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[This corrects the article DOI: 10.1371/journal.pone.0060674.].
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Gaucher disease (GD) is characterized by a marked phenotypic and genetic diversity. It is caused by the functional deficiency of the lysosomal enzyme ß-glucocerebrosidase (GCase), which in most instances results from mutations in the GBA1 gene and over 500 different disease causing mutations have been described. We present the biochemical and molecular findings in 141 GD cases (14 were siblings) with the three types of the disorder diagnosed in Greece over the last 35 years. 111/141 (78%) GD patients were of Greek origin. The remaining patients were Albanian (24/141; 17%), Syrian (2/141; 1.4%), Egyptian (2/141; 1.4%), Italian (1/141; 0.7%) and Polish (1/141; 0.7%). Mutation analysis identified 28 different mutations and 37 different genotypes. Seven of the mutations were not previously reported (T231I, D283N, N462Y, LI75P, F81L, Y135S and T482K). The most frequent mutations were N370S, D409H;H255Q and L444P. Mutation D409H;H255Q was only identified in Greek and Albanian patients. Sixteen mutations, including the novel ones, were identified only in one allele. Although the N370S mutation was identified only in type 1 patients, not all of type 1 patients carried this mutation. Our results highlight the heterogeneity of Gaucher disease and support the Balkan origin of the double mutant allele D409H;H255Q.
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Lysosomal Storage Diseases (LSDs) are rare genetic diseases, the majority of which are caused by specific lysosomal enzyme deficiencies and all are characterized by malfunctioning lysosomes. Lysosomes are key regulators of many different cellular processes and are vital for the function of the immune system. Several studies have shown the coexistence of LSDs and immune abnormalities. In this study, we investigated the presence of autoantibodies in the plasma of patients with Gaucher disease (GD; nâ¯=â¯6), Sanfilippo Syndrome B (SFB; nâ¯=â¯8) and Niemann - Pick type C disease (NPC; nâ¯=â¯5) before and following Miglustat treatment (nâ¯=â¯3). All were examined for antibodies to antigens of Hep-2 cells and antiganglioside antibodies (AGSA). No autoantibodies were detected in GD patients. 3/8 SFB patients showed only AGSA (2/3 IgM / IgG; 1/3 IgG), 3/8 only anti-Sm E/F and 2/8 showed both IgM / IgG or IgG AGSA and anti-Sm E/F. 3/5 NPC patients showed AGSA (2/3 IgM and IgG, 1/3 IgM) and one anti-Sm E/F and IgM AGSA. Following treatment one patient with no AGSA developed IgM AGSA and two with both IgG and IgM showed only IgG AGSA. In our study, investigating similar numbers of patients, autoantibodies were observed in NPC and SFB patients but not in GD patients. Our findings suggest that, independently of the development of an autoimmune disease in patients with LSDs, there seems to be an autoimmune activation that differs in different disorders. Further studies including more patients, also at different stages of disease and treatment, are needed in order to get further insight into the immune irregularities associated with different LSDs and their significance.
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Niemann-Pick type C disease (NPC) is a neurovisceral lysosomal storage disease caused by mutations in either the NPC1 or the NPC2 gene. It is a cellular lipid trafficking disorder characterized by the accumulation of unesterified cholesterol and various sphingolipids in the lysosomes and late endosomes, and it exhibits a broad clinical spectrum. Today, over 420 disease-causing mutations have been identified in the NPC1 and the NPC2 genes. We present the clinical, biochemical, and molecular findings in 14 cases diagnosed in Greece during the last 28 years. Age at diagnosis ranged from 2.5 months to 48 years. Systemic manifestations were present in 7/14 patients. All developed neurological manifestations (age of onset 5 months to 42 years). Six patients are still alive (age: 5-50 years). Classical filipin staining pattern was observed in all but four patients (3 NPC1, 1 NPC2). The rate of LDL-induced cholesteryl ester formation was severely reduced in 4/7 and significantly reduced in 3/7 patients studied. Increased chitotriosidase activity was observed in 9/12 patients. Mutation analysis in 11 unrelated patients identified 12 different mutations in the NPC1 gene: eight previously described p.E1089K (c.3265G>A), p.F284Lfs*26 (c.852delT), p.A1132P(c.3394G>C), del promoter region and exons 1-10, p.R1186H (c.3557G>A), p.P1007A (c.3019C>G), p.Q92R(c.275A>G),p.S940L (c.2819C>T), and four novel ones: (p.N701K fs*13 (c.2102-2103insA), p.K1057R (c.3170A>G), IVS23+3insT(c.3591+3insT), p.C1119*(c.3357T>C); and the previously described IVS2+5G>A(c.190+5G>A) mutation in the NPC2 gene. All patients were of Greek origin. Assuming a birth rate of 100,000/year, a rough incidence estimate for NPC disease in Greece would be 0.5/100,000 births.
