Effects of sporoderm-broken germination activated ganoderma spores on promotion of proliferation and/or differentiation of neural stem cells in injured spinal cord
The present invention provides a method for promoting proliferation and/or differentiation of neural stem cells in a mammal, which comprises orally administering sporoderm-broken germination activated Ganoderma lucidum spores (GASP) to the mammal. The effects of GASP on proliferation and/or differentiation of neural stem cells are particularly prominent in mammals after a spinal cord injury.
The present invention is a Continuation-In-Part (CIP) Application of U.S. patent application Ser. No. 10/631,809, filed on Aug. 1, 2003, which is herein incorporated by reference.
FIELD OF THE INVENTIONThe present invention relates to a method for promoting proliferation and/or differentiation of neural stem cells in a mammal, particularly after the mammal has a spinal cord injury, by administering to the mammal sporoderm-broken germination activated ganoderma spores (GASP).
BACKGROUND OF THE INVENTIONThe spinal cord coordinates the body's movement and sensation to and from the brain. It is a complex organ containing nerve cells (also known as neurons), and supporting cells (also known as neuroglial cells or glial cells). A typical neuron consists of a cell body, containing the nucleus and the surrounding cytoplasm (perikaryon); several short radiating processes (dendrites); and one long process (the axon), which terminates in twiglike branches (telodendrons) and may have branches (collaterals) projecting along its course. The axon together with its covering or sheath forms the nerve fiber. The neuroglial or glial cells form the supporting structure of nervous tissue. There are three types of glial cells, which are astrocytes, oligodendrocytes, and microglia. Astrocytes and oligodendrocytes (collectively macroglia) are of ectodermal origin. These cells far outnumber neurons in the brain and spinal cord and perform many essential functions. The oligodendrocyte creates the myelin sheaths that insulate axons and improve the speed and reliability of nerve signal transmission. Astrocytes, large star-shaped glial cells, regulate the composition of the fluids that surround neurons. Some of these cells also form scar tissue after injury. Microglia are of mesodermal origin. They are smaller cells that become activated in response to injury and help clean up waste products. All of these glial cells produce substances that support neuron survival and influence axon growth.
Many axons in the spinal cord are covered by sheaths of an insulating substance called myelin, which gives them a whitish appearance; therefore, the region in which they lie is called “white matter.” The neurons themselves, with their tree-like dendrites that receive signals from other neurons, make up “gray matter.” This gray matter lies in a butterfly-shaped region in the center of the spinal cord. Like the brain, the spinal cord is enclosed in three membranes (meninges): the pia mater, the arachnoid, and the dura mater. The spinal cord is then surrounded by rings of bone called vertebra.
The spinal cord and the brain together make up the central nervous system (CNS). Unlike neurons of the peripheral nervous system (PNS), which carry signals to the limbs, torso, and other parts of the body, it was generally believe that new neurons could not be generated in the adult mammalian brain and neurons of the CNS do not regenerate after injury. More recently, findings that neurons can be renewed in certain regions of the adult CNS, e.g., in the olfactory bulb, where signals from neurons from the organ of smell reach the brain and in the dentate gyrus of hippocampus, have been reported. Since neurons are unable to divide, the addition of new neurons suggested the existence of immature cells, i.e., progenitor or stem cells, which may generate neurons.
Stem cells are undifferentiated cells capable of proliferation, self-maintenance, and production of a large number of differentiated, functional progeny, regenerating tissue after injury. Stem cells isolated from the early embryonic blastula, i.e., prior to gastrulation, can produce cell types of all different lineages (Keller, Curr. Opin. Cell Biol., 7:862-869 (1995)). Stem cells in adults, however, play the role of replacing cells which have been lost by natural cell death, injury or disease. When transplanted into the brain, these neural stem cells survived and differentiated into neurons and neuroglial cells. (Johansson et al., Cell (1999), 96(1): 25-34). There are also certain numbers of neural stem cells in the spinal cord. They are proliferating cells in the ependyma of the spinal central canal in adult mammals. (Frisen et al., J. Cell Biol. (1995), 131:453-464; Yamamoto et al., Exp. Neurol. (2001), 172:115-127)). A method for isolation of ependymal neural stem cells have recently been described in U.S. Pat. No. 6,541,247.
