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1.
Front Mol Neurosci ; 14: 618360, 2021.
Article in English | MEDLINE | ID: mdl-34040503

ABSTRACT

Mucopolysaccharidosis type I (MPS I) is an inherited metabolic disorder caused by deficiency of the lysosomal enzyme alpha-L-iduronidase (IDUA). The two current treatments [hematopoietic stem cell transplantation (HSCT) and enzyme replacement therapy (ERT)], are insufficiently effective in addressing neurologic disease, in part due to the inability of lysosomal enzyme to cross the blood brain barrier. With a goal to more effectively treat neurologic disease, we have investigated the effectiveness of AAV-mediated IDUA gene delivery to the brain using several different routes of administration. Animals were treated by either direct intracerebroventricular (ICV) injection, by intrathecal (IT) infusion into the cerebrospinal fluid, or by intranasal (IN) instillation of AAV9-IDUA vector. AAV9-IDUA was administered to IDUA-deficient mice that were either immunosuppressed with cyclophosphamide (CP), or immunotolerized at birth by weekly injections of human iduronidase. In animals treated by ICV or IT administration, levels of IDUA enzyme ranged from 3- to 1000-fold that of wild type levels in all parts of the microdissected brain. In animals administered vector intranasally, enzyme levels were 100-fold that of wild type in the olfactory bulb, but enzyme expression was close to wild type levels in other parts of the brain. Glycosaminoglycan levels were reduced to normal in ICV and IT treated mice, and in IN treated mice they were normalized in the olfactory bulb, or reduced in other parts of the brain. Immunohistochemical analysis showed extensive IDUA expression in all parts of the brain of ICV treated mice, while IT treated animals showed transduction that was primarily restricted to the hind brain with some sporadic labeling seen in the mid- and fore brain. At 6 months of age, animals were tested for spatial navigation, memory, and neurocognitive function in the Barnes maze; all treated animals were indistinguishable from normal heterozygous control animals, while untreated IDUA deficient animals exhibited significant learning and spatial navigation deficits. We conclude that IT and IN routes are acceptable and alternate routes of administration, respectively, of AAV vector delivery to the brain with effective IDUA expression, while all three routes of administration prevent the emergence of neurocognitive deficiency in a mouse MPS I model.

2.
Cell Transplant ; 26(8): 1472-1482, 2017 08.
Article in English | MEDLINE | ID: mdl-28901182

ABSTRACT

Spinal cord injury (SCI) is a widely disabling condition, constraining those affected by it to wheelchairs and requiring intense daily care and assistance. Cell replacement therapies, targeting regeneration of cells in the injured cord, are currently gaining momentum in the field of SCI research. Previous studies indicate that mesenchymal stem cells (MSCs) can reduce functional deficits through immunomodulation and production of trophic factors in a variety of neurological disorders. The present study assessed the efficacy of transplanted bone marrow-derived MSCs at different concentrations and locations for promoting functional recovery following SCI. Although effects were modest, MSCs facilitated an increase in the base of support, as measured by increased distance between the plantar surface of the hind paws, following incomplete contusive SCI, and reduced the density of astroglial scarring. Varying the concentrations or locations of transplanted cells did not provide additional benefits on these measures. These findings indicate that MSC transplants are safe at relatively high concentrations and confer therapeutic benefits that, when used as an adjunctive treatment, could significantly enhance functional recovery following SCI.


Subject(s)
Bone Marrow/metabolism , Cell- and Tissue-Based Therapy/methods , Mesenchymal Stem Cells/metabolism , Spinal Cord Injuries/therapy , Animals , Disease Models, Animal , Male , Mesenchymal Stem Cells/cytology , Rats , Rats, Sprague-Dawley
3.
Restor Neurol Neurosci ; 35(4): 395-411, 2017.
Article in English | MEDLINE | ID: mdl-28598857

