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Frances M. Platt - One of the best experts on this subject based on the ideXlab platform.
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NSAIDs Increase Survival in the Sandhoff Disease Mouse: Synergy with N-butyldeoxynojirimycin
2016Co-Authors: Frances M. PlattAbstract:The GM2 gangliosidoses are caused by incomplete catabolism of GM2 ganglioside in the lysosome, leading to progressive storage and a neurodegenerative clinical course. An inflammatory response (microglial activation, macrophage infiltra-tion, oxidative damage) has been found to be a consequence of GM2 storage in the brain, although it remains unclear whether this contributes to pathogenesis or Disease progression. In this study, we treated Sandhoff Disease mice with nonsteroidal antiinflammatory drugs (indomethacin, aspirin, and ibuprofen) and antioxidants (L-ascorbic acid and -tochopherol acetate). The treated mice lived significantly longer than untreated littermates (12–23%, p < 0.0001) and showed a slower rate of Disease progression (p < 0.001). When aspirin treatment was combined with substrate reduction therapy, synergy resulted (11%, p < 0.05) with a maximum improvement of 73 % in survival (p < 0.00001). This study demonstrates that inflammation contributes to Disease progression and identifies antiinflammatory and antioxidant ther-apies as a potential adjunctive approach to slow the clinical course of this and related disorders. Ann Neurol 2004;56:642–649 The glycosphingolipid (GSL) lysosomal storage dis-eases arise due to inherited defects in the genes en-coding the enzymes, or their cofactors, responsible for the catabolism of GSLs in the lysosome.1,2 Withi
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deprivation therapy and
2016Co-Authors: Mylvaganam Jeyakumar, Terry D. Butters, Richard L Proia, Raymond A. Dwek, Francine Norflus, Cynthia J. Tifft, Mario Cortina-borja, Hugh V. Perry, Frances M. PlattAbstract:survival in Sandhoff Disease mice receiving a combination of substrat
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macroautophagy is not directly involved in the metabolism of amyloid precursor protein
Journal of Biological Chemistry, 2010Co-Authors: Barry Boland, David Smith, Declan Mooney, Sonia S Jung, Dominic M Walsh, Frances M. PlattAbstract:Alterations in the metabolism of amyloid precursor protein (APP) are believed to play a central role in Alzheimer Disease pathogenesis. Burgeoning data indicate that APP is proteolytically processed in endosomal-autophagic-lysosomal compartments. In this study, we used both in vivo and in vitro paradigms to determine whether alterations in macroautophagy affect APP metabolism. Three mouse models of glycosphingolipid storage Diseases, namely Niemann-Pick type C1, GM1 gangliosidosis, and Sandhoff Disease, had mTOR-independent increases in the autophagic vacuole (AV)-associated protein, LC3-II, indicative of impaired lysosomal flux. APP C-terminal fragments (APP-CTFs) were also increased in brains of the three mouse models; however, discrepancies between LC3-II and APP-CTFs were seen between primary (GM1 gangliosidosis and Sandhoff Disease) and secondary (Niemann-Pick type C1) lysosomal storage models. APP-CTFs were proportionately higher than LC3-II in cerebellar regions of GM1 gangliosidosis and Sandhoff Disease, although LC3-II increased before APP-CTFs in brains of NPC1 mice. Endogenous murine Aβ40 from RIPA-soluble extracts was increased in brains of all three mice. The in vivo relationship between AV and APP-CTF accumulation was also seen in cultured neurons treated with agents that impair primary (chloroquine and leupeptin + pepstatin) and secondary (U18666A and vinblastine) lysosomal flux. However, Aβ secretion was unaffected by agents that induced autophagy (rapamycin) or impaired AV clearance, and LC3-II-positive AVs predominantly co-localized with degradative LAMP-1-positive lysosomes. These data suggest that neuronal macroautophagy does not directly regulate APP metabolism but highlights the important anti-amyloidogenic role of lysosomal proteolysis in post-secretase APP-CTF catabolism.