The existence of neural stem cells in the adult mammalian CNS was first demonstrated by culturing cells from the adult rat brain and spinal cord. Under certain culture conditions, a population of multipotent neural stem cells can be propagated. (Reynolds et al., Science (1992) 255:1707-1710). Under these conditions, single cells proliferate in vitro and the progeny forms a cluster of aggregated cells. Such cell clones detach from the culture dish after a few days in vitro. The cells continue to proliferate and form a characteristic spheroid cell aggregate, referred to as a neurosphere, of tightly clustered cells, all of which are derived from a single cell. Most of the cells in the neurosphere express nestin, an intermediate filament found in neuroepithelial stem cells, (Lendahl et al., Cell (1990), 60:585-595), but not markers typical for differentiated cells. These undifferentiated cells rapidly differentiate if plated on an adhesive substrate or if serum is added to the culture medium. Importantly, a clone of cells derived from a single cell can generate neurons, astrocytes and oligodendrocytes, demonstrating that at least the initial cell was multipotent (Reynolds et al., Science, ibid.). Moreover, if a cell clone is dissociated, many of the cells will form new clusters of undifferentiated multipotent cells, thus fulfilling the criteria for being stem cells.
The development of the mammalian CNS begins in the early stage of fetal development and continues until the post-natal period. (U.S. Pat. No. 6,497,872). The first step in neural development is cell birth, which is the precise temporal and spatial sequence in which stem cells and stem cell progeny (i.e., daughter stem cells and progenitor cells) proliferate. Proliferating cells will give rise to neuroblasts, glioblasts and new stem cells.
The second step is a period of cell type differentiation and migration when undifferentiated progenitor cells differentiate into neuroblasts and gliolblasts which give rise to neurons and glial cells and migrate to their final positions. Cells which are derived from the neural tube give rise to neurons and glia of the CNS, while cells derived from the neural crest give rise to the cells of the peripheral nervous system (PNS). Certain factors present during development, such as nerve growth factor (NGF), promote the growth of neural cells. NGF is secreted by cells of the neural crest and stimulates the sprouting and growth of the neuronal axons.
The third step in development occurs when cells acquire specific phenotypic qualities, such as the expression of particular neurotransmitters. At this time, neurons also extend processes which synapse on their targets. Neurons are generated primarily during the fetal period, while oligodendrocytes and astrocytes are generated during the early post-natal period. By the late post-natal period, the CNS has its full complement of nerve cells.
The final step of CNS development is selective cell death, wherein the degeneration and death of specific cells, fibers and synaptic connections “fine-tune” the complex circuitry of the nervous system. This “fine-tuning” continues throughout the life of the host. Later in life, selective degeneration due to aging, infection and other unknown etiologies can lead to neurodegenerative diseases.
There is increasing evidence that nervous system injuries may affect stem cells in the adult CNS. After both spinal cord and brain injuries, nestin expression is increased in cells lining the central canal and in the subventricular zone, respectively. (Frisen et al., J. Cell Biol., Ibid.). These cells may be derived from stem cells. With time, nestin expressing cells are seen progressively further from the central canal and the lateral ventricle and these cells express astrocytic markers. (Frisen et al., J. Cell Biol., ibid.). These data demonstrate that stem cells or progenitor cells residing by the ventricular system are induced to proliferate, migrate toward the site of the injury and differentiate to astrocytes.
Currently, no methods are available in clinical practice to stimulate generation of new cells in the nervous system. Transplantation of cells from human embryos or animals have been tested clinically with some encouraging results. However, these methods have several problems, mainly ethical and immunological, which makes it very unlikely that they will be used in any larger number of patients. Accordingly, the discovery of a proliferation and differentiation factor on neural stem cells in the adult CNS of mammals is important and may make it possible to develop strategies to stimulate generation of new neurons or glial cells.