ABSTRACT

PURPOSE: Utilizing genetic overexpression of trophic molecules in cell populations has been a promising strategy to develop cell replacement therapies for spinal cord injury (SCI). Over-expressing the chemokine, stromal derived factor-1 (SDF-1α), which has chemotactic effects on many cells of the nervous system, offers a promising strategy to promote axonal regrowth following SCI. The purpose of this study was to explore the effects of human SDF-1α, when overexpressed by mesenchymal stem cells (MSCs), on axonal growth and motor behavior in a contusive rat model of SCI. METHODS: Using a transwell migration assay, the paracrine effects of MSCs, which were engineered to secrete human SDF-1α (SDF-1-MSCs), were assessed on cultured neural stem cells (NSCs). For in vivo analyses, the SDF-1-MSCs, unaltered MSCs, or Hanks Buffered Saline Solution (vehicle) were injected into the lesion epicenter of rats at 9-days post-SCI. Behavior was analyzed for 7-weeks post-injury, using the Basso, Beattie, and Bresnahan (BBB) scale of locomotor functions. Immunohistochemistry was performed to evaluate major histopathological outcomes, including gliosis, inflammation, white matter sparing, and cavitation. New axonal outgrowth was characterized using immunohistochemistry against the neuron specific growth-associated protein-43 (GAP-43). RESULTS: The results of these experiments demonstrate that the overexpression of SDF-1α by MSCs can enhance the migration of NSCs in vitro. Although only modest functional improvements were observed following transplantation of SDF-1-MSCs, a significant reduction in cavitation surrounding the lesion, and an increased density of GAP-43-positive axons inside the SCI lesion/graft site were found. CONCLUSION: The results from these experiments support the potential role for utilizing SDF-1α as a treatment for enhancing growth and regeneration of axons after traumatic SCI.


Subject(s)
Axons/metabolism , Chemokine CXCL12/metabolism , Mesenchymal Stem Cell Transplantation , Mesenchymal Stem Cells/metabolism , Spinal Cord Injuries/therapy , Spinal Cord Regeneration , Animals , Axons/pathology , Cell Culture Techniques , Cell Engineering , Cell Movement , Chemokine CXCL12/genetics , Disease Models, Animal , Female , GAP-43 Protein/metabolism , Humans , Male , Mesenchymal Stem Cell Transplantation/methods , Mesenchymal Stem Cells/pathology , Motor Activity/physiology , Neural Stem Cells/metabolism , Neural Stem Cells/pathology , Proof of Concept Study , Rats, Sprague-Dawley , Recovery of Function/physiology , Spinal Cord/metabolism , Spinal Cord/pathology , Spinal Cord Injuries/metabolism , Spinal Cord Injuries/pathology
4.
Hum Gene Ther ; 28(8): 626-638, 2017 08.
Article in English | MEDLINE | ID: mdl-28478695

ABSTRACT

Mucopolysaccharidosis type II (MPS II; Hunter syndrome) is a rare X-linked recessive lysosomal disorder caused by defective iduronate-2-sulfatase (IDS), resulting in accumulation of heparan sulfate and dermatan sulfate glycosaminoglycans (GAGs). Enzyme replacement is the only Food and Drug Administration-approved therapy available for MPS II, but it is expensive and does not improve neurologic outcomes in MPS II patients. This study evaluated the effectiveness of adeno-associated virus (AAV) vector encoding human IDS delivered intracerebroventricularly in a murine model of MPS II. Supraphysiological levels of IDS were observed in the circulation (160-fold higher than wild type) for at least 28 weeks post injection and in most tested peripheral organs (up to 270-fold) at 10 months post injection. In contrast, only low levels of IDS were observed (7-40% of wild type) in all areas of the brain. Sustained IDS expression had a profound effect on normalization of GAG in all tested tissues and on prevention of hepatomegaly. Additionally, sustained IDS expression in the central nervous system (CNS) had a prominent effect in preventing neurocognitive deficit in MPS II mice treated at 2 months of age. This study demonstrates that CNS-directed, AAV9 mediated gene transfer is a potentially effective treatment for Hunter syndrome, as well as other monogenic disorders with neurologic involvement.


Subject(s)
Dependovirus/genetics , Gene Transfer Techniques , Genetic Therapy , Genetic Vectors/genetics , Iduronate Sulfatase/genetics , Mucopolysaccharidosis II/genetics , Mucopolysaccharidosis II/psychology , Animals , Central Nervous System/metabolism , Cognition , Disease Models, Animal , Enzyme Activation , Female , Gene Expression , Gene Order , Genetic Vectors/administration & dosage , Glycosaminoglycans/metabolism , Humans , Iduronate Sulfatase/blood , Iduronate Sulfatase/metabolism , Male , Mice , Mucopolysaccharidosis II/blood , Mucopolysaccharidosis II/therapy , Neuropsychological Tests , Pilot Projects , Time Factors , Tissue Distribution , Transduction, Genetic
5.
Cell Transplant ; 25(8): 1473-88, 2016.
Article in English | MEDLINE | ID: mdl-26996530