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n butyldeoxygalactonojirimycin reduces brain ganglioside and gm2 content in neonatal Sandhoff Disease mice
Neurochemistry International, 2008Co-Authors: Rena C Baek, Frances M. Platt, J L Kasperzyk, Thomas N SeyfriedAbstract:Abstract Sandhoff Disease involves the CNS accumulation of ganglioside GM2 and asialo-GM2 (GA2) due to inherited defects in the β-subunit gene of β-hexosaminidase A and B ( Hexb gene). Accumulation of these glycosphingolipids (GSLs) produces progressive neurodegeneration, ultimately leading to death. Substrate reduction therapy (SRT) aims to decrease the rate of glycosphingolipid (GSL) biosynthesis to compensate for the impaired rate of catabolism. The imino sugar, N -butyldeoxygalactonojirimycin ( N B-DGJ) inhibits the first committed step in GSL biosynthesis. N B-DGJ treatment, administered from postnatal day 2 (p-2) to p-5 (600 mg/kg/day)), significantly reduced total brain ganglioside and GM2 content in the Sandhoff Disease ( Hexb −/− ) mice, but did not reduce the content of GA2. We also found that N B-DGJ treatment caused a slight, but significant elevation in brain sialidase activity. The drug had no adverse effects on viability, body weight, brain weight, or brain water content in the mice. No significant alterations in neutral lipids or acidic phospholipids were observed in the N B-DGJ-treated Hexb −/− mice. Our results show that N B-DGJ is effective in reducing total brain ganglioside and GM2 content at early neonatal ages.
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improved outcome of n butyldeoxygalactonojirimycin mediated substrate reduction therapy in a mouse model of Sandhoff Disease
Neurobiology of Disease, 2004Co-Authors: Ulrika Andersson, Mylvaganam Jeyakumar, Mario Cortina Borja, Terry D. Butters, Raymond A. Dwek, David Smith, Frances M. PlattAbstract:Abstract Sandhoff Disease is a severe neurodegenerative glycosphingolipid (GSL) lysosomal storage disorder, currently without treatment options. One therapeutic approach under investigation is substrate reduction therapy (SRT). By partially inhibiting GSL biosynthesis, the impaired rate of GSL catabolism is balanced by a slower rate of influx of GSLs into the lysosome. In a previous study, we reported the beneficial effects of treating Sandhoff Disease mice with the glucose analogue N-butyldeoxynojirimycin (NB-DNJ), a compound that inhibits the first step of GSL biosynthesis catalysed by the ceramide specific glucosyltransferase. NB-DNJ, however, exhibits adverse effects at high doses such as weight loss and GI tract distress (due to glucosidase inhibition). This might limit the therapeutic potential of NB-DNJ for treating Diseases affecting the CNS where high dose therapy may be required to achieve therapeutic levels of the drug in the brain. In the present study, a more selective compound, the galactose analogue N-butyldeoxygalactonojirimycin (NB-DGJ), was evaluated in the Sandhoff Disease mouse model. Treatment with NB-DGJ showed greater therapeutic efficacy than NB-DNJ with no detectable side effects. The ability to escalate the dose of NB-DGJ, leading to extended life expectancy and increased delay in symptom onset, demonstrates the greater therapeutic potential of NB-DGJ for the treatment of the human gangliosidoses.
Richard L Proia - One of the best experts on this subject based on the ideXlab platform.
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deprivation therapy and
2016Co-Authors: Mylvaganam Jeyakumar, Terry D. Butters, Richard L Proia, Raymond A. Dwek, Francine Norflus, Cynthia J. Tifft, Mario Cortina-borja, Hugh V. Perry, Frances M. PlattAbstract:survival in Sandhoff Disease mice receiving a combination of substrat
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sphingosine kinase 1 s1p receptor signaling axis controls glial proliferation in mice with Sandhoff Disease
Human Molecular Genetics, 2008Co-Authors: Kiyomi Mizugishi, Meryem Bektas, Roger Sandhoff, Richard L ProiaAbstract:Sphingosine-1-phosphate (S1P) is a lipid-signaling molecule produced by sphingosine kinase in response to a wide number of stimuli. By acting through a family of widely expressed G protein-coupled receptors, S1P regulates diverse physiological processes. Here we examined the role of S1P signaling in neurodegeneration using a mouse model of Sandhoff Disease, a prototypical neuronopathic lysosomal storage disorder. When sphingosine kinase 1 (Sphk1) was deleted in Sandhoff Disease mice, a milder Disease course occurred, with decreased proliferation of glial cells and less-pronounced astrogliosis. A similar result of milder Disease course and reduced astroglial proliferation was obtained by deletion of the gene for the S1P3 receptor, a G protein-coupled receptor enriched in astrocytes. Our studies demonstrate a functional role of S1P synthesis and receptor expression in astrocyte proliferation leading to astrogliosis during the terminal stages of neurodegeneration in Sandhoff Disease mice. Because astrocyte responses are involved in many types of neurodegeneration, the Sphk1/S1P receptor signaling axis may be generally important during the pathogenesis of neurodegenerative Diseases.