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Several observations suggest that disturbed homeostasis of α-Synuclein (α-Syn) may provide a link between Gaucher disease (GD) and Parkinson's disease (PD). We recently reported increased dimerization of α-Syn in the red blood cell (RBC) membrane of patients with GD. Several studies indicate a crucial relationship between lipids, oxidative stress and α-Syn status. Here we investigated the relationship between the observed increased dimerization of α-Syn in the cell membranes of RBCs, cells devoid of lysosomes and lacking lysosomal enzyme synthesis, and the lipid abnormalities and oxidative stress already described in GD. Correlation studies showed that in GD the α-Syn dimer/monomer ratio is positively correlated with the levels of glucosylceramide (GlcCer) and the glucosylceramide/ceramide (GlcCer/Cer) ratio and negatively with the levels of malonyldialdehyde (MDA) and plasmalogens. In conclusion, we have shown that the increased tendency of α-Syn to form dimers in the RBC membrane of patients with GD, is correlated with both the level of lipids, including GlcCer, the primary lipid abnormality in GD, and the increased oxidative stress observed in this disorder. The study of other tissues, and in particular brain, will be important in order to elucidate the significance of these findings regarding the link between GD and PD.
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Eritrócitos/metabolismo , Doença de Gaucher/sangue , Lipídeos/sangue , Estresse Oxidativo , alfa-Sinucleína/sangue , Adolescente , Adulto , Idoso , Estudos de Casos e Controles , Ceramidas/sangue , Pré-Escolar , Dimerização , Humanos , Malondialdeído/sangue , Pessoa de Meia-Idade , Plasmalogênios/sangue , Adulto Jovem , alfa-Sinucleína/químicaRESUMO
Krabbe disease is an autosomal recessive neurodegenerative lysosomal storage disease caused by the deficiency of ß-galactocerebrosidase. This deficiency results in the impaired degradation of ß-galactocerebroside, a major myelin lipid, and of galactosylsphingosine. Based on the age of onset of neurological symptoms, an infantile form (90% patients) and late-onset forms (10% patients) of the disease are recognized. Over 130 disease-causing mutations have been identified in the ß-galactocerebrosidase gene. We present the biochemical and molecular findings in 19 cases of Krabbe disease, 17 of them unrelated, diagnosed in Greece over the last 30 years. ß-Galactocerebrosidase activity assayed in leukocyte homogenates using either the tritium-labeled or the fluorescent substrate was diagnostic for all. Increased plasma chitotriosidase activity was found in 11/15 patients.Mutational analysis, carried out in 11 unrelated cases, identified seven different mutations, four previously described (p.I250T, c.1161+6532_polyA+9kbdel, p.K139del, p.D187V) and three novel mutations (p.D610A, c.583-1 G>C, p.W132X), and seven distinct genotypes. The most prevalent mutation was mutation p.I250T, first described in a patient of Greek origin. It accounted for 36.4% (8/22) of the mutant alleles. The second most frequent mutation was c.1161+6532_polyA+9kbdel that accounted for 22.7% (5/22) of the mutant alleles. The observed frequency was lower than that described in Northern European countries and closer to that described in Italian patients.
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Plasmalogens represent a unique class of phospholipids. Reduced red blood cell plasmalogen levels in Gaucher disease patients were reported, correlating to total disease burden. The relation between plasmalogen abnormalities in Gaucher disease patients and primary glycosphingolipid abnormalities, malonyldialdehyde levels, an indicator of lipid peroxidation, and the total antioxidant status was further investigated. Significant reduction of C16:0 and C18:0 plasmalogens in red blood cells of Gaucher disease patients was confirmed. In parallel, a significant increase in the glucosylceramide/ceramide ratio in red blood cell membranes, as well as an average 200-fold increase in plasma glucosylsphingosine levels was observed. Red blood cell malonyldialdehyde levels were significantly increased in patients, whereas their total antioxidant status was significantly reduced. A negative correlation between plasmalogen species and glucosylceramide, ceramide, glucosylceramide/ceramide ratio, glucosylsphingosine and malonyldialdehyde, significant for the C16:0 species and all the above parameters with the exception of malonyldialdehyde levels, was found along with a positive non-significant correlation with the total antioxidant status. Our results indicate that increased lipid peroxidation and reduced total antioxidant status exist in Gaucher disease patients. They demonstrate a clear link between plasmalogen levels and the primary glycolipid abnormalities characterizing the disorder and an association with the increased oxidative stress observed in Gaucher disease patients.