ABSTRACT

Despite the high prevalence and devastating outcome, there remain a few options for treatment of ischemic stroke. Currently available treatments are limited by a short time window for treatment and marginal efficacy when used. We have tested a human umbilical cord blood-derived stem cell line that has been shown to result in a significant reduction in stroke infarct volume as well as improved functional recovery following stroke in the rat. In the present study we address the mechanism of action and compared the therapeutic efficacy of high- versus low-passage nonhematopoietic umbilical cord blood stem cells (nh-UCBSCs). Using the middle cerebral arterial occlusion (MCAo) model of stroke in Sprague-Dawley rats, we administered nh-UCBSC by intravenous (IV) injection 2 days following stroke induction. These human cells were injected into rats without any immune suppression, and no adverse reactions were detected. Both behavioral and histological analyses have shown that the administration of these cells reduces the infarct volume by 50% as well as improves the functional outcome of these rats following stroke for both high- and low-passaged nh-UCBSCs. Flow cytometry analysis of immune cells present in the brains of normal rats, rats with ischemic brain injury, and ischemic animals with nh-UCBSC treatment confirmed infiltration of macrophages and T cells consequent to ischemia and reduction to normal levels with nh-UCBSC treatment. Flow cytometry also revealed a restoration of normal levels of microglia in the brain following treatment. These data suggest that nh-UCBSCs may act by inhibiting immune cell migration into the brain from the periphery and possibly by inhibition of immune cell activation within the brain. nh-UCBSCs exhibit great potential for treatment of stroke, including the fact that they are associated with an increased therapeutic time window, no known ill-effects, and that they can be expanded to high numbers for, and stored for, treatment.


Subject(s)
Brain Ischemia/therapy , Fetal Blood/cytology , Stem Cells/cytology , Stroke/therapy , Animals , Cells, Cultured , Disease Models, Animal , Female , Flow Cytometry , Humans , Infarction, Middle Cerebral Artery/therapy , Macrophages/cytology , Microglia/cytology , Rats , Rats, Sprague-Dawley , Recovery of Function
6.
Neurosurgery ; 74(1): 99-111, 2014 Jan.
Article in English | MEDLINE | ID: mdl-24077583

ABSTRACT

BACKGROUND: Hurler disease (mucopolysaccharidosis type I [MPS-I]) is an inherited metabolic disorder characterized by deficiency of the lysosomal enzyme α-L-iduronidase (IDUA). Currently, the only therapies for MPS-I, enzyme replacement and hematopoietic stem cell transplantation, are generally ineffective for central nervous system manifestations. OBJECTIVE: To test whether brain-targeted gene therapy with recombinant adeno-associated virus (rAAV5)-IDUA vectors in an MPS-I transgenic mouse model would reverse the pathological hallmarks. METHODS: Gene therapy approaches were compared using intraventricular or endovascular delivery with a marker (rAAV5-green fluorescent protein) or therapeutic (rAAV5-IDUA) vector. To improve the efficiency of brain delivery, we tested different applications of hyperosmolar mannitol to disrupt the blood-brain barrier or ependymal-brain interface. RESULTS: Intraventricular delivery of 1 × 10 viral particles of rAAV5-IDUA with systemic 5 g/kg mannitol co-administration resulted in IDUA expression throughout the brain, with global enzyme activity >200% of the baseline level in age-matched, wild-type mice. Endovascular delivery of 1 × 10 viral particles of rAAV5-IDUA to the carotid artery with 29.1% mannitol blood-brain barrier disruption resulted in mainly ipsilateral brain IDUA expression and ipsilateral brain enzyme activity 42% of that in wild-type mice. Quantitative assays for glycosaminoglycans showed a significant decrease in both hemispheres after intraventricular delivery and in the ipsilateral hemisphere after endovascular delivery compared with untreated MPS-I mice. Immunohistochemistry for ganglioside GM3, another disease marker, showed reversal of neuronal inclusions in areas with IDUA co-expression in both delivery methods. CONCLUSION: Physiologically relevant biochemical correction is possible with neurosurgical or endovascular gene therapy approaches for MPS-I. Intraventricular or endovascular delivery of rAAV5-IDUA was effective in reversing brain pathology, but in the latter method, effects were limited to the ipsilateral hemisphere.


