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Dette review omhandler den genetiske baggrund for gangliosidosis type 1 og type 2 hos den Japanske hunderace Shiba Inu. Gangliosidosis er en arvelig letal sygdom, der påvirker hele nervesystemet idet nerveceller gradvist nedbrydes. De to typer skyldes deletion mutationer i GLB1(GM1) og HEXB (GM2) generne. For at undgå syge individer med gangliosidosis, kan gentestning måske være vejen frem. TaqMan er en god testmetode, der kan bruges i evt. fremtidigt avlsarbejde. GM1 og GM2 hos Shiba Inu, minder meget om det sygdomsbillede vi ser i andre hunderacer og arter, som katte, får, kvæg og mennesker. GM1 hos Shiba Inu minder især meget om den juvenile humane type af GM1 gangliosidosis der ses hos mennesker. Shibaen kan være en god dyremodel for GM1 gangliosidosis hos mennesker på baggrund af sygdomsbillede og GLB1-genet.

Dette review omhandler den genetiske baggrund for gangliosidosis type 1 og type 2 hos den Japanske hunderace Shiba Inu. Gangliosidosis er en arvelig letal sygdom, der påvirker hele nervesystemet idet nerveceller gradvist nedbrydes. De to typer skyldes deletion mutationer i GLB1(GM1) og HEXB (GM2) generne. For at undgå syge individer med gangliosidosis, kan gentestning måske være vejen frem. TaqMan er en god testmetode, der kan bruges i evt. fremtidigt avlsarbejde. GM1 og GM2 hos Shiba Inu, minder meget om det sygdomsbillede vi ser i andre hunderacer og arter, som katte, får, kvæg og mennesker. GM1 hos Shiba Inu minder især meget om den juvenile humane type af GM1 gangliosidosis der ses hos mennesker. Shibaen kan være en god dyremodel for GM1 gangliosidosis hos mennesker på baggrund af sygdomsbillede og GLB1-genet.

Mammalian sperm undergo several maturation steps after leaving the testis to become competent for fertilization. Important changes occur in sperm within the female reproductive tract, although the molecular mechanisms underlying these processes remain unclear. To investigate sperm membrane remodeling upon sperm maturation, we developed transgenic mouse lines carrying glycosylphosphatidylinositol (GPI)-anchored enhanced green fluorescent protein (EGFP-GPI) and traced the fate of this fluorescent protein during the fertility-acquiring process in sperm in vitro and in vivo. When the GFP-labeled sperm were treated with compounds for promoting the acrosome reaction, EGFP-GPI was released from the sperm surface crosslinked with characteristic relocation of a lipid raft marker ganglioside GM1. Sperm ejaculated into the uterus strongly expressed EGFP-GPI in the head region, whereas a part of the oviductal sperm lost fluorescence in a manner that was dependent on the presence of angiotensin-converting enzyme (ACE). Moreover, sperm on the zona pellucida of eggs in the oviduct were all found to have low levels of GFP. These results suggest that sperm undergoing GPI-anchored protein release associated with reorganization of lipid rafts and the acrosome reaction acquire fertilization potential.

Gangliosides are target receptors for bacterial entry, yet those present in human milk exhibit a protective role against bacterial infection. Here, we show that treatment with ganglioside mixture at a concentration of 100 microg/mL resulted in significant inhibition of the vacuole formation activity of Helicobacter pylori vacuolating cytotoxin (VacA) in gastric epithelial cancer AZ-521 cells. All gangliosides (GM1, GM2, GM3, GD1a, GD1b, GD3 and GT1b) examined showed good neutralizing capacity against VacA. A pull-down assay was performed using lyso-GM1 coupled to Sepharose as the tagged polysaccharide polymer to capture VacA from H. pylori culture supernatant. GM1-VacA complexes were successfully precipitated, suggesting that GM1 binds directly to VacA. The hydrodynamic binding of lyso-GM1 and VacA measured by fluorescence correlation spectroscopy had a K(d) value of 190 nM. VacA also bound to lyso-GM1 at pH 2 corresponding to the physiological pH of human stomach. Collectively, these results showed that direct binding of H. pylori VacA to free gangliosides neutralizes the toxin activity of VacA. These findings offer an alternative insight into the role of gangliosides in VacA toxicity and the pathogenesis of H. pylori.