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Doença de Gaucher/metabolismo , Metabolismo dos Lipídeos , Estresse Oxidativo , Plasmalogênios/metabolismo , Adolescente , Adulto , Idoso , Estudos de Casos e Controles , Ceramidas/metabolismo , Criança , Pré-Escolar , Eritrócitos/metabolismo , Doença de Gaucher/sangue , Glucosilceramidas/metabolismo , Humanos , Peroxidação de Lipídeos , Pessoa de Meia-Idade , Adulto JovemRESUMO
To date, a plethora of studies have provided evidence favoring an association between Gaucher disease (GD) and Parkinson's disease (PD). GD, the most common lysosomal storage disorder, results from the diminished activity of the lysosomal enzyme ß-glucocerebrosidase (GCase), caused by mutations in the ß-glucocerebrosidase gene (GBA). Alpha-synuclein (ASYN), a presynaptic protein, has been strongly implicated in PD pathogenesis. ASYN may in part be degraded by the lysosomes and may itself aberrantly impact lysosomal function. Therefore, a putative link between deficient GCase and ASYN, involving lysosomal dysfunction, has been proposed to be responsible for the risk for PD conferred by GBA mutations. In this current work, we aimed to investigate the effects of pharmacological inhibition of GCase on ASYN accumulation/aggregation, as well as on lysosomal function, in differentiated SH-SY5Y cells and in primary neuronal cultures. Following profound inhibition of the enzyme activity, we did not find significant alterations in ASYN levels, or any changes in the clearance or formation of its oligomeric species. We further observed no significant impairment of the lysosomal degradation machinery. These findings suggest that additional interaction pathways together with aberrant GCase and ASYN must govern this complex relation between GD and PD.
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Glucosilceramidase/deficiência , Lisossomos/metabolismo , Neurônios/metabolismo , alfa-Sinucleína/metabolismo , Animais , Autofagia/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Doença de Gaucher/genética , Doença de Gaucher/metabolismo , Glucosilceramidase/antagonistas & inibidores , Humanos , Inositol/análogos & derivados , Inositol/farmacologia , Neurônios/citologia , Neurônios/efeitos dos fármacos , Doença de Parkinson/genética , Doença de Parkinson/metabolismo , Multimerização Proteica , Ratos , alfa-Sinucleína/química , alfa-Sinucleína/genéticaAssuntos
Intolerância à Frutose/diagnóstico , Frutose-Bifosfato Aldolase/deficiência , Frutose/metabolismo , Lisossomos/enzimologia , Transferrina/metabolismo , Dieta com Restrição de Carboidratos , Diagnóstico Precoce , Intolerância à Frutose/dietoterapia , Intolerância à Frutose/enzimologia , Frutose-Bifosfato Aldolase/genética , Humanos , Focalização Isoelétrica , Plasma , Isoformas de Proteínas/metabolismoAssuntos
Doença de Gaucher/diagnóstico , Doença de Gaucher/patologia , Encéfalo/patologia , Análise Mutacional de DNA , Morte , Ecocardiografia , Feminino , Doença de Gaucher/genética , Grécia , Hexosaminidases/sangue , Humanos , Recém-Nascido , Masculino , Pele/patologia , beta-Glucosidase/metabolismoRESUMO
BACKGROUND: Variable increases in the plasma activity of different lysosomal enzymes have been reported in patients with congenital disorders of glycosylation (CDG). In particular, elevated plasma aspartylglucosaminidase activity (AGA) has been found in the majority of CDG type I patients. We report on the plasma activity of AGA and other lysosomal enzymes in patients with different types of primary and secondary CDG defects. METHODS: AGA, alpha-mannosidase, beta-mannosidase and beta-hexosaminidase activities were assayed in the plasma of patients with CDGI (4CDGIa, 4CDGIx) and CDGIIx (5, all with a combined N- and O-glycosylation defect), classical galactosemia (GALT) (n=3) and hereditary fructose intolerance (HFI) (n=2). RESULTS: Increased AGA and beta-hexosaminidase activities were found in all and 7/8 of the GDGI patients respectively. All enzymic activities were normal in the CDGIIx patients. Elevated AGA and beta-hexosaminidase activity was also seen in GALT and HFI patients before treatment, when transferrin isoelectric focusing (TfIEF) patterns were also abnormal. CONCLUSIONS: Increased AGA plasma activity, although a consistent finding in CDGI patients, is not specific to this group of disorders since it is also observed in untreated cases of GALT and HFI. Furthermore, plasma AGA activity cannot serve as a marker for CDGII disorders. In conjunction with TfIEF it could be used in the follow up of GALT and HFI patients.