Subject(s)
Genetic Therapy/methods , Iduronidase/administration & dosage , Mucopolysaccharidosis I/pathology , Animals , Dependovirus/genetics , Disease Models, Animal , Genetic Vectors , Humans , Iduronidase/genetics , Injections, Intra-Arterial , Injections, Intraventricular , Mice , Mice, Transgenic
7.
Cell Transplant ; 21(7): 1577-93, 2012.
Article in English | MEDLINE | ID: mdl-22472595

ABSTRACT

Mucopolysaccharidosis type I (MPS-I; Hurler syndrome) is an inborn error of metabolism caused by lack of the functional lysosomal glycosaminoglycan (GAG)-degrading enzyme α-L-iduronidase (IDUA). Without treatment, the resulting GAG accumulation causes multisystem dysfunction and death within the first decade. Current treatments include allogeneic hematopoietic stem cell transplantation (HSCT) and enzyme replacement therapy. HSCT ameliorates clinical features and extends life but is not available to all patients, and inadequately corrects the most devastating features of the disease including mental retardation and skeletal deformities. Recent developments suggest that stem cells can be used to deliver needed enzymes to the central nervous system. To test this concept, we transplanted bone marrow-derived normal adult human MultiStem® cells into the cerebral lateral ventricles of immunodeficient MPS-I neonatal mice. Transplanted cells and human-specific DNA were detected in the hippocampal formation, striatum, and other areas of the central nervous system. Brain tissue assays revealed significant long-term decrease in GAG levels in the hippocampus and striatum. Sensorimotor testing 6 months after transplantation demonstrated significantly improved rotarod performance of transplanted mice in comparison to nontransplanted and sham-transplanted control animals. These results suggest that a single injection of MultiStem cells into the cerebral ventricles of neonatal MPS-I mice induces sustained reduction in GAG accumulation within the brain, and modest long-term improvement in sensorimotor function.


Subject(s)
Bone Marrow Cells/cytology , Mucopolysaccharidosis I/therapy , Multipotent Stem Cells/transplantation , Animals , Animals, Newborn , Brain/metabolism , Corpus Striatum/metabolism , Disease Models, Animal , Feedback, Sensory/physiology , Glycosaminoglycans/metabolism , Hippocampus/metabolism , Humans , Infusions, Intraventricular , Mice , Mice, Inbred NOD , Mice, SCID , Motor Activity/physiology , Mucopolysaccharidosis I/metabolism , Mucopolysaccharidosis I/pathology , Multipotent Stem Cells/cytology , Transplantation, Heterologous
8.
Mol Genet Metab ; 106(1): 131-4, 2012 May.
Article in English | MEDLINE | ID: mdl-22420937

ABSTRACT

Here we provide the first evidence that therapeutic levels of a lysosomal enzyme can bypass the blood-brain barrier following intranasal administration. α-L-iduronidase (IDUA) activity was detected throughout the brains of IDUA-deficient mice following a single intranasal treatment with concentrated Aldurazyme® (laronidase) and was also detected after intranasal treatment with an adeno-associated virus (AAV) vector expressing human IDUA. These results suggest that intranasal routes of delivery may be efficacious in the treatment of lysosomal storage disorders.


Subject(s)
Blood-Brain Barrier , Central Nervous System/drug effects , Iduronidase/administration & dosage , Iduronidase/genetics , Mucopolysaccharidosis I/drug therapy , Administration, Intranasal , Animals , Brain/drug effects , Dependovirus/genetics , Disease Models, Animal , Drug Delivery Systems , Genetic Vectors/administration & dosage , Humans , Lysosomes/enzymology , Mice , Mice, Transgenic , Recombinant Proteins/administration & dosage
9.
Neurobiol Dis ; 43(1): 123-33, 2011 Jul.
Article in English | MEDLINE | ID: mdl-21397026