Gangliosides are target receptors for bacterial entry, yet those present in human milk exhibit a protective role against bacterial infection. Here, we show that treatment with ganglioside mixture at a concentration of 100 microg/mL resulted in significant inhibition of the vacuole formation activity of Helicobacter pylori vacuolating cytotoxin (VacA) in gastric epithelial cancer AZ-521 cells. All gangliosides (GM1, GM2, GM3, GD1a, GD1b, GD3 and GT1b) examined showed good neutralizing capacity against VacA. A pull-down assay was performed using lyso-GM1 coupled to Sepharose as the tagged polysaccharide polymer to capture VacA from H. pylori culture supernatant. GM1-VacA complexes were successfully precipitated, suggesting that GM1 binds directly to VacA. The hydrodynamic binding of lyso-GM1 and VacA measured by fluorescence correlation spectroscopy had a K(d) value of 190 nM. VacA also bound to lyso-GM1 at pH 2 corresponding to the physiological pH of human stomach. Collectively, these results showed that direct binding of H. pylori VacA to free gangliosides neutralizes the toxin activity of VacA. These findings offer an alternative insight into the role of gangliosides in VacA toxicity and the pathogenesis of H. pylori.

Gangliosides are target receptors for bacterial entry, yet those present in human milk exhibit a protective role against bacterial infection. Here, we show that treatment with ganglioside mixture at a concentration of 100 microg/mL resulted in significant inhibition of the vacuole formation activity of Helicobacter pylori vacuolating cytotoxin (VacA) in gastric epithelial cancer AZ-521 cells. All gangliosides (GM1, GM2, GM3, GD1a, GD1b, GD3 and GT1b) examined showed good neutralizing capacity against VacA. A pull-down assay was performed using lyso-GM1 coupled to Sepharose as the tagged polysaccharide polymer to capture VacA from H. pylori culture supernatant. GM1-VacA complexes were successfully precipitated, suggesting that GM1 binds directly to VacA. The hydrodynamic binding of lyso-GM1 and VacA measured by fluorescence correlation spectroscopy had a K(d) value of 190 nM. VacA also bound to lyso-GM1 at pH 2 corresponding to the physiological pH of human stomach. Collectively, these results showed that direct binding of H. pylori VacA to free gangliosides neutralizes the toxin activity of VacA. These findings offer an alternative insight into the role of gangliosides in VacA toxicity and the pathogenesis of H. pylori., Glycobiology, 20(6), pp.668-678; 2010

TTNatural and semisynthetic gangliosides protect neurons from toxin-induced cell death and salvage neurons after toxin exposure. The hydrophilic property of gangliosides restricts their blood-brain barrier (BBB) permeability, which hinders their use as neuroprotective agents. Gangliosides semisynthetic derivatives with improved cytoprotective properties and BBB permeability can be produced. Even with gangliosides great therapeutic promise, no study has examined ganglioside functional group derivatives that would provide cytoprotection AND effectively cross the BBB; information that would provide a basis for future studies of neuroprotective mechanisms. This study examined the ability of ganglioside derivatives to be cytoprotective in vitro models using the dopaminergic neurotoxin, 1-methyl-4-phenylpyridinium (MPP+) and the SH-SY5Y cell line. Derivatives determined to have therapeutic potential were to be tested in vitro for their ability to cross a brain capillary endothelial cell culture model of the BBB. Finally, derivatives that were both cytoprotective and that effectively crossed the in vitro BBB model were to be tested in vivo for their ability to neuroprotect dopaminergic neurons in both chronic and acute neurotoxicity models using the MPP+ precursor, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). The hypothesis is that changes in ganglioside ceramide and/or oligosaccharide functional groups can improve neuroprotection through changes in cytoprotection and BBB transcytosis., The original document contains color images.

