Our goal is to develop a cellular strategy for repairing the damage seen in children's myelin disease, Multiple Sclerosis and other neurological diseases.

 

Stem Cell Plasticity?

Dr. Jacek Kwiecien writes that Dr. Jonas Frisen questions the current view of stem cell plasticity and gives an insight into the current thinking about the subject. However, he cautions, in vivo experimentation is avoided, therefore the discussion is purely academic if applied research cannot be even addressed.


posted by Canadian Myelin Research Initiative at 12:31 PM

 

Stem Cell Plasticity?

Dr. Jacek Kwiecien says the following article by Dr. Jonas Frisen gives us an idea what is the current status of thinking in the stem cell community. Frisen critically assess, given the last knowledge, the promise of stem cell plasticity. Dr. Kwiecien warns however, that the paper does not address in vivo experimentation and reminds us that the discussion is purely academic if applied research is not addressed.






posted by Canadian Myelin Research Initiative at 12:20 PM

 

Adult-stem-cell research shows some limits

Fri Aug 23, 9:06 AM ET
Elizabeth Weise USA TODAY

Despite earlier research that adult stem cells from bone marrow could successfully change into brain cells, research out today suggests the change isn't as easy as once believed. That could put a damper on hopes that adult cells might take the place of those from embryos in research to cure brain diseases and spinal cord injuries.


posted by Canadian Myelin Research Initiative at 1:10 AM

 
Dear Wayne,

I trust this not finds you and your family well. I apologize for not writing to you earlier. As you may know, I retired from the UBC as of December 31, 2001and am emeritus professor of neurology and allowed to keep my laboratory at the UBC Hospital. I am currently spending more time in Suwon, Korea working mostly in the area of immortalized human neural stem cells.

Most recently my collaborators in Japan were successful in treating mucopolysaccharidoses VII mice with our human neural stem cells (over-expressing beta-glucuronidase gene). The manuscript was submitted to the journal Gene Therapy a couple of weeks ago. I attach title and abstract pages of the paper for your reference. We have produced immortalized human neural stem cells (ours carry normal human karyotype and express neural stem cell markers) transfected with tyrosin hydroxylase/ GTP CH genes (for Parkinson disease), choline acetyltransferase (for spinal cord injury and ALS) and also with GDNF or BDNF (for stroke, Alzheimer and other neurological disorders). We were also successful in generating a large number of neurons from the stem cells by introducing a master gene for neurogenesis (NeuroD) into our immortalized neural stem cells.

I will be in Vancouver until the middle of September.

With kindest regards, Seung Kim, MD

Brain transplantation of genetically engineered human neural stem cells correct
lysosomal storage in mucopolysaccharidosis VII mouse


Xing-Li Meng1, Jin-Song Shen1, Toya Ohashi1,2, Seung U. Kim3,4 and Yoshikatsu Eto1,2
1Department of Gene Therapy, Institute of DNA Medicine, 2Department of Pediatrics, The Jikei University School of Medicine, Tokyo, Japan; 3Brain Disease Research Center, Ajou University, Suwon, Korea and 4Division of Neurology, Department of Medicine, University of British Columbia, Vancouver, Canada.

Corresponding author: Toya Ohashi, M.D.
Department of Gene Therapy, Institute of DNA Medicine
The Jikei University School of Medicine
Japan

Summary

Therapeutic potential of human neural stem cells (NSCs) in neurological disorders was investigated in a mouse model of human mucopolysaccharidoses type VII (MPSVII) that is caused by b -glucuronidase deficiency. An immortalized human NSC line, HB1.F3, was transduced retro virally to overexpress human b -glucuronidase in vitro, and transplanted into the brain of newborn MPSVII mice intraventricularlly. In NSC-grafted MPSVII mouse brain at 25 days post-transplantation, a number of donor human NSCs were identified in the ventricular zone and in brain parenchyma, and increased level of b -glucuronidase enzyme activity was detected as compared to non-transplanted MPSVII brains. The levels of glycosaminoglycans, substrates of b -glucuronidase, in NSC-grafted MPSVII brains were significantly decreased to 50% of those in non-transplanted MPSVII brains and close to those of wild type normal mouse brains. Widespread clearance of lysosomal storage was observed in various brain regions of the MPSVII mice receiving NSC grafts. These results demonstrate that the human NSCs over expressing b -glucuronidase are capable of correcting lysosomal storage in the mutant mouse brain caused by deficiency of the enzyme following brain transplantation and should prove that the human NSCs would serve as an ideal gene transfer vehicle for the treatment of diffuse CNS lesions in human lysosomal storage diseases.

Keywords: beta-glucuronidase, brain transplantation, gene transfer, human neural stem cells,
mucopolysaccharosis VII, mutant mouse, Sly syndrome
Introduction

The mammalian central nervous system (CNS) harbors neural stem cells (NSCs) throughout development and in the adulthood.1-3 These CNS primordial cells showed the capacities of self-renewal, multipotency and remarkable plasticity in vivo and in vitro.4-7 NSCs should serve as powerful tools in the study of normal development of the CNS, the mechanisms involving neural integration and processes of brain repair and in cell replacement and gene transfer for treatment of patients with neurological disorders.
Previous studies have reported that murine NSCs were utilized for cell replacement/ gene transfer therapies in animal models of neurological diseases; murine NSCs showed robust migration in the recipients� brain after transplantation, produced deficient enzyme8 and formed myelin or replaced the lost neurons in the hosts by shifting their differentiation mode to a certain neural cell lineage.9,10 Neuropathological and functional improvements were also observed in some of these experimental animals.8,9 Human NSCs, as their murine counterparts, respond to developmental cues, differentiate in vivo after grafted into the adult rat CNS11,12 or the fetal primate cerebral germinal zone13 and express foreign genes after transplanted into neonatal mouse brain14 Recent studies have demonstrated that human NSCs grafted into the brain of parkinsonian rats survive long-term and produce clinical improvement,15 successful delivery of b -glucuronidase gene in adult nude mouse,16 and functional improvement in aged rat brain.17 In the present study, we wish to investigate if brain transplantation of human NSCs can efficiently deliver therapeutic genes and improve diffuse neuropathology of the CNS in mouse model of mucopolysaccharidosis type VII.

Mucopolysaccharidoses type VII (MPSVII), also referred as Sly syndrome,18 results from the deficiency in enzyme activity of b -glucuronidase (GUSB, EC.3.2.1.31), a tetrameric lysosomal acid hydrolase, and is inherited as an autosomal recessive trait. The subsequent lysosomal storage of undegraded substrates, glycosaminoglycans (GAGs), in most tissues leads to cellular and organ dysfunction. The murine MPSVII, an authentic model of human MPSVII, shares many of the pathological and clinical features of human MPSVII. 19-21 MPSVII mouse shows progressive abnormalities such as dysmorphic face, poor mobility, skeletal deformities, dwarfism, corneal clouding and a shortened life span. The well-described neuropathology in the MPSVII mouse brain and the convenience in performing biochemical, histochemical and pathological analysis22 make murine MPSVII an ideal system in the assessment of therapeutic efficacies of novel gene transfer and cell replacement approaches to the CNS.

In the present study, human NSC line, HB1.F3, immortalized with v-myc,23 was transduced with a retrovirus carrying human b -glucuronidase cDNA and transplanted into the brain of newborn MPSVII mice. Human NSCs showed the excellent therapeutic potential by serving as an efficient gene transfer vehicle and improving neuropathology of diffuse CNS lesions in the MPSVII mouse brain.

posted by Canadian Myelin Research Initiative at 12:41 PM

 

The cells set up shop and built a new immune system that didn't attack his myelin.

Nobody knows what causes MS, but it seems triggered when the body becomes allergic to itself and the misdirected immune system attacks the central nervous system--specifically, the myelin sheaths, the thin layers of fatty cells that wrap around and insulate nerve fibers of the brain and spinal cord.

Repeated attacks bring about a loss of the protective coating of myelin, which halts nerve cells from communicating much the way that frayed electrical wires short out and stop carrying current. MS is a crippler, not usually a killer, but the progression of the disease cannot be predicted and depends on the individual.

Northwestern Memorial has pioneered stem cell transplants for this and other devastating neurological disorders. Dr. Richard K. Burt, the hospital's director of immunotherapy, performed the first procedure in the U.S. in 1997. Based on the program's success so far, the National Institute of Allergy and Infectious Diseases awarded the hospital a $9.2 million contract for further research on stem cell transplants for autoimmune diseases.

posted by Canadian Myelin Research Initiative at 11:39 PM

 
Dr. Jacek Kwiecien writes, "Needless to say, the optimism transpiring from this editorial is not supported by new knowledge and the fact that between the cells in the dish and the human patients there is a missing link which is appropriate animal model(s). This is especially true for the myelination research.

Versatile Cells Against Intractable Diseases

Science Online - Constance Holden


If popular accounts are to be believed, these versatile cells hold cures for a variety of ailments, chief among them neurological disorders such as Parkinson's and Alzheimer's diseases. With such publicity, it's not surprising that patients are clamoring for treatments that are as yet barely conceptualized in the lab. "The expectation of my patients is that this will be ready tomorrow or in a year," says Jeffrey Rothstein, who does research on amyotrophic lateral sclerosis (ALS) at Johns Hopkins University in Baltimore. In reality, he and other scientists say, even without political obstacles, the closest treatments are years away, and some will take decades.

ALS
At Project ALS--a privately funded venture at Harvard, Cornell, Columbia, and Johns Hopkins universities--collaborators are trying to find out which types of cells would be best. The Johns Hopkins group is focusing on the potential of a line of pluripotent stem cells, called embryoid body-derived (EBD) cells, derived from the germ cells of aborted fetuses. Johns Hopkins researchers generated considerable excitement last year when they showed that EBD cells were able to restore some mobility to rats with paralyzed hindlimbs. Unlike ES cells, these do not appear to trigger tumor formation, says Rothstein. What's more, says Dawson, ethical problems are minimal: "You would probably only need one fetus to treat tens of thousands of patients."

At Harvard, Evan Snyder of Children's Hospital and Beth Israel Deaconess Medical Center and Robert H. Brown of Massachusetts General Hospital in Boston are comparing not only EBD cells but also fetal spinal progenitor cells, umbilical cord blood cells, human adult stem cells called fibroblasts, a line of stem cells derived from fetal brains, mouse neural stem cells, fetal pig neurons (which resemble human neurons), and even skin cells (fibroblasts). These cells, representing varying stages of development, are being injected in an ALS mouse model to see which type does best in producing or rescuing motor neurons.

Although no treatment is yet in sight, the Johns Hopkins researchers are already testing the safety of EBD-derived neurons in 28 African green monkeys. "We meet constantly with FDA [the Food and Drug Administration]" to be sure all the rules are being followed, says Rothstein, so researchers can move quickly into clinical trials if a promising treatment emerges.

Multiple sclerosis
Stem cells might ultimately provide some benefit, albeit limited, to MS patients. The problems are multiple. First, MS is both an autoimmune and a neurological disorder. Finding the right type of precursor cell for therapy might be relatively simple, because the disease attacks not the neurons but the tiny cells called oligodendrocytes that make the protective coating of myelin on axons, the long fibers that conduct impulses from cell bodies. Because they're homogeneous and easy to identify, Steven Goldman of Cornell University believes they will be one of the first neural cell types to be used in stem-cell therapy.

But damage sometimes extends beyond the myelin sheaths to the underlying neurons. Furthermore, "adding cells may be adding fuel to the fire, unless the underlying inflammatory process is approached," says Goldman. For this reason, congenital myelin diseases might be easier targets, he says.

Safety
Even if scientists can conquer the complexities and create replacement neurons from stem cells, huge regulatory hurdles remain. ES cells are the most daunting: Scientists must devise ways to cultivate them that don't involve contact with potentially contaminating mouse cells. Geron Corp. in Menlo Park, California, which is working to create dopamine-producing cells, says it has already developed a culture medium free of mouse cells.

More difficult will be ensuring that the cells are safe, because a single remaining undifferentiated cell could lead to tumor formation in the patient. Dawson of Johns Hopkins speculates, for example, that the FDA "may want us to engineer in a kill switch" so introduced cells could be turned off if they started misbehaving after transplantation. Then there is the problem of overcoming a patient's immune response to foreign cells, although researchers say this is not a major obstacle with the brain, which has been called "immune privileged."

In short, there's a long road ahead. Nonetheless, says Isacson, "I am still surprised at how much remodeling is going on in the brain." He is confident that someday researchers can learn how to tap into that plasticity.

.



posted by Canadian Myelin Research Initiative at 11:43 AM

 
Parthenogenetic embryo

US scientists have given details of how they managed to get the unfertilised eggs of monkeys to start dividing like embryos - a process known as parthenogenesis - and then "harvest" them for special cells.

Dr. Hollands reports, "Primate parthenogenesis is now well described in the literature and I did some experiments many years ago which indicated the human potential of the technique. Further work is needed but it is defintiely a potential source of stem cells. On the ethical side we would have to wait and see what the committees came up with. The embryos cannot go to term so the rules for them should surely be different".

posted by Canadian Myelin Research Initiative at 11:06 PM

 
Dr. Hollands discusses Embryonic stem cells: Where to next?