ABSTRACT

The mucopolysaccharidoses (MPSs) are a group of 11 storage diseases caused by disruptions in glycosaminoglycan (GAG) catabolism, leading to their accumulation in lysosomes. Resultant multisystemic disease is manifested by growth delay, hepatosplenomegaly, skeletal dysplasias, cardiopulmonary obstruction, and, in severe MPS I, II, III, and VII, progressive neurocognitive decline. Some MPSs are treated by allogeneic hematopoietic stem cell transplantation (HSCT) and/or recombinant enzyme replacement therapy (ERT), but effectiveness is limited by central nervous system (CNS) access across the blood-brain barrier. To provide a high level of gene product to the CNS, we tested neonatal intracerebroventricular (ICV) infusion of an adeno-associated virus (AAV) serotype 8 vector transducing the human α-L-iduronidase gene in MPS I mice. Supranormal levels of iduronidase activity in the brain (including 40× normal levels in the hippocampus) were associated with transduction of neurons in motor and limbic areas identifiable by immunofluorescence staining. The treatment prevented accumulation of GAG and GM3 ganglioside storage materials and emergence of neurocognitive dysfunction in a modified Morris water maze test. The results suggest the potential of improved outcome for MPSs and other neurological diseases when a high level of gene expression can be achieved by direct, early administration of vector to the CNS.


Subject(s)
Genetic Therapy/methods , Iduronidase/administration & dosage , Iduronidase/genetics , Mucopolysaccharidosis I/prevention & control , Mucopolysaccharidosis I/therapy , Transduction, Genetic/methods , Adenoviridae/genetics , Animals , Animals, Newborn , Disease Models, Animal , Female , Humans , Iduronidase/biosynthesis , Infusions, Intraventricular , Male , Mice , Mice, Mutant Strains , Mucopolysaccharidosis I/genetics , Up-Regulation/genetics
10.
Proc Natl Acad Sci U S A ; 108(1): 260-5, 2011 Jan 04.
Article in English | MEDLINE | ID: mdl-21173221

ABSTRACT

Trinucleotide expansions cause disease by both protein- and RNA-mediated mechanisms. Unexpectedly, we discovered that CAG expansion constructs express homopolymeric polyglutamine, polyalanine, and polyserine proteins in the absence of an ATG start codon. This repeat-associated non-ATG translation (RAN translation) occurs across long, hairpin-forming repeats in transfected cells or when expansion constructs are integrated into the genome in lentiviral-transduced cells and brains. Additionally, we show that RAN translation across human spinocerebellar ataxia type 8 (SCA8) and myotonic dystrophy type 1 (DM1) CAG expansion transcripts results in the accumulation of SCA8 polyalanine and DM1 polyglutamine expansion proteins in previously established SCA8 and DM1 mouse models and human tissue. These results have implications for understanding fundamental mechanisms of gene expression. Moreover, these toxic, unexpected, homopolymeric proteins now should be considered in pathogenic models of microsatellite disorders.


Subject(s)
Protein Biosynthesis/genetics , Spinocerebellar Ataxias/genetics , Trinucleotide Repeat Expansion/genetics , Amino Acid Sequence , Blotting, Northern , Cell Line , Cloning, Molecular , Codon, Initiator/genetics , DNA Primers/genetics , Fluorescent Antibody Technique , Genetic Vectors , Humans , Immunoblotting , Immunohistochemistry , Immunoprecipitation , Lentivirus , Mass Spectrometry , Molecular Sequence Data , Mutagenesis , Myotonic Dystrophy/genetics , Peptides/genetics , Peptides/metabolism , Protein Biosynthesis/physiology , Reverse Transcriptase Polymerase Chain Reaction
11.
Brain Res Bull ; 74(6): 429-38, 2007 Nov 01.
Article in English | MEDLINE | ID: mdl-17920451

ABSTRACT

Mucopolysaccharidosis type I (MPS-I or Hurler syndrome) is an inherited deficiency of the lysosomal glycosaminoglycan (GAG)-degrading enzyme alpha-l-iduronidase (IDUA) in which GAG accumulation causes progressive multi-system dysfunction and death. Early allogeneic hematopoietic stem cell transplantation (HSCT) ameliorates clinical features and extends life but is not available to all patients, and inadequately corrects its most devastating features including mental retardation and skeletal deformities. To test novel therapies, we characterized an immunodeficient MPS-I mouse model less likely to develop immune reactions to transplanted human or gene-corrected cells or secreted IDUA. In the liver, spleen, heart, lung, kidney and brain of NOD/SCID/MPS-I mice IDUA was undetectable, and reduced to half in heterozygotes. MPS-I mice developed marked GAG accumulation (3-38-fold) in these organs. Neuropathological examination showed GM(3) ganglioside accumulation in the striatum, cerebral peduncles, cerebellum and ventral brainstem of MPS-I mice. Urinary GAG excretion (6.5-fold higher in MPS-I mice) provided a non-invasive and reliable method suitable for serially following the biochemical efficacy of therapeutic interventions. We identified and validated using rigorous biostatistical methods, a highly reproducible method for evaluating sensorimotor function and motor skills development. This Rotarod test revealed marked abnormalities in sensorimotor integration involving the cerebellum, striatum, proprioceptive pathways, motor cortex, and in acquisition of motor coordination. NOD/SCID/MPS-I mice exhibit many of the clinical, skeletal, pathological and behavioral abnormalities of human MPS-I, and provide an extremely suitable animal model for assessing the systemic and neurological effects of human stem cell transplantation and gene therapeutic approaches, using the above techniques to measure efficacy.


Subject(s)
Disease Models, Animal , Genetic Therapy , Mucopolysaccharidoses/pathology , Mucopolysaccharidoses/physiopathology , Mucopolysaccharidoses/therapy , Stem Cell Transplantation , Animals , Glycosaminoglycans/metabolism , Humans , Immunohistochemistry , Mice , Mice, Mutant Strains
12.
Ann N Y Acad Sci ; 1049: 84-96, 2005 May.
Article in English | MEDLINE | ID: mdl-15965109

ABSTRACT

Umbilical cord blood is a rich source of hematopoietic stem cells. It is routinely used for transplantation to repopulate cells of the immune system. Recent studies, however, have demonstrated that intravenous infusions of umbilical cord blood can ameliorate neurologic deficits associated with ischemic brain injury in rodents. Moreover, the infused cells penetrate into the parenchyma of the brain and adopt phenotypic characteristics typical of neural cells. In the present study we tested the hypothesis that the administration of umbilical cord blood can also diminish neurologic deficits caused by intracerebral hemorrhage (ICH). Intracerebral hemorrhage is a major cause of morbidity and mortality, and at the present time there are no adequate therapies that can minimize the consequences of this cerebrovascular event. ICH was induced in rats by intrastriatal injections of collagenase to cause bleeding in the striatum. Twenty-four hours after the induction of ICH rats received intravenous saphenous vein infusions of human umbilical cord blood (2.4 x 10(6) to 3.2 to 10(6) cells). Animals were evaluated using a battery of tests at day 1 after ICH, but before the administration of umbilical cord blood, and at days 7, and 14 after ICH (days 6 and 13, respectively, after cord blood administration). These tests included a neurological severity test, a stepping test, and an elevated body-swing test. Animals with umbilical cord blood infusions exhibited significant improvements in (1) the neurologic severity test at 6 and 13 days after cord blood infusion in comparison to saline-treated animals (P < 0.05); (2) the stepping test at day 6 (P < 0.05); and (3) the elevated body-swing test at day 13 (P< 0.05). These results demonstrate that the administration of human umbilical cord blood cells can ameliorate neurologic deficits associated with intracerebral hemorrhage.


Subject(s)
Brain Injuries/therapy , Cerebral Hemorrhage/therapy , Cord Blood Stem Cell Transplantation , Fetal Blood/cytology , Hematopoietic Stem Cells/physiology , Animals , Behavior, Animal , Brain Injuries/pathology , Cerebral Hemorrhage/pathology , Disease Models, Animal , Hematopoietic Stem Cells/cytology , Humans , Rats , Rats, Sprague-Dawley , Transplantation, Heterologous
13.
Stem Cells Dev ; 14(6): 722-33, 2005 Dec.
Article in English | MEDLINE | ID: mdl-16433627

ABSTRACT

Umbilical cord blood (UCB) is a rich source of hematopoetic stem cells (HSCs). We have isolated a novel cell line population of stem cells from human UCB that exhibit properties of self-renewal, but do not have cell-surface markers that are typically found on HSCs. Analysis of transcripts revealed that these cells express transcription factors Oct-4, Rex-1, and Sox-2 that are typically expressed by stem cells. We refer to these novel cells as nonhematopoietic umbilical cord blood stem cells (nh-UCBSCs). Previous studies have shown that the intravenous infusion of UCBCs can ameliorate neurological deficits arising from ischemic brain injury. The identity of the cells that mediate this restorative effect, however, has yet to be determined. We postulate that nh-UCBSCs may be a source of the UCB cells that can mediate these effects. To test this hypothesis, we intravenously injected one million human nh-UCBSCs into rats 48 h after transient unilateral middle cerebral artery occlusion. Animals in other experimental groups received either saline injections or injections of RN33b neural stem cells. Animals were tested for neurological function before the infusion of nh-UCBSCs and at various time periods afterwards using a battery of behavioral tests. In limb placement tests, animals treated with nh-UCBSCs exhibited mean scores that were significantly better than animals treated with RN33b neural stem cells or saline. Similarly, in stepping tests, nh-UCBSC-treated animals again exhibited significantly better performance than the other experimental groups of animals. Analysis of infarct volume revealed that ischemic animals treated with nh-UCBSCs exhibited a 50% reduction in lesion volume in comparison to saline-treated controls. Histological analysis of brain tissue further revealed the presence of cells that stained for human nuclei. Some human nuclei-positive cells were also co-labeled for NeuN, indicating that the transplanted cells expressed markers of a neuronal phenotype. Cells expressing the human nuclei marker within the brain, however, were rather scant, suggesting that the restorative effects of nh-UCBSCs may be mediated by mechanisms other than cell replacement. To test this hypothesis, nh-UCBSCs were directly transplanted into the brain parenchyma after ischemic brain injury. Sprouting of nerve fibers from the nondamaged hemisphere into the ischemically damaged side of the brain was assessed by anterograde tracing using biotinylated dextran amine (BDA). Animals with nh-UCBSC transplants exhibited significantly greater densities of BDA-positive cells in the damaged side of the brain compared to animals with intraparenchymal saline injections. These results suggest that restorative effects observed with nh-UCBSC treatment following ischemic brain injury may be mediated by trophic actions that result in the reorganization of host nerve fiber connections within the injured brain.


Subject(s)
Brain Ischemia/therapy , Cord Blood Stem Cell Transplantation , Fetal Blood/cytology , Fetal Blood/metabolism , Animals , Brain Infarction/pathology , Cell Line , Humans , Karyotyping , Nerve Fibers/physiology , Rats
14.
Proc Natl Acad Sci U S A ; 100(10): 6087-92, 2003 May 13.
Article in English | MEDLINE | ID: mdl-12721362

ABSTRACT

Tauroursodeoxycholic acid (TUDCA), an endogenous bile acid, modulates cell death by interrupting classic pathways of apoptosis. Intracerebral hemorrhage (ICH) is a devastating acute neurological disorder, without effective treatment, in which a significant loss of neuronal cells is thought to occur by apoptosis. In this study, we evaluated whether TUDCA can reduce brain injury and improve neurological function after ICH in rats. Administration of TUDCA before or up to 6 h after stereotaxic collagenase injection into the striatum reduced lesion volumes at 2 days by as much as 50%. Apoptosis was approximately 50% decreased in the area immediately surrounding the hematoma and was associated with a similar inhibition of caspase activity. These changes were also associated with improved neurobehavioral deficits as assessed by rotational asymmetry, limb placement, and stepping ability. Furthermore, TUDCA treatment modulated expression of certain Bcl-2 family members, as well as NF-kappaB activity. In addition to its protective action at the mitochondrial membrane, TUDCA also activated the Akt-1protein kinase Balpha survival pathway and induced Bad phosphorylation at Ser-136. In conclusion, reduction of brain injury underlies the wide-range neuroprotective effects of TUDCA after ICH. Thus, given its clinical safety, TUDCA may provide a potentially useful treatment in patients with hemorrhagic stroke and perhaps other acute brain injuries associated with cell death by apoptosis.


Subject(s)
Apoptosis/drug effects , Brain/pathology , Cerebral Hemorrhage/prevention & control , Taurochenodeoxycholic Acid/therapeutic use , Animals , Bile/metabolism , Brain/drug effects , Brain/metabolism , Caspases/metabolism , Cerebral Hemorrhage/chemically induced , Cerebral Hemorrhage/pathology , Cholagogues and Choleretics/therapeutic use , Collagenases , DNA Primers , Disease Models, Animal , Female , In Situ Nick-End Labeling , Proto-Oncogene Proteins c-bcl-2/genetics , Rats , Rats, Sprague-Dawley , Reverse Transcriptase Polymerase Chain Reaction , Substrate Specificity
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