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Aspartilglucosilaminase/sangue , Erros Inatos do Metabolismo da Frutose/enzimologia , Galactosemias/enzimologia , Adolescente , Criança , Pré-Escolar , Intolerância à Frutose/sangue , Intolerância à Frutose/diagnóstico , Intolerância à Frutose/enzimologia , Erros Inatos do Metabolismo da Frutose/sangue , Erros Inatos do Metabolismo da Frutose/diagnóstico , Galactosemias/sangue , Glicosilação , Humanos , Lactente , Lisossomos/enzimologia , alfa-Manosidase/sangue , beta-Manosidase/sangue , beta-N-Acetil-Hexosaminidases/sangueRESUMO
Plasmalogens represent a unique type of phospholipids characterized by the presence of a vinyl-ether bond at the sn-1 position of the glycerol backbone. Peroxisomes are essential in their biosynthesis. Their suggested functions include protection against oxidative stress, participation in signal transduction, membrane fusion events, cholesterol transport and membrane trafficking, processes known to be disturbed in sphingolipidoses. We here report on red blood cell membrane plasmalogen levels in Gaucher disease patients. Plasmalogen levels were measured as their dimethylacetal derivatives (DMA) by gas chromatography in lipid extracts of erythrocytes from 15 patients. Their relative amount was estimated as the ratio between C18:0 DMA and methylstearate (C18:0), as well as C16:0 DMA and methylpalmitate (C16:0). Statistically significant lower levels of both plasmalogen species were observed in Gaucher disease patients compared to normal individuals. Furthermore, a negative correlation between plasmalogen levels and chitotriosidase was observed in the patients, which was statistically significant for the C18:0 species. Upon therapy, a significant rise of plasmalogen levels and fall in chitotriosidase activity was observed. However, C18:0 DMA/C18:0 was still significantly lower in Gaucher disease patients compared to controls and the negative correlation to chitotriosidase persisted. At both time points there was no indication of an overt peroxisomal dysfunction, very long chain fatty acid, phytanate and pristanate levels being normal. In conclusion, reduced plasmalogen levels that show a significant rise following treatment and a negative correlation to total disease burden, as expressed by chitotriosidase activity, are observed in Gaucher disease.
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Membrana Eritrocítica/química , Doença de Gaucher/sangue , Plasmalogênios/análise , Adolescente , Adulto , Estudos de Casos e Controles , Criança , Pré-Escolar , Cromatografia Gasosa , Ácidos Graxos/análise , Doença de Gaucher/tratamento farmacológico , Hexosaminidases/sangue , Hexosaminidases/metabolismo , Humanos , Lactente , Pessoa de Meia-IdadeRESUMO
Gaucher disease is an autosomal recessive lysosomal storage disease that is mainly due to mutations in the GBA gene. Most of the mutant alleles described so far bear a single mutation. However, there are a few alleles bearing two or more DNA changes. It has been reported that patients homozygous for the [D409H;H255Q] double mutant allele (HGVS-approved nomenclature, p.[D448H;H294Q]) present a more severe phenotype than patients homozygous for the relatively common D409H mutation. In this study, we confirmed the detrimental cumulative effect of these two mutations at the enzymatic activity level by the heterologous expression of the single and double mutant alleles. Additionally, we found a high frequency of the [D409H;H255Q] allele in patients from the Balkans and the Adriatic area of Italy. This prompted us to perform a haplotype analysis, using five microsatellite polymorphisms close to the GBA gene, to determine the origin of this allele. The results of the 37 chromosomes analysed showed that most of them share a common haplotype and are consistent with a single origin in the Balkans and the Adriatic area of Italy for the [D409H;H255Q] allele.
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Doença de Gaucher/genética , Glucosilceramidase/genética , Análise Mutacional de DNA , Europa Oriental , Grécia , Haplótipos , Homozigoto , Humanos , Itália , Repetições de Microssