatment results in an average improvement of about 25 percent over the post-injury performance in both sensory (ability to feel) and motor (ability to move) outcomes. Unfortunately, the variation from one animal to another varied widely.

For sensory outcomes the degree of improvement tended to increase with the number of cells implanted. Such dose-responsive results tend to indicate that the improvements are actually due to the stem cells, and therefore, this stem cell-mediated effect represents a genuine biological effect.

The authors also measured the effects of bias. Simply put, if the experimenters knew which animals were treated and which were untreated, then they might be more likely to report improvements in the stem cell-treated animals. They also examined the way that the stem cells were cultured, the way that the spinal injury was generated and the way that outcomes were measured. In each case, important lessons were learned that should help inform and refine the design of future animal studies.

The meta-analysis also revealed some surprises that should provoke further investigations. For example, there was little evidence that female animals showed any beneficial sensory effects as a result of stem cell treatments. Also, the efficacy of the stem cell treatment seemed to not depend on whether immunosuppressive drugs were administered or not.

The authors conclude, “Extensive recent preclinical literature suggests that stem cell-based therapies may offer promise; however the impact of compromised internal validity and publication bias means that efficacy is likely to be somewhat lower than reported here.”

Even though human clinical trials are in the works, such trials will continue to be informed by preclinical studies on laboratory animals.

Stem Cell-Mediated Scarring of the Spinal Cord Aids in Recovery


After injury to the spinal cord, glial cells and neural stem cells in the spinal cord contribute to the formation of the “glial scar.” This glial scar is rich in molecules known as chondroitin sulfate proteoglycans (CSPGs) that are known to repel growing axons. Therefore, the glial scar is viewed as a major impediment to spinal cord regeneration.

However, new work from the Karolinska Institutet in Solna, Sweden has confirmed that the glial scar actually works to contain the damage within the spinal cord. Far from impairing spinal cord recovery, the stem cell-mediated formation of the glial scar confines the damage to a discrete portion of the spinal cord and prevents it from spreading.

Trauma to the spinal cord can sever those nerve fibers that conduct nerve impulses to from the brain to skeletal muscles below the level of spinal cord injury. Depending on where the spinal cord is injured and the severity of the injury, spinal cord injuries can lead to a various degrees of paralysis. Such paralysis is often permanent, since the severed nerves do not grow back.

The absence of neural regeneration required an explanation, since cultured neurons whose axons are severed can regenerate both in culture and in a living creatures (for an excellent review, see Nishio T. Axonal regeneration and neural network reconstruction in mammalian CNS. J Neurol. 2009 Aug;256 Suppl 3:306-9). Thus, neuroscientists have concluded that the injured spinal contains a variety of molecules that inhibit axonal outgrowth and regeneration.

This hypothesis has been demonstrated since many axon growth inhibitors have been isolated from the injured spinal cord (see Schwab ME (2002) Repairing the injured spinal cord. Science 295:1029–1031). Such molecules include proteins like Nogo, Myelin-Associated Glycoprotein (MAG), and Oligodendrocyte-Myelin Glycoprotein (OMgp). However, as the Nishio review points out, axons from severed nerved have been seen growing throughout the central nervous system. Therefore, most of the blame for a lack of regrowth has been pinned on the glial scar.

A new study by Jonas Frisén of the Department of Cell and Molecular Biology and his colleagues has shown that the neural stem cell population in the spinal cord are the main contributors to the glial scar. However, when glial scar formation was prevented after spinal cord injury, the injured area in the spinal cord expanded and more nerve fibers were severed. Furthermore, in their mouse model, a great number of nerve cells died in those mice that did not make glial scars when compared to those mice that were able to produce a normal glial scar.

Ependymal cell incorporation of 5-ethynyl-2′-deoxyuridine is reduced in the absence of Ras genes in intact spinal cord (A and B) and 7 days after injury (C to E). Arrowheads and arrows point to proliferating recombined (A and C) and unrecombined (C and D) ependymal cells, respectively. Injury-induced migration is blocked in rasless ependymal cells (F). Sagittal view of the lesion site 14 weeks after injury in a FoxJ1 control mouse (G) and FoxJ1-rasless mice (H to J). Recombined ependymal cells express YFP in (A) to (D), and cell nuclei are labeled with 4′,6-diamidino-2-phenylindole (DAPI) and appear blue. *P < 0.05, **P < 0.01; Student’s t test. Error bars show SEM. Scale bars represent 10 μm in (A) to (D) and 200 μm in (G) to (J). GFAP, glial fibrillary acidic protein.
Ependymal cell incorporation of 5-ethynyl-2′-deoxyuridine is reduced in the absence of Ras genes in intact spinal cord (A and B) and 7 days after injury (C to E). Arrowheads and arrows point to proliferating recombined (A and C) and unrecombined (C and D) ependymal cells, respectively. Injury-induced migration is blocked in rasless ependymal cells (F). Sagittal view of the lesion site 14 weeks after injury in a FoxJ1 control mouse (G) and FoxJ1-rasless mice (H to J). Recombined ependymal cells express YFP in (A) to (D), and cell nuclei are labeled with 4′,6-diamidino-2-phenylindole (DAPI) and appear blue. *P < 0.05, **P < 0.01; Student’s t test. Error bars show SEM. Scale bars represent 10 μm in (A) to (D) and 200 μm in (G) to (J). GFAP, glial fibrillary acidic protein.

“It turned out that scarring from stem cells was necessary for stabilizing the injury and preventing it from spreading,” said Frisén. “Scar tissue also facilitated the survival of damaged nerve cells. Our results suggest that more rather than less stem cell scarring could limit the consequences of a spinal cord injury.”

According to earlier animal studies, recovery can be improved by transplanting stem cells to the injured spinal cord. These new findings suggest that stimulating the spinal cord’s own stem cells could offer an alternative to cell transplantation therapies.

This paper appeared in the journal Science, 1 November 2013: 637-640, and the first author was Hanna Sabelström. This interesting paper might be leaving one thing out when it comes to spinal cord regeneration.  Once the acute phase of spinal cord injury is completed and the chronic phase begins, the glial scar does in fact prevent spinal cord regeneration.  This is the main reason Chinese researchers have used chondroitinase enzymes to digest the scar in combination with transplantations on stem cells.  By weakening the repulsive effects of the glial scar, these stem cells can form axons that grow through the scar.  Also, olfactory ensheathing cells or OECs seem to be able to shepherd axons through the scar, although the degree of regeneration with these cells has been modest, but definitely real.  Therefore, negotiating axonal regeneration through the glial scar remains a major challenge of spinal cord injury.  Thus, while the glial scar definitely has short-term benefits, for the purposes or long-term regeneration, it is a barrier all the same.

Radio Interview About my New Book


I was interviewed by the campus radio station (89.3 The Message) about my recently published book, The Stem Cell Epistles,

Stem Cell Epistles

It has been archived here. Enjoy.

Biphasic Electrical Stimulation Increases Stem Cell Survival


One of the challenges of stem cell-based therapies is cell survival. Once stem cells are implanted into a foreign site, many of them tend to pack up and die before they can do any good. For this reason, many scientists have examined strategies to improve stem cell survival.

A new technique that improves stem cells survival have been discovered by Yubo Fan and his colleagues at Beihang University School of Biological Science and Medical Engineering. This non-chemical technique, biphasic electrical stimulation (BES) might become important for spinal cord injury patients in the near future.

The BES incubation system. (a) Schematic diagram of a longitudinal section of the incubation chamber including: the upper and lower electric conductive glass plates (FTO glass), a closed silicone gasket, the incubation chamber, and a pair of electrode wires; (b) Schematic diagram of a longitudinal section of the entire BES incubation system including the incubation chamber, the fluid inflow-outflow system, the air filter system, a pair of electrode wires, and a fixed cover and base. Conditions of BES: the NPCs were exposed to 12 h of BES at 25mV/mm and 50mV/mm electric field strengths with a pulse-burst pattern and 8ms pulses (20% duty cycle). Cells that were not exposed to BES served as controls. (A color version of this figure is available in the online journal)
The BES incubation system. (a) Schematic diagram of a longitudinal
section of the incubation chamber including: the upper and lower electric  conductive glass plates (FTO glass), a closed silicone gasket, the incubation
chamber, and a pair of electrode wires; (b) Schematic diagram of a longitudinal
section of the entire BES incubation system including the incubation chamber,
the fluid inflow-outflow system, the air filter system, a pair of electrode wires, and
a fixed cover and base. Conditions of BES: the NPCs were exposed to 12 h of
BES at 25mV/mm and 50mV/mm electric field strengths with a pulse-burst
pattern and 8ms pulses (20% duty cycle). Cells that were not exposed to BES
served as controls. 

Spinal cord injury affects approximately 250,000 Americans, with 52% being paraplegic and 47% quadriplegic. There are 11,000 new spinal cord injuries each year and 82% are male.

Stem cell transplantions into the spinal cord to regenerate severed neurons and associated cells provides a potentially powerful treatment. However, once stem cells are implanted into the injured spinal cord, many of them die. Cell death is probably a consequence of several factors such as a local immune response, hypoxia (lack of oxygen), and probably most importantly, limited quantities of growth factors.

Fan said of his work, “We’ve shown for the very first time that BES may provide insight into preventing growth factor deprivation-triggered apoptosis in olfactory bulb precursor cells. These findings suggest that BES may thus be used as a strategy to improve cell survival and prevent cell apoptosis (programmed cell death) in stem cell-based transplantation therapies.”

The olfactory bulb is in green in this mouse brain.
The olfactory bulb is in green in this mouse brain.

Since electrical stimulation dramatically accelerates the speed of axonal regeneration and target innervation and positively modulates the functional recovery of injured nerves, Fan decided to test BES. His results showed that BES upregulated all the sorts of responses in stem cells that you would normally see with growth factors. Thus BES can increase stem cell survival without exogenous chemicals or genetic engineering.

Fan and his team examined the effects of BES on olfactory bulb neural precursor cells and they found that 12 hours of BES exposure protected cells from dying after growth factor deprivation. How did BES do this? Fan and other showed that BES stimulated a growth factor pathway called the PI3K/Akt signaling cascade. BES also increase the output of brain-derived neurotrophic factor.

“What was especially surprising and exciting,” said Fan, “was that a non-chemical procedure can prevent apoptosis in stem cell therapy for spinal cord patients.” Fan continued: “How BES precisely regulates the survival of exogenous stem cells is still unknown but will be an extremely novel area of research on spinal cord injury in the future.”

BES alters the ultrastructure of NPCs. The ultrastructural morphological changes of cells were investigated by TEM. In the control group (unstimulated), cells had a necrotic appearance: most cells lost the normal cellular structure with a consequent release of cell contents. In the 25mV/mm and 50mV/mm BES groups, the NPCs showed an apoptotic morphology with nuclear fragmentation and condensation
BES alters the ultrastructure of NPCs. The ultrastructural morphological changes of cells were investigated by TEM. In the control group (unstimulated), cells had a necrotic appearance: most cells lost the normal cellular structure with a consequent release of cell contents. In the 25mV/mm and 50mV/mm BES groups, the NPCs showed an apoptotic morphology with nuclear fragmentation and condensation

BES can improve the survival of neural precursor cells and will provide the survival of neural precursor cells and will provide the basis or future studies that could lead to novel therapies for patients with spinal cord injury.

Neural Stem Cells Improve Spinal Injuries in Rats


Disclaimer:  I am reporting on this experiment because of its significance for people with spinal cord-injuries even though I remain appalled at the manner in which the stem cells were acquired.

An international research team has reported that a single set of injections of human neural stem cells had provided significant neuronal regeneration and improvement of function in rats impaired by acute spinal cord injury.

Dr. Martin Marsala, who is professor of anesthesiology at the University of California, San Diego, with colleagues from academic institutions in Slovakia, the Czech Republic, and the Netherlands, used neural stem cells derived from an aborted human fetus to treat spinal cord-injured rats.

Sprague-Dawley rats received spinal cord injuries at the level of the third lumbar vertebra by means of compression. Such injuries render the rats incapable of using their hind legs. They cannot climb a ladder, walk a catwalk or perform other tasks that require the effective use of their hind legs.

The stem cells that were transplanted into the spinal cords of these rats were NSI-566RSC cells, which were provided by the biotechnology company Neuralstem. These cells were initially isolated from the spinal cord of an eight-week old human fetus whose life was terminated through elective abortion. These cells have been grown in culture and split many times. They are a neural stem cell culture that has the capacity to form neurons and glia.

The rats were broken into six groups, and four of these groups received spinal cord injuries. One of these spinal cord-injured groups received injections of were injured NSI-566RSC cells (12 injections total, about 20,000 cells per microliter of fluid injected), another received injections of only fluid, and the third group received no injections. The final spinal cord-injured group of rats received injections of NSI-566RSC cells that had been genetically engineered to express a green glowing protein. Another group of rats were operated on, but no spinal cord injury was given to these animals, and the final group of rats were never operated on.

All rats that received injections of cells were administered powerful drugs to prevent their immune systems from rejecting the administered human cells before the injections (methylprednisolone acetate for those who are interested at 10 mg / kg), and after the stem cell injections (tacrolimus at 1.5 mg / kg).

The results were significant and exciting. In the words of Marsala, “The primary benefits were improvement in the positioning and control of paws during walking tests and suppression of muscle spasticity.” Spasticity refers to an exaggerated muscle tone or uncontrolled spasms of muscles. Spasticity is a serious and common complication of traumatic injury. It can cause severe cramping and uncontrolled contractions of muscles, which increases the patient’s pain and decreases their control.

First, it is clear from several control experiments that the injection procedure did not affect the spinal cord function of these animals, since the sham injected rats had perfectly normal use of their hind limbs and normal sensory function of their limbs. Thus the injection procedure is innocuous. Also, the use of the drugs to suppress the immune response were also equally unimportant when it came to the spinal cord health of the rats.

Two months after the stem cell injections, the rats were subjected to the “catwalk test,” in which the animals walked a narrow path and their paw position was assessed. As you can see in the figure below, the stem cell-injected rats have a paw position that is far more similar to the normal rats than to the spinal cord injured rats.

Improvement in hind paw positioning and muscle spasticity in SCI animals grafted with HSSC. A: CatWalk gait analysis of hind paw positioning at two months after treatment. In comparison to SCI control animals, a significant improvement was seen in HSSC-grafted animals. B1-B3: An example of paw step images taken from the CatWalk software in naïve (B1), SCI-control (B2) and SCI-HSSC-treated animals (B3). Note a large paw footprint overlap between the front and hind paws in naïve animals (B1) but a substantial dissociation in footprint overlap in SCI controls (B2). An improvement in paw placement in SCI-HSSC-treated animals can be seen (B3). C: Statistical analysis showed significant suppression of spasticity response (expressed as a muscle resistance ratio: values at two months versus seven days post injury in ‘HIGH spasticity’ HSSC-treated animals if compared to ‘HIGH spasticity’ controls). D: To identify the presence of muscle spasticity in fully awake animals, the hind-paw ankle is rotated 40° at a velocity of 80°/second. Spasticity is identified by exacerbated EMG activity measured in the gastrocnemius muscle and corresponding increase in muscle resistance. In control SCI animals with developed spasticity (that is, ‘high spasticity’/HIGH group), no change in spasticity response if compared to seven days post-vehicle injection was seen at two months (compare D1 to D3). In contrast to SCI control animals, a decrease in spasticity response was seen in SCI-HSSC-treated animals at two months after cell injections (compare D4 to D6). To identify mechanical resistance, animals are anesthetized with isoflurane at the end of the recording session and the contribution of mechanical resistance (which is, isoflurane non-sensitive) is calculated. (D2, D5: data expressed as mean ± SEM; one-way ANOVAs). ANOVA, analysis of variance; EMG, electromyography; HSSC, human fetal spinal cord-derived neural stem cells; SCI, spinal cord injury; SEM, standard error of the mean.
Improvement in hind paw positioning and muscle spasticity in SCI animals grafted with HSSC. A: CatWalk gait analysis of hind paw positioning at two months after treatment. In comparison to SCI control animals, a significant improvement was seen in HSSC-grafted animals. B1-B3: An example of paw step images taken from the CatWalk software in naïve (B1), SCI-control (B2) and SCI-HSSC-treated animals (B3). Note a large paw footprint overlap between the front and hind paws in naïve animals (B1) but a substantial dissociation in footprint overlap in SCI controls (B2). An improvement in paw placement in SCI-HSSC-treated animals can be seen (B3). C: Statistical analysis showed significant suppression of spasticity response (expressed as a muscle resistance ratio: values at two months versus seven days post injury in ‘HIGH spasticity’ HSSC-treated animals if compared to ‘HIGH spasticity’ controls). D: To identify the presence of muscle spasticity in fully awake animals, the hind-paw ankle is rotated 40° at a velocity of 80°/second. Spasticity is identified by exacerbated EMG activity measured in the gastrocnemius muscle and corresponding increase in muscle resistance. In control SCI animals with developed spasticity (that is, ‘high spasticity’/HIGH group), no change in spasticity response if compared to seven days post-vehicle injection was seen at two months (compare D1 to D3). In contrast to SCI control animals, a decrease in spasticity response was seen in SCI-HSSC-treated animals at two months after cell injections (compare D4 to D6). To identify mechanical resistance, animals are anesthetized with isoflurane at the end of the recording session and the contribution of mechanical resistance (which is, isoflurane non-sensitive) is calculated. (D2, D5: data expressed as mean ± SEM; one-way ANOVAs). ANOVA, analysis of variance; EMG, electromyography; HSSC, human fetal spinal cord-derived neural stem cells; SCI, spinal cord injury; SEM, standard error of the mean.

Secondly, when muscle spasticity was measured, the stem cell-injected rats showed definite decreases in muscle spasticity. The spinal cord-injured rats that received no stem cell injections showed no such changes.

Sensory assessments also showed improvements in the stem cell-treated rats, but the improvements were not stellar. Nevertheless, the stem cell-treated rats progressively improved in their sensory sensitivity whereas the non-treated spinal cord-injured rats consistently showed no such improvement.

Amelioration of hypoesthesia in SCI-HSSC-grafted animals. Baseline and biweekly assessments of perceptive thresholds for (A) mechanical and (B) thermal stimuli, applied below the level of injury, showed a trend towards progressive recovery in SCI-HSSC-grafted animals. C: When expressed as percentages of the maximal possible effect for mechanical and thermal perceptive thresholds improvements, SCI-HSSC-treated animals showed significant improvements in sensory function for both mechanical and thermal components. (A-C: data expressed as mean ± SEM; A-B: repeated measures ANOVAs; C: Student t-tests). ANOVA, analysis of variance; HSSC, human fetal spinal cord-derived neural stem cells; SCI, spinal cord injury; SEM, standard error of the mean.
Amelioration of hypoesthesia in SCI-HSSC-grafted animals. Baseline and biweekly assessments of perceptive thresholds for (A) mechanical and (B) thermal stimuli, applied below the level of injury, showed a trend towards progressive recovery in SCI-HSSC-grafted animals. C: When expressed as percentages of the maximal possible effect for mechanical and thermal perceptive thresholds improvements, SCI-HSSC-treated animals showed significant improvements in sensory function for both mechanical and thermal components. (A-C: data expressed as mean ± SEM; A-B: repeated measures ANOVAs; C: Student t-tests). ANOVA, analysis of variance; HSSC, human fetal spinal cord-derived neural stem cells; SCI, spinal cord injury; SEM, standard error of the mean.