Another barrier which must be crossed is the potential problem which lies in the number of embryonic stem cells which would be required to provide a successful transplant into one recipient. It is well known that it is often difficult to obtain sufficient cells for a transplant from an umbilical cord blood sample. The problem will be amplified when using embryonic cells unless the properties of the embryonic cells are found to be very different. It may therefore be necessary to amplify embryonic stem cell numbers in vitro prior to transplantation perhaps using cytokines. Such studies on adult stem cells have had varying levels of success and this is an area which will require extensive research prior to any clinical applications.

posted by Canadian Myelin Research Initiative at 10:59 PM

 
Dr. Peter Hollands worked with Dr. Robert G. Edwards in the 1980's laying the groundwork for the current studies on embryonic stem cells. Drs Edwards and Patrick C. Steptoe of Cambridge pioneered in vitro fertilization techniques that led to the birth of the first �test tube baby� in 1978 in Great Britain. In addition, Dr. Hollands has worked extensively on adult stem cells. He plans to resume his embryonic stem cell research. CMRI is considering collaboration.

Dr. Hollands intends to derive the cells from human embryos donated to research in the IVF clinic in which he practices. Moreover, Dr. Hollands is keen to look at the potential of therapeutic cloning and parthenogenetic activation as possible sources of embryos for stem cell derivation. Parthenogenesis in particular may be useful allaying the reservations of ethicists and legal constraints, placed on human embryo research in the UK. . Interested in spinal cord repair following damage and also the potential of these cells in the treatment of conditions such as Parkinsonism, Dr. Hollands intends to apply for human trials. Another interest is re-myelination
processess applicable to multiple sclerosis.


"I have a very open mind regarding applications and strongly believe in the enormous potential of these neuronal stem cells" he says.


posted by Canadian Myelin Research Initiative at 9:48 PM

 
While this is very good news for moving toward human clinical trials similar issues will need to be addressed regarding the human platform. Ethics Committees will still, I believe, tread cautiously concerning the testing of such cellular lines. Diseases like CJD (Mad Cow) and HIV viruses will remain a concern in any clinical trial and will most certainly elevate the cost.

Yahoo! News - First Fully Human Embryonic Stem Cell Line Created

The standard technique for creating human embryonic stem cell lines has been to extract cells from an embryo and grow them atop embryonic mouse cells, known as "feeder" cells, which excrete nutritional or growth factors that sustain the cell lines. But this close association with mouse cells raises safety questions, as the cell lines could transfer animal viruses to people.


posted by Canadian Myelin Research Initiative at 4:40 PM

 
Dr. Jacek Kwiecien of McMaster University, Hamilton, Ontario, Canada writes:

I also have been active in re-myelination work and determined that neurospheres from the subependyma in adult rat and adult mouse are a poor source of myelinating cells. Myelin sheaths which I could find in the LES rats transplanted and then maintained for 1 month, were well compacted and without any abnormalities indicating degeneration. Also, immune response was not observed in LES rats transplanted with either mouse or rat neurospheres. This work was presented as a poster during the IXth International Symposium on Neural Regeneration in Pacific Grove in December 2001.

Presently, students in Dr. Doering�s lab are isolating neurospheres from neonatal mouse and I will transplant them into LES rats in mid-July. Neonatal rat neurospheres were transplanted into 10 LES rats 6 weeks ago for 1 month and I am waiting for the spinal cords from 5 rats to be sectioned to examine degree of migration of these cells. Other 5 rats' spinal cord will be examined electron microscopically for quality of myelin sheaths. This work is to develop a model of robust re-myelination in the LES rat to use it as a testing method for growth factors and drugs enhancing formation of myelin. With Dr. Doering's collaboration I will submit the grant proposal to NSERC to continue on this basic research.

In the meantime, I would like to repose that human cell candidates should be tested on the ES rats for their ability to myelinate. Since Dr. Snyder's cells are still not available to me, a major contamination made it impossible to transfer them from Harvard to McMaster back in February and I have not heard from Evan and his lab co-workers that situation improved.

Therefore, I would like to obtain a line of human embryonic stem cells from University of Wisconsin to test the ability of these cells to form myelin in the LES rat.



posted by Canadian Myelin Research Initiative at 4:24 PM