Recently, Cheung et al., FEBS Lett. (2000), 486: 291-296, disclose that Ganoderma lucidum extract induces neuronal differentiation in vitro, using a primary neuronal cell system (i.e., PC12 cells). The PC12 cells are derived from rat pheochromocytoma cells which respond to the nerve growth factor (NGF). The Ganoderma lucidum extract described by Cheung et al. is made from the fruit bodies of the Ganoderma lucidum.
In the invention to be presented in the following sections, a novel use of sporoderm-broken germination-activated Ganoderma spores (GASP) from Ganoderma lucidum as an effective, safe and practical alternative to induce the proliferation of self neural stem cells of the injured CNS and migration to the injured area and further differentiation into neurons for replacement of damaged or lost neurons is described. In the parent application of the instant invention, we have described the novel use of GASP in promoting the motor neuron survival and axon regeneration and its use in treatment of related spinal cord injury. The GASP have also previously been disclosed for use in treating patients with cancer, AIDS, hepatitis, diabetes, and cardiovascular diseases, and can prevent or inhibit free radical oxidation and hepatotoxic effects. See U.S. Pat. Nos. 6,316,002 and 6,468,542, which are incorporated herein by reference. A further benefit of using the GASP is that they are non-toxic so that higher dosage can be prescribed to the patients.
SUMMARY OF THE INVENTIONThe present invention provides a method for promoting proliferation and/or differentiation of neural stem cells in a mammal, by administering an effective amount of sporoderm-broken, germination activated Ganoderma spores (GASP) into a mammal, preferably human. Gandoderm (Ganoderma lucidum Leyss ex Fr. Karst) is a polyporous fungus. It belongs to the class of Basidiomycetes, the family of Polypolaceae, and the genus of Ganoderma. The GASP is especially effective on promoting proliferation and/or differentiation of neural stem cell after the mammal has a spinal cord injury. The causes for a spinal cord injury include, but are not limited to, compression or severance of the spinal cord, trauma (such as car accident, violence, falls, sports etc.), or a disease (such as polio, spina bifida, or Friedreich's Ataxia). In addition, the spinal cord injury can be due to damage or death of neurons within the injured spinal cord or crush of axons within the injured spinal cord. The GASP are preferred to be orally administered to a mammal, including a human, in the amount of about 0.5-15 g per kg of body weight per day, most favorably about 8 g per kg of body weight per day.
BRIEF DESCRIPTION OF DRAWINGS
The present invention provides a method for promoting the proliferation and/or differentiation of neural stem cells in a mammal, particularly after a spinal cord injury. The method involves administering an effective amount of germination activated Ganoderma spore powder (GASP) into a mammal having spinal cord injury.
Spinal cord injury has been a serious problem in humans, which, in many case, leads to complete or incomplete loss of motion or sensory function. The seriousness of spinal cord injury is primarily due to the difficulty of recovery from injury because the neurons in the spinal cord are very specialized and unable to divide and create new cells. In the United States, approximately 450,000 people live with spinal cord injury, and there are about 10,000 new cases every year, mostly involving males between ages of 16 to 30. Although spinal cord is well protected, spinal cord injury does occur due to causes such as trauma (e.g., car accident, violence, falls, sports) and disease (e.g., polio, spina bifida, Friedreich's Ataxia).
Ganoderma (Ganoderma lucidum Leyss ex Fr. Karst) is a polyporous fungus which belongs to the class Basidiomycetes, the family Polypolaceae, and the genus Ganoderma. Ganoderma spores are tiny and mist-like spores of 5˜8 μm in sizes which have extremely hard and resilient, double-layer epispores, thus making them difficult to break open. The spores contain high concentrations of many bioactive substances, including, but are not limited to, polyunsaturated fatty acids, polysaccharides, vitamins, sterols, trace minerals, amino acids, and triterpenes. The GASP used in the present invention are sporoderm-broken (i.e., the double-layer epispores of the spores are broken so that the bioactive substances within the spores are released), which is produced by the method described in U.S. Pat. No. 6,316,002 (“the '002 patent). The entire content of the '002 patent is herein incorporated by reference. Through the unique spore-breaking method described in the '002 patent, the bioactive substances within the GASP are recovered in high yields and the functional activities of the bioactive substances are successfully preserved.
As shown below is a general description of the method used in the '002 patent, which leads to the production of the GASP:
I. Soaking to induce germination: Mature and perfect spores of Ganoderma lucidum were carefully selected to carry out a soaking process to induce germination. Spores were kept in clear or distilled water, biological saline solution, or other nutritional solutions that could enable the spores of Ganoderma lucidum to germinate rapidly. Examples of nutritional solutions include coconut juice or a 1-5% malt extract solution, 0.5-25% extracts of Ganoderma lucidum sporocarps or Ganoderma lucidum capillitia, 0.1-5% of culture solution containing biotin, 0.1-3% of culture solution containing monobasic potassium phosphate and magnesium sulfate. The choice of solution would depend on the soaking time required, the amount of spores to be processed and other such factors as availability of materials. One or more of the above germination solutions could be used, with the amount added being 0.1-5 times the weight of the spores of Ganoderma lucidum. The soaking time can be determined according to the temperature of the water, and usually the soaking was carried out for 30 min to 8 h with the temperature of the water at 20-43° C. Preferably soaking times were 2-4 hours, and temperature of the water was 25-35° C.
II. Activation culture: The spores of Ganoderma lucidum were removed from the soaking solution and excess water was eliminated by allowing it to drip. The spores were then placed in a well-ventilated culturing box at a constant temperature and humidity so that spore activation culture could be carried out. The relative humidity of the culture was generally set at 65-98%, the culture temperature at 18-48° C. and the activation time lasted from 30 min to 24 h. Preferably humidity is 85-97% and temperature is 25-35° C. Using the method provided by the present invention, the activation of spores of Ganoderma lucidum reached a rate of more than 95%. During activation, the cell walls of the spores of red Ganoderma lucidum were clearly softened such that it was easier to penetrate the cell walls of the spores.
III. Treatment of the epispores: After the germination activation process, the spores were treated by enzymolysis. This process was carried out at a low temperature and under conditions such that enzyme activity was maintained, using chitinase, cellulase, or other enzymes, which are commonly used in the industry. The process was complete when the epispores lost their resilience and became brittle. Alternatively, physical treatments were carried out to penetrate the cell walls, for example, micronization, roll pressing, grinding, super high pressure microstream treatment, and other mechanical methods commonly used in the industry could be carried out, with a penetration rate of over 99%.
IV. Drying or extraction: Drying was carried out at low temperature using standard methods including freeze-drying or vacuum-drying etc., which are commonly used in the industry. The obtained product had a moisture content less than 4%. After drying, the bioactive substances were extracted by water or alcohol, or by thin film condensation. The extracted bioactive substances could be further purified by dialysis to ensure no contamination in the final products.
V. Pharmaceutical formulations of the bioactive substances: The bioactive substances can then be made into purified powders, extract pastes, solutions for injection, or for oral consumption. The invention also encompasses the manufacture of pharmaceutical preparations of the active substances, using well-known expedients and methods of manufacture known in the art. In addition, the bioactive substances can be dosed by any convenient method including tablets, capsules, solutions, suppositories, nasal sprays, paranterals, or injection devices. The choice of method for administration is determined by the clinical situation of the patient. The bioactive substances of the present invention, produced by the methods described, include active genes, inducers of the biotic potential promotor, inducers of the multicellular activator, inducers of interferon, lactone A, ganoderma polysaccharide, ganoderma spore fatty acids, ganoderma spore long chain alkyl hydrocarbon, ganoderma triterpenes, sterols, superoxide dismutase, vitamin E, active glycoprotein, certain growth factors, ganoderma acid A, superoxide dismutases (SOD), active glycoproteins, multiple active enzymes, and growth factors and so on. These bioactive substances, in a whole, contribute to the therapeutic uses described in the later sections.
GASP are non-toxic. The preferred method for administering GASP is through oral uptake. Currently, GASP are approved by the Food and Drug Administration (FDA) to be used as dietary supplement in the capsule form under the name of Enhanvol® and Holistol, sold by Enhan Technology Holdings International Co., Ltd. in Hong Kong. Each capsule of GASP contains 0.3 g of GASP. The recommended dosage of GASP, when used as dietary supplement, is 4 times every day, 4 capsules each time. Thus, for an adult of 60 kg, the daily dosage of GASP as dietary supplement is at about 0.08 g/kg of body weight per day.
It has been shown, however, that no physiological and pathological abnormalities were found when 8 g/kg/day of GASP were given to patients and animals. 0.5 g to 15 g/kg/day of GASP have been given to animals and demonstrated significant effects on treatment of spinal cord injury. However, it is understood that the dosage for any particular patient depends upon a variety of factors, including age, body weight, general health, sex, diet, time of administration, route of administration, rate of excretion, drug combination and the severity of the disease. For these reasons, dosing is left to the discretion of the skilled clinician.
The present invention is conducted using a rat model. According to Metz et al., J Neurotrauma (2000), 17(1):1-17), which is herein incorporated by reference, the functional, electrophysiological and morphological outcome parameters following spinal cord injury in rats can be extrapolated for those in humans. Metz et al. collected data from human patients with chronic spinal cord injury and compared them to those of rats with contusion spinal cord injury induced by a weight-drop. The results suggest an analogous relationship in rats and humans with respect to functional, electrophysiological, and morphological outcomes, which demonstrates that rat can serve as an adequate animal model for research on functional and morphological changes after spinal cord injury and the effects of new treatment strategies.
In the present invention, five (5) fluorescent neruonal markers were used, which are BrdU, nestin, neurofilament (NF), oligodendrocytin, and glial fibrillary acidic protein (GFAP), to explore the proliferation of cells in general (based on the staining of BrdU), the undifferentiated neural stem cell (based on the staining of nestin), the newly generated neurons from neural stem cell (based on the staining of NF), the newly generated oligodendrocytes from neural stem cell (based on the staining of oligodendrocytin), and the newly generated astrocytes (based on the staining of GFAP).
The following experimental designs are illustrative, but not limiting the scope of the present invention. Reasonable variations, such as those occur to reasonable artisan, can be made herein without departing from the scope of the present invention.
Experiment 1 Preparation of Spinal Hemisection Animal Model, Injection of BrdU, and Treatment of GASPAnimal grouping. Twenty-six (26) female SD rats (150-180 g) were purchased from the Experimental Animal Center of the Sun Yat-sen University. The animals were separated into 3 groups: the normal control group (2 rats), the injured/control group (12 rats: 2 rats in the 1-day group, 5 rats each in the 2-week and 4-week groups) and the injured/treatment group (12 rats: 2 rats in the 1-day group, 5 rats each in the 2-week and 4-week groups).
Animal Model preparation. 1% Pentobarbital sodium (35˜40 mg/kg) was injected intraperitoneally to anesthetize the rats in the injured/control group and the injured/treatment group. The skin on the back of the rat was cut open at the centerline. The thoracic (T) 12 spinal segment was exposed under a surgical microscope and hemisection of the right side of the spinal cord was performed. After surgery, penicillin was injected intramuscularly.
BrdU injection and treatment: The animals in all three groups received intraperitoneal injection of BrdU (50 mg/kg) twice a day for consecutive 10 days. After the spinal injury, the animals in the injured/treatment group were administered twice a day via stomach a GASP solution at 8 g/kg/day. The injured/control group was administered via stomach a 5% sodium carboxycellulose solution (i.e., the solvent in the GASP solution). The GASP is provided by Enhan Technology Holdings International Co., Ltd. The thimidine analogue, 5-BrdU, is commonly used to study DNA synthesis and cell proliferation. It is a nonradioactive molecule. BrdU is substituted stoichiometrically for thymidine in DNA during the S-phase (synthesis) of cell growth where the cellular DNA content is doubling between the G1 and G2 cell phases. A measure of the incorporated BrdU is related to the new DNA content, i.e., the status or location of the cell in the S-phase. Thus, the detection of BrdU-substituted DNA is functionally related to synthesis of DNA during the S-phase of the cell cycle.
Experiment 2 Perfusion, Fixation and SamplingAt 1 day, 2 weeks, and 4 weeks after the spinal injury, the animals in the corresponding groups were anesthetized by an intraperitoneal injection of 1% pentobarbital sodium. The chest of the anesthetized animal was opened and a catheter was inserted from the left ventricle to the aorta. The animal was first rapidly perfused with physiologic saline, then followed with perfusion of 4% paraformaldehyde in 0.1 M PBS (pH 7.4) to fix the tissue of the animal. The T8-L1 spinal segment was harvested, placed in a fresh fixation solution and fixed for 4 hours, and then placed in a 30% sucrose solution at 4° C. overnight. The T10, T11, T12, T13 and L1 spinal segments were continuously frozen-sectioned at cross sections to slices of 30 μm thickness.
Experiment 3 Fluorescent ImmunohistochemistryThe cross-section slices of spinal cords were first rinsed three times with 0.01 M PBS and each time the rinsing lasted for about 5 minutes. The spinal slices were then soaked in 2 N HCl for 30 minutes at 37° C., followed-by 0.1 M sodium borate buffer (pH 8.3) for 15 minutes and 20.2% Triton X-100 solution at 37° C. for 30 minutes. The spinal slices were then incubated in normal sheep serum at 37° C. for 20 minutes. Mouse antiBrdU primary antibody was added dropwise and the spinal slices were further incubated at 37° C. for 2 hours. The spinal slices were rinsed with 0.01 M PBS three times and each time the rinsing lasted for 5 minutes. The biotinylated secondary antibody was then added dropwise and the spinal slices were incubating at 37° C. for 30 minutes. The spinal slices were rinsed with 0.01 M PBS three times and each time the rinsing lasted for 5 minutes. The SABC-cys3 fluorescent complex solution was added dropwise and the spinal slices were incubating at 37° C. for 30 minutes. The spinal slices were rinsed with 0.01 M PBS three times while each time the rinsing lasted for 5 minutes. For negative control, the primary antibody was omitted and replaced with PBS. The rest of the procedures were the same as shown above.
After completing the BrdU staining, the spinal slices of each rat were divided into 4, each separately treated with the fluorescent immunohistochemistry stains for nestin (neural stem cell marker), NF (neuron marker), oligodendrocytin (oligodendrocyte marker) and GFAP (astroglial cell marker), respectively.
Under fluorescent microscope at 10×20 magnification, the numbers of BrdU-positive cells in the ependyma of the central canal of the cross-sectioned spinal slices were counted. For each rat, ten spinal slices were counted to determine the average number of BrdU-positive cells in the ependyma of the central canal per spinal slice. The calculated results of each group were expressed as the average numbers of BrdU-positive cells in the ependyma of the central canal±standard deviation. The t-test on the average numbers of two samples was performed.
Results of Examples 1-3:
1. Normal Ependyma in the Spinal Cord Central Canal
In the spinal cross sections of the neck and thoracic regions of the normal control group, the ependymal cells of the central canal appeared as a round shape, but as an oval shape in the lumbar region. For the purpose of locating the proliferation of the ependymal cells, mainly the BrdU-positive cells in the ependyma were counted. This was because most of the BrdU-positive cells were located at the ependyma (
2. Ependymal Cell Proliferation and Nestin Expression in the Spinal Cord Central Canal after Spinal Hemisection
At 1 day after the spinal injury, the ependymal cell layer of the central canal was thickened. The BrdU-positive cells markedly increased after spinal hemisection (
t test: pairwise comparison among the 3 groups, P < 0.05
At 1 day after the spinal hemisection, nestin-positive cells in the ependyma of the central canal increased. There were also nestin-positive cells in the spinal white matter and gray matter (
At 2 weeks and 4 weeks after the spinal injury, the ependymal cell layer of the spinal central canal of the injured/treatment group was thickened, having more BrdU-positive cells than those of the injured/control group. From the results of the BrdU and nestin, BrdU and NF, BrdU and oligodendrocytin, and BrdU and GFAP dual immunohistochemical stains, it was found that there were no cells positive with dual stains in the ependyma of the central canal. However, in the spinal white matter of the injured/treatment group there were a small number of cells positive with dual stains of BrdU and nestin (
Discussion:
The above study confirmed that at 1 day after the spinal hemisection, the ependymal cells of the central canal proliferated, but the number of proliferated cells declined as the time progressed. The closer to the spinal injured site, the more the proliferation of the ependymal cells was. Proliferated cells not only existed in the ependymal cells of the central canal, but also in the spinal white matter and gray matter. Meanwhile, some proliferated cells expressed nestin, indicating these cells had differentiated into neural stem cells. These findings agreed with the report of Yamamoto et al., Exp. Neurol., 2001; 172(1):115-127, that neural stem cells not only existed in the ependyma of the spinal central canal, but also existed in other parts of the spinal cord. In addition, the numbers of proliferated ependymal cells of the central canal in rats of the injured/treatment group were higher than those of the injured/control group, indicating GASP have the effects of promoting cell proliferation of the ependymal cells of the central canal in the injured spinal cord.
The results of the present study confirmed that after the spinal injury, a small number of proliferated ependymal cells could differentiate into neural stem cells, neuron-like cells, oligodendrocyte-like cells and astroglial-like cells which possibly involved in the repair of the injured spinal cord.
CONCLUSIONGASP is effective in promoting the proliferation and/or differentiation of neural stem cells in rats having spinal cord injury.
While the invention has been described by way of examples and in term of the preferred embodiments, it is to be understood that the invention is not limited to the disclosed embodiments. On the contrary, it is intended to cover various modifications as would be apparent to those skilled in the art. Therefore, the scope of the appended claims should be accorded the broadest interpretation so as to encompass all such modifications.
Claims
1. A method for promoting proliferation of neural stem cells in a mammal comprising:
- administering an effective amount of sporoderm-broken germination activated Ganoderma spores (GASP) to a mammal.
2. The method according to claim 1, wherein said promotion of proliferation of neural stem cells occur after a spinal cord injury.
3. The method according to claim 1, wherein said GASP are orally administered to said mammal.
4. The method according to claim 1, wherein said mammal is human.
5. The method according to claim 1, wherein said spinal cord injury is caused by compression or severance of the spinal cord.
6. The method according to claim 1, wherein said spinal cord injury is caused by a trauma.
7. The method according to claim 1, wherein said spinal cord injury is caused by a disease.
8. A method for promoting differentiation of neural stem cells in a mammal comprising:
- administering an effective amount of sporoderm-broken germination activated Ganoderma spores (GASP) to a mammal.
9. The method according to claim 8, wherein said neural stem cells having spinal cord injury.
10. The method according to claim 8, wherein said promotion of differentiation of said neural stem cells occur after a spinal cord injury.
11. The method according to claim 8, wherein said neural stem cells are differentiated into neurons.
12. The method according to claim 8, wherein said neural stem cells are differentiated into astrocytes.
13. The method according to claim 8, wherein said neural stem cells are differentiated into oligodendrocytes.
14. The method according to claim 8, wherein said GASP are orally administered to said mammal.
15. The method according to claim 8, wherein said mammal is human.
16. The method according to claim 8, wherein said spinal cord injury is caused by compression or severance of the spinal cord.
17. The method according to claim 8, wherein said spinal cord injury is caused by a trauma.
18. The method according to claim 8, wherein said spinal cord injury is caused by a disease.
19. A method for promoting proliferating and differentiating neural stem cells in a mammal comprising:
- administering an effective amount of sporoderm-broken germination activated Ganoderma spores (GASP) to a mammal.
20. The method according to claim 19, wherein said proliferating and differentiating of said neural stem cells occur after a spinal cord injury.
Type: Application
Filed: Jan 8, 2004
Publication Date: Feb 3, 2005
Inventors: Chee-Keung Chung (Kowloon), Siu-Kan Tong (Kowloon)
Application Number: 10/752,685