G(M1)-gangliosidosis is a lysosomal storage disorder caused by a deficiency of beta-galactosidase (GLB1). The GLB1 gene gives rise to the GLB1 lysosomal enzyme and to the elastin binding protein (EBP), involved in elastic fiber deposition. GLB1 forms a complex with protective protein cathepsin A (PPCA), alpha neuraminidase (NEU1), and galactosamine 6-sulphate sulfatase (GALNS) inside lysosomes, while EBP binds to PPCA and NEU1 on the cell surface. We investigated the function of the GLB1 and EBP mutated proteins by analyzing the clinical, genetic, and cellular data of 11 G(M1)-gangliosidosis patients. Their molecular analysis, followed by expression studies, lead to the identification of four new and 10 known GLB1 mutations. Some common amino acid substitutions [c.1445G>A (p.Arg482H), c.622C>T (p.Arg208His), c.175C>T (p.Arg59Cys) and c.176G>A (p.Arg59His)] were present in the GLB1 enzyme of several patients, all of Mediterranean origin, suggesting a common origin. Western blotting analyses against GLB1, EBP, and PPCA proteins showed that the identified mutations affect GLB1 enzyme activity and/or stability. The c.1445G>A (p.Arg482His), c.175C>T (p.Arg59Cys), c.733+2T>C, c.1736G>A (p.Gly579Asp), and c.1051C>T (p.Arg351X) GLB1 mutations, affect the stabilization of PPCA probably because they hamper the interaction between GLB1/EBP and PPCA within the multiprotein complex. The amount of EBP was normal, but the detection of impaired elastogenesis in such patients suggests an alteration in its function. We conclude that the presence of genetic lesions in both GLB1 and EBP coding region does not directly predict impaired elastogenesis and that elastic fiber assembly has to be evaluated specifically in each case. Nevertheless, the degree of EBP involvement may be linked to specific clinical findings.

G(M1)-gangliosidosis is a lysosomal storage disorder caused by a deficiency of beta-galactosidase (GLB1). The GLB1 gene gives rise to the GLB1 lysosomal enzyme and to the elastin binding protein (EBP), involved in elastic fiber deposition. GLB1 forms a complex with protective protein cathepsin A (PPCA), alpha neuraminidase (NEU1), and galactosamine 6-sulphate sulfatase (GALNS) inside lysosomes, while EBP binds to PPCA and NEU1 on the cell surface. We investigated the function of the GLB1 and EBP mutated proteins by analyzing the clinical, genetic, and cellular data of 11 G(M1)-gangliosidosis patients. Their molecular analysis, followed by expression studies, lead to the identification of four new and 10 known GLB1 mutations. Some common amino acid substitutions [c.1445G>A (p.Arg482H), c.622C>T (p.Arg208His), c.175C>T (p.Arg59Cys) and c.176G>A (p.Arg59His)] were present in the GLB1 enzyme of several patients, all of Mediterranean origin, suggesting a common origin. Western blotting analyses against GLB1, EBP, and PPCA proteins showed that the identified mutations affect GLB1 enzyme activity and/or stability. The c.1445G>A (p.Arg482His), c.175C>T (p.Arg59Cys), c.733+2T>C, c.1736G>A (p.Gly579Asp), and c.1051C>T (p.Arg351X) GLB1 mutations, affect the stabilization of PPCA probably because they hamper the interaction between GLB1/EBP and PPCA within the multiprotein complex. The amount of EBP was normal, but the detection of impaired elastogenesis in such patients suggests an alteration in its function. We conclude that the presence of genetic lesions in both GLB1 and EBP coding region does not directly predict impaired elastogenesis and that elastic fiber assembly has to be evaluated specifically in each case. Nevertheless, the degree of EBP involvement may be linked to specific clinical findings.