Rapid detection of GM1 ganglioside in cerebrospinal fluid in dogs with GM1 gangliosidosis using matrix-assisted laser desorption ionization time-of-flight mass spectrometry.

The concentration of GM1 (monosialotetrahexosyl ganglioside) in cerebrospinal fluid (CSF) is markedly increased in dogs with GM1 gangliosidosis due to GM1 accumulation in the central nervous system and leakage to the CSF. The present study established a rapid and simple method for detection of accumulated GM1 in the CSF in dogs with GM1 gangliosidosis using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI TOF MS) and discusses the usefulness of this method for the rapid diagnosis and/or high-risk screening of this disease in domestic animals. Cerebrospinal fluid was collected from normal dogs and 4- to 11-month-old Shiba dogs with GM1 gangliosidosis. The MALDI TOF MS analysis was carried out in combination with a special sample plate and a simple desalting step on the plate. Specific signs of GM1 could be detected in the standard GM1 solutions at concentrations of 50 nmol/l or more. The signs were also clearly detected in CSF (131-618 nmol/l) in affected dogs, but not in normal canine CSF (12 ± 5 nmol/l, mean ± standard deviation). The results demonstrated that MALDI TOF MS can detect GM1 accumulated in canine CSF even in the early stage of the disease. In conclusion, the rapid detection of increased CSF GM1 using MALDI TOF MS is a useful method for diagnosis and/or screening for canine GM1 gangliosidosis.

Cerebrospinal fluid biomarkers showing neurodegeneration in dogs with GM1 gangliosidosis: possible use for assessment of a therapeutic regimen.

The present study investigated cerebrospinal fluid (CSF) biomarkers for estimating degeneration of the central nervous system (CNS) in experimental dogs with GM1 gangliosidosis and preliminarily evaluated the efficacy of long-term glucocorticoid therapy for GM1 gangliosidosis using the biomarkers identified here. GM1 gangliosidosis, a lysosomal storage disease that affects the brain and multiple systemic organs, is due to an autosomal recessively inherited deficiency of acid beta-galactosidase activity. Pathogenesis of GM1 gangliosidosis may include neuronal apoptosis and abnormal axoplasmic transport and inflammatory response, which are perhaps consequent to massive neuronal storage of GM1 ganglioside. In the present study, we assessed some possible CSF biomarkers, such as GM1 ganglioside, aspartate aminotransferase (AST), lactate dehydrogenase (LDH), neuron-specific enolase (NSE) and myelin basic protein (MBP). Periodic studies demonstrated that GM1 ganglioside concentration, activities of AST and LDH, and concentrations of NSE and MBP in CSF were significantly higher in dogs with GM1 gangliosidosis than those in control dogs, and their changes were well related with the months of age and clinical course. In conclusion, GM1 ganglioside, AST, LDH, NSE and MBP could be utilized as CSF biomarkers showing CNS degeneration in dogs with GM1 gangliosidosis to evaluate the efficacy of novel therapies proposed for this disease. In addition, we preliminarily treated an affected dog with long-term oral administration of prednisolone and evaluated the efficacy of this therapeutic trial using CSF biomarkers determined in the present study. However, this treatment did not change either the clinical course or the CSF biomarkers of the affected dog, suggesting that glucocorticoid therapy would not be effective for treating GM1 gangliosidosis.

Use of amnion and placenta in neonatal screening for canine GM1-gangliosidosis and the risk of diagnostic misclassifications.

BACKGROUND: A closed breeding colony of Shiba dogs with GM1-gangliosidosis is maintained at Hokkaido University (Sapporo, Japan). Neonatal genotyping is essential to control the breeding colony genetically as an animal model for the human disease. OBJECTIVES: The purpose of the present study was to determine the utility of amnion and placenta in the neonatal screening or diagnosis for canine GM1-gangliosidosis. METHODS: Twenty neonatal Shiba dogs of a pedigree with GM1- gangliosidosis were differentiated into 3 genotypes--normal, heterozygous, and affected dogs--by using a previously reported DNA mutation assay. Acid beta-galactosidase activity was measured in amnion and placenta and compared among the 3 genotypes. RESULTS: The level of beta-galactosidase activity in the amnion of affected dogs was negligible and <2% of the mean activity in normal dogs; there was no significant difference among the 3 genotypes. In placenta, beta-galactosidase activity was significantly different among all the genotypes; however, there was wide overlap in enzyme activity between normal and heterozygous dogs. The level of activity in affected dogs was relatively high and >10% of the mean activity in normal dogs. The DNA mutation assay gave correct information about genotype with genomic DNA extracted from amnion but ambiguous information with DNA from placenta. CONCLUSIONS: Amnion and placenta were not useful as enzyme sources in neonatal screening in canine GM1-gangliosidosis because of the risk of misdiagnosis. DNA from amnion is applicable as a template for genotyping, whereas placenta should not be used because canine placenta contains maternal cells.