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deletion of macrophage inflammatory protein 1α retards neurodegeneration in Sandhoff Disease mice
Proceedings of the National Academy of Sciences of the United States of America, 2004Co-Authors: Yunping Wu, Richard L ProiaAbstract:Sandhoff Disease is a prototypical lysosomal storage disorder in which a heritable deficiency of a lysosomal enzyme, β-hexosaminidase, results in the storage of the enzyme's substrates in lysosomes. As with many of the other lysosomal storage Diseases, neurodegeneration is a prominent feature. Although the cellular and molecular pathways that underlie the neurodegenerative process are not yet fully understood, macrophage/microglial-mediated inflammation has been suggested as one possible mechanism. We now show that the expanded macrophage/microglial population in the CNS of Sandhoff Disease mice is compounded by the infiltration of cells from the periphery. Coincident with the cellular infiltration was an increased expression of macrophage-inflammatory protein 1α (MIP-1α), a leukocyte chemokine, in astrocytes. Deletion of MIP-1α expression resulted in a substantial decrease in infiltration and macrophage/microglial-associated pathology together with neuronal apoptosis in Sandhoff Disease mice. These mice without MIP-1α showed improved neurologic status and a longer lifespan. The results indicate that the pathogenesis of Sandhoff Disease involves an increase in MIP-1α that induces monocytes to infiltrate the CNS, expand the activated macrophage/microglial population, and trigger apoptosis of neurons, resulting in a rapid neurodegenerative course.
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enhanced survival in Sandhoff Disease mice receiving a combination of substrate deprivation therapy and bone marrow transplantation
Blood, 2001Co-Authors: Mylvaganam Jeyakumar, Terry D. Butters, Richard L Proia, Raymond A. Dwek, Francine Norflus, Cynthia J. Tifft, Mario Cortinaborja, V H Perry, Frances M. PlattAbstract:Sandhoff Disease is a lysosomal storage disorder characterized by GM2 ganglioside accumulation in the central nervous system (CNS) and periphery. It results from mutations in the HEXB gene, causing a deficiency in β-hexosaminidase. Bone marrow transplantation (BMT), which augments enzyme levels, and substrate deprivation (using the glycosphingolipid biosynthesis inhibitor N -butyldeoxynojirimycin [ N B-DNJ]) independently have been shown to extend life expectancy in a mouse model of Sandhoff Disease. The efficacy of combining these 2 therapies was evaluated. Sandhoff Disease mice treated with BMT and N B-DNJ survived significantly longer than those treated with BMT or N B-DNJ alone. When the mice were subdivided into 2 groups on the basis of their donor bone marrow–derived CNS enzyme levels, the high enzyme group exhibited a greater degree of synergy (25%) than the group as a whole (13%). Combination therapy may therefore be the strategy of choice for treating the infantile onset Disease variants.
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Bone marrow transplantation prolongs life span and ameliorates neurologic manifestations in Sandhoff Disease mice.
The Journal of clinical investigation, 1998Co-Authors: Francine Norflus, Cynthia J. Tifft, Michael P. Mcdonald, Gregory Goldstein, Jacqueline N. Crawley, Alexander Hoffmann, Kinuko Suzuki, Richard L ProiaAbstract:The GM2 gangliosidoses are a group of severe, neurodegenerative conditions that include Tay-Sachs Disease, Sandhoff Disease, and the GM2 activator deficiency. Bone marrow transplantation (BMT) was examined as a potential treatment for these disorders using a Sandhoff Disease mouse model. BMT extended the life span of these mice from approximately 4.5 mo to up to 8 mo and slowed their neurologic deterioration. BMT also corrected biochemical deficiencies in somatic tissues as indicated by decreased excretion of urinary oligosaccharides, and lower glycolipid storage and increased levels of beta-hexosaminidase activity in visceral organs. Even with neurologic improvement, neither clear reduction of brain glycolipid storage nor improvement in neuronal pathology could be detected, suggesting a complex pathogenic mechanism. Histological analysis revealed beta-hexosaminidase-positive cells in the central nervous system and visceral organs with a concomitant reduction of colloidal iron-positive macrophages. These results may be important for the design of treatment approaches for the GM2 gangliosidoses.
Victoria J. Mccurdy - One of the best experts on this subject based on the ideXlab platform.
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Original Article AAV-Mediated Gene Delivery in a Feline Model of Sandhoff Disease Corrects Lysosomal Storage in the Central Nervous System
2016Co-Authors: Hannah E Rockwell, Victoria J. Mccurdy, Allison M Bradbury, Aime K. Johnson, Diane U Wilson, Henry J Baker, Samuel C. Eaton, Ashley N. R, Heather L. Gray-edwards, Judith A HudsonAbstract:Sandhoff Disease (SD) is an autosomal recessive neurodegenerative Disease caused by a mutation in the gene for the b-subunit of b-N-acetylhexosaminidase (Hex), resulting in the inability to catabolize ganglioside GM2 within the lysosomes. SD presents with an accumulation of GM2 and its asialo derivative GA2, primarily in the central nervous system. Myelin-enriched glycolipids, cerebrosides and sulfatides, are also decreased in SD corresponding with dysmyelination. At present, no treatment exists for SD. Previous studies have shown the therapeutic benefit of adeno-associated virus (AAV) vector-mediated gene therapy in the treatment of SD in murine and feline models. In this study, we treated presymptomatic SD cats with AAVrh8 vectors expressing feline Hex in the thalamus combined with intracerebroventricular (Thal/ICV) injections. Treated animals showed clearly improved neurologic function and quality of life, manifested in part by prevention or attenu-ation of whole-body tremors characteristic of untreated animals. Hex activity was significantly elevated, whereas storage of GM2 and GA2 was significantly decreased in tissue samples taken from the cortex, cerebellum, thalamus, and cervical spinal cord. Treatment also increased levels of myelin-enriched cerebrosides and sulfatides in the cortex and thalamus. This study demonstrates the therapeutic potential of AAV for feline SD and suggests a similar potential for human SD patients
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aav mediated gene delivery in a feline model of Sandhoff Disease corrects lysosomal storage in the central nervous system
Asn Neuro, 2015Co-Authors: Hannah E Rockwell, Victoria J. Mccurdy, Allison M Bradbury, Heather L Grayedwards, Ashley N Randle, Aime K. Johnson, Samuel Eaton, Diane U Wilson, Henry J Baker, Judith A HudsonAbstract:Sandhoff Disease (SD) is an autosomal recessive neurodegenerative Disease caused by a mutation in the gene for the b-subunit of b-N-acetylhexosaminidase (Hex), resulting in the inability to catabolize ganglioside GM2 within the lysosomes. SD presents with an accumulation of GM2 and its asialo derivative GA2, primarily in the central nervous system. Myelinenriched glycolipids, cerebrosides and sulfatides, are also decreased in SD corresponding with dysmyelination. At present, no treatment exists for SD. Previous studies have shown the therapeutic benefit of adeno-associated virus (AAV) vectormediated gene therapy in the treatment of SD in murine and feline models. In this study, we treated presymptomatic SD cats with AAVrh8 vectors expressing feline Hex in the thalamus combined with intracerebroventricular (Thal/ICV) injections. Treated animals showed clearly improved neurologic function and quality of life, manifested in part by prevention or attenuation of whole-body tremors characteristic of untreated animals. Hex activity was significantly elevated, whereas storage of GM2 and GA2 was significantly decreased in tissue samples taken from the cortex, cerebellum, thalamus, and cervical spinal cord. Treatment also increased levels of myelin-enriched cerebrosides and sulfatides in the cortex and thalamus. This study demonstrates the therapeutic potential of AAV for feline SD and suggests a similar potential for human SD patients.
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widespread correction of central nervous system Disease after intracranial gene therapy in a feline model of Sandhoff Disease
Gene Therapy, 2015Co-Authors: Victoria J. Mccurdy, Brandon L. Brunson, Misako Hwang, Hannah E Rockwell, Allison M Bradbury, Ashley N Randle, Julian R. Arthur, Aime K. Johnson, Heather L GrayedwardsAbstract:Widespread correction of central nervous system Disease after intracranial gene therapy in a feline model of Sandhoff Disease
Cynthia J. Tifft - One of the best experts on this subject based on the ideXlab platform.
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deprivation therapy and
2016Co-Authors: Mylvaganam Jeyakumar, Terry D. Butters, Richard L Proia, Raymond A. Dwek, Francine Norflus, Cynthia J. Tifft, Mario Cortina-borja, Hugh V. Perry, Frances M. PlattAbstract:survival in Sandhoff Disease mice receiving a combination of substrat
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enhanced survival in Sandhoff Disease mice receiving a combination of substrate deprivation therapy and bone marrow transplantation
Blood, 2001Co-Authors: Mylvaganam Jeyakumar, Terry D. Butters, Richard L Proia, Raymond A. Dwek, Francine Norflus, Cynthia J. Tifft, Mario Cortinaborja, V H Perry, Frances M. PlattAbstract:Sandhoff Disease is a lysosomal storage disorder characterized by GM2 ganglioside accumulation in the central nervous system (CNS) and periphery. It results from mutations in the HEXB gene, causing a deficiency in β-hexosaminidase. Bone marrow transplantation (BMT), which augments enzyme levels, and substrate deprivation (using the glycosphingolipid biosynthesis inhibitor N -butyldeoxynojirimycin [ N B-DNJ]) independently have been shown to extend life expectancy in a mouse model of Sandhoff Disease. The efficacy of combining these 2 therapies was evaluated. Sandhoff Disease mice treated with BMT and N B-DNJ survived significantly longer than those treated with BMT or N B-DNJ alone. When the mice were subdivided into 2 groups on the basis of their donor bone marrow–derived CNS enzyme levels, the high enzyme group exhibited a greater degree of synergy (25%) than the group as a whole (13%). Combination therapy may therefore be the strategy of choice for treating the infantile onset Disease variants.
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Bone marrow transplantation prolongs life span and ameliorates neurologic manifestations in Sandhoff Disease mice.
The Journal of clinical investigation, 1998Co-Authors: Francine Norflus, Cynthia J. Tifft, Michael P. Mcdonald, Gregory Goldstein, Jacqueline N. Crawley, Alexander Hoffmann, Kinuko Suzuki, Richard L ProiaAbstract:The GM2 gangliosidoses are a group of severe, neurodegenerative conditions that include Tay-Sachs Disease, Sandhoff Disease, and the GM2 activator deficiency. Bone marrow transplantation (BMT) was examined as a potential treatment for these disorders using a Sandhoff Disease mouse model. BMT extended the life span of these mice from approximately 4.5 mo to up to 8 mo and slowed their neurologic deterioration. BMT also corrected biochemical deficiencies in somatic tissues as indicated by decreased excretion of urinary oligosaccharides, and lower glycolipid storage and increased levels of beta-hexosaminidase activity in visceral organs. Even with neurologic improvement, neither clear reduction of brain glycolipid storage nor improvement in neuronal pathology could be detected, suggesting a complex pathogenic mechanism. Histological analysis revealed beta-hexosaminidase-positive cells in the central nervous system and visceral organs with a concomitant reduction of colloidal iron-positive macrophages. These results may be important for the design of treatment approaches for the GM2 gangliosidoses.
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mice lacking both subunits of lysosomal β hexosaminidase display gangliosidosis and mucopolysaccharidosis
Nature Genetics, 1996Co-Authors: Kazunori Sango, Cynthia J. Tifft, Michael P. Mcdonald, Jacqueline N. Crawley, Michelle L. Mack, Elisa Skop, Christopher M. Starr, Alexander HoffmannAbstract:The GM2 gangliosidoses, Tay-Sachs and Sandhoff Diseases, are caused by mutations in the HEXA (α-subunit) and HEXB (β-subunit) genes, respectively. Each gene encodes a subunit for the heterodimeric lysosomal enzyme, (β-hexosaminidase A (αβ), as well as for the homodimers β-hexosaminidase B (ββ) and S (αα). In this study, we have produced mice that have both Hexa and Hexb genes disrupted through interbreeding Tay-Sachs1 (Hexa−/−) and Sandhoff2 (Hexb-/-) Disease model mice. Lacking both the α and β-subunits these ‘double knockout’ mice displayed a total deficiency of all forms of lysosomal β-hexosaminidase including the small amount of β-hexosaminidase S present in the Sandhoff Disease model mice. More surprisingly, these mice showed the phenotypic, pathologic and biochemical features of the mucopolysaccharidoses, lysosomal storage Diseases caused by the accumulation of glycosaminoglycans. The mucopolysaccharidosis phenotype is not seen in the Tay-Sachs or Sandhoff Disease model mice or in the corresponding human patients3,4,5. This result demonstrates that glycosaminoglycans are crucial substrates for β-hexosaminidase and that their lack of storage in Tay-Sachs and Sandhoff Diseases is due to functional redundancy in the β-hexosaminidase enzyme system.
Kohji Itoh - One of the best experts on this subject based on the ideXlab platform.
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Lyso-GM2 Ganglioside: A Possible Biomarker of Tay-Sachs Disease and Sandhoff Disease
PloS one, 2011Co-Authors: Takashi Kodama, Keisuke Kitakaze, Kohji Itoh, Takahiro Tsukimura, Tadayasu Togawa, Ikuo Kawashima, Yo-ichi Ishida, Minoru SuzukiAbstract:To find a new biomarker of Tay-Sachs Disease and Sandhoff Disease. The lyso-GM2 ganglioside (lyso-GM2) levels in the brain and plasma in Sandhoff mice were measured by means of high performance liquid chromatography and the effect of a modified hexosaminidase (Hex) B exhibiting Hex A-like activity was examined. Then, the lyso-GM2 concentrations in human plasma samples were determined. The lyso-GM2 levels in the brain and plasma in Sandhoff mice were apparently increased compared with those in wild-type mice, and they decreased on intracerebroventricular administration of the modified Hex B. The lyso-GM2 levels in plasma of patients with Tay-Sachs Disease and Sandhoff Disease were increased, and the increase in lyso-GM2 was associated with a decrease in Hex A activity. Lyso-GM2 is expected to be a potential biomarker of Tay-Sachs Disease and Sandhoff Disease.
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Prostaglandin E2 Reverses Aberrant Production of an Inflammatory Chemokine by Microglia from Sandhoff Disease Model Mice through the cAMP-PKA Pathway
2011Co-Authors: Eri Kawashita, Daisuke Tsuji, Masahiro Toyoshima, Yosuke Kanno, Hiroyuki Matsuno, Kohji ItohAbstract:Background: Sandhoff Disease (SD) is a neurodegenerative lysosomal b-hexosaminidase (Hex) deficiency involving excessive accumulation of undegraded substrates, including terminal GlcNAc-oligosaccharides and GM2 ganglioside. Microglia-mediated neuroinflammation contributes to the pathogenesis and progression of SD. Our previous study demonstrated that MIP-1a, a putative pathogenic factor for SD, is up-regulated in microglial cells derived from SD model mice (SD-Mg) through activation of Akt and JNK. Methodology/Principal Findings: In this study, we first demonstrated that prostaglandin E2 (PGE2), which is one of the lipid mediators derived from arachidonic acid and is known to suppress activation of microglia, reduced the aberrant MIP-1a production by SD-Mg to the same level as by WT-Mg. PGE2 also attenuated the activation of Akt and JNK. The inhibition of MIP-1a production and the activation of Akt and JNK occurred through the EP2 and 4/cAMP/PKA signaling pathway in the murine microglia derived from SD model mice. Conclusions/Significance: We propose that PGE2 plays a role as a negative regulator of MIP-1a production in th
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mechanism of abnormal growth in astrocytes derived from a mouse model of gm2 gangliosidosis
Journal of Neurochemistry, 2009Co-Authors: Nagako Kawashima, Kohji Itoh, Daisuke Tsuji, Tetsuya Okuda, Kenichi NakayamaAbstract:Sandhoff Disease is a progressive neurodegenerative disorder caused by mutations in the HEXB gene which encodes the β-subunit of N-acetyl-β-hexosaminidase A and B, resulting in the accumulation of the ganglioside GM2. We isolated astrocytes from the neonatal brain of Sandhoff Disease model mice in which the N-acetyl-β-hexosaminidase β-subunit gene is genetically disrupted (ASD). Glycolipid profiles revealed that GM2/GA2 accumulated in the lysosomes and not on the cell surface of ASD astrocytes. In addition, GM3 was increased on the cell surface. We found remarkable differences in the cell proliferation of ASD astrocytes when compared with cells isolated from wild-type mice, with a faster growth rate of ASD cells. In addition, we observed increased extracellular, signal-regulated kinase (ERK) phosphorylation in ASD cells, but Akt phosphorylation was decreased. Furthermore, the phosphorylation of ERK in ASD cells was not dependent upon extracellular growth factors. Treatment of ASD astrocytes with recombinant N-acetyl-β-hexosaminidase A resulted in a decrease of their growth rate and ERK phosphorylation. These results indicated that the up-regulation of ERK phosphorylation and the increase in proliferation of ASD astrocytes were dependent upon GM2/GA2 accumulation. These findings may represent a mechanism in linking the nerve cell death and reactive gliosis observed in Sandhoff Disease.
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Therapeutic evaluation of GM2 gangliosidoses by ELISA using anti-GM2 ganglioside antibodies
Clinica Chimica Acta, 2006Co-Authors: Yukari Higashine, Hitoshi Sakuraba, Kazuhiko Matsuoka, Kohji ItohAbstract:Abstract Background GM2 gangliosidoses, including Tay-Sachs Disease, Sandhoff Disease and the AB variant, comprise deficiencies of β-hexosaminidase isozymes and GM2 ganglioside activator protein associated with accumulation of GM2 ganglioside (GM2) in lysosomes and neurosomatic clinical manifestations. A simple assay system for intracellular quantification of GM2 is required to evaluate the therapeutic effects on GM2-gangliosidoses. Methods We newly established a cell–ELISA system involving anti-GM2 monoclonal antibodies for measuring GM2 storage in fibroblasts from Tay-Sachs and Sandhoff Disease patients. Results We succeeded in detecting the corrective effect of enzyme replacement on elimination of GM2 in the cells with this ELISA system. Conclusions This simple and sensitive system should be useful as additional diagnosis tool as well as therapeutic evaluation of GM2 gangliosidoses.
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Inefficiency in GM2 Ganglioside Elimination by Human Lysosomal β-Hexosaminidase β-Subunit Gene Transfer to Fibroblastic Cell Line Derived from Sandhoff Disease Model Mice
Biological & pharmaceutical bulletin, 2006Co-Authors: Tomohiro Itakura, Eri Kawashita, Shoji Yamanaka, Hitoshi Sakuraba, Yukari Higashine, Yasuhiro Ishibashi, Aya Kuroki, Kohji ItohAbstract:Sandhoff Disease (SD) is an autosomal recessive GM2 gangliosidosis caused by the defect of lysosomal beta-hexosaminidase (Hex) beta-subunit gene associated with neurosomatic manifestations. Therapeutic effects of Hex subunit gene transduction have been examined on Sandhoff Disease model mice (SD mice) produced by the allelic disruption of Hexb gene encoding the murine beta-subunit. We demonstrate here that elimination of GM2 ganglioside (GM2) accumulated in the fibroblastic cell line derived from SD mice (FSD) did not occur when the HEXB gene only was transfected. In contrast, a significant increase in the HexB (betabeta homodimer) activity toward neutral substrates, including GA2 (asialo-GM2) and oligosaccharides carrying the terminal N-acetylglucosamine residues at their non-reducing ends (GlcNAc-oligosaccharides) was observed. Immunoblotting with anti-human HexA (alphabeta heterodimer) serum after native polyacrylamide gel electrophoresis (Native-PAGE) revealed that the human HEXB gene product could hardly form the chimeric HexA through associating with the murine alpha-subunit. However, co-introduction of the HEXA encoding the human alpha-subunit and HEXB genes caused significant corrective effect on the GM2 degradation by producing the human HexA. These results indicate that the recombinant human HexA could interspeciesly associate with the murine GM2 activator protein to degrade GM2 accumulated in the FSD cells. Thus, therapeutic effects of the recombinant human HexA isozyme but not human HEXB gene product could be evaluated by using the SD mice.