What were the implanted cells doing? To answer this question, Marsala and his co-workers examined tissue sections of spinal cords from the rats implanted with the glowing green stem cells. According to Marsala, the implanted neural stem cells are stimulating host neuron regeneration and partially replacing the function of lost neurons.

Marsala explained: “Grafted spinal stem cells are a rich source of different growth factors which can have a neuroprotective effect and can promote sprouting of nerve fibers of host neurons. We have demonstrated that grafted neurons can develop contacts with the host neurons and, to some extent, restore the connectivity between centers, above and below the injury, which are involved in motor and sensory processing.”

The implanted neural stem cells definitely showed extensive integration with the spinal nerves of the host rats. Again Marsala, “In all cell-grafted animals, there was a robust engraftment and neuronal maturation of grafted human neurons was noted.” Marsala continued: “Importantly cysts or cavities were not present in any cell-treated animal. The injury-caused cavity was completely filled by grafted cells.”

Effective cavity-filling effect by transplanted cells in SCI HSSC-injected animals. At the end of the two-month post-treatment survival, animals were perfusion fixed with 4% PFA, the spinal column dissected and MRI-imaged in situ before spinal cord dissection for further histological processing. A, B: Three-dimensional MRI images of spinal cord segments in animals with previous traumatic injury and treated with spinal HSSC (A) or media (B) injections. Note the near complete injected-cells cavity-filling effect in HSSC-treated animals. A1, A2, B1, B2: To validate the presence of grafted cells or cavitation at the epicenter of injury, the same region was histologically processed, semi-thin plastic sections prepared and compared to the corresponding MRI image (compare A1 to A2 and B1 to B2). C: Two-dimensional MRI image taken from a naïve-non-injured animal. D: Quantification of the cavity and scar volume from serial MRI images showed significantly decreased cavity and scar volumes in SCI-HSSC-injected animals if compared to media-injected SCI controls. (D: data expressed as mean ± SEM; Student t-tests), (Scale Bars: A, B: 5 mm; A1, A2, B1, B2, C: 3 mm). HSSC, human fetal spinal cord-derived neural stem cells; MRI, magnetic resonance imaging; PFA, paraformaldehyde; SCI, spinal cord injury; SEM, standard error of the mean.
Effective cavity-filling effect by transplanted cells in SCI HSSC-injected animals. At the end of the two-month post-treatment survival, animals were perfusion fixed with 4% PFA, the spinal column dissected and MRI-imaged in situ before spinal cord dissection for further histological processing. A, B: Three-dimensional MRI images of spinal cord segments in animals with previous traumatic injury and treated with spinal HSSC (A) or media (B) injections. Note the near complete injected-cells cavity-filling effect in HSSC-treated animals. A1, A2, B1, B2: To validate the presence of grafted cells or cavitation at the epicenter of injury, the same region was histologically processed, semi-thin plastic sections prepared and compared to the corresponding MRI image (compare A1 to A2 and B1 to B2). C: Two-dimensional MRI image taken from a naïve-non-injured animal. D: Quantification of the cavity and scar volume from serial MRI images showed significantly decreased cavity and scar volumes in SCI-HSSC-injected animals if compared to media-injected SCI controls. (D: data expressed as mean ± SEM; Student t-tests), (Scale Bars: A, B: 5 mm; A1, A2, B1, B2, C: 3 mm). HSSC, human fetal spinal cord-derived neural stem cells; MRI, magnetic resonance imaging; PFA, paraformaldehyde; SCI, spinal cord injury; SEM, standard error of the mean.

Marsala’s goal is to used a neuronal stem cell line derived from a patient-specific induced pluripotent stem cell line in a clinical trial. For now, the UC San Diego Institutional Review Board or IRB is reviewing a small phase 1 clinical trial to test the safety and efficacy of this neural stem cell line in patients with spinal cord injuries who have no feeling or motor function below the level of the spinal cord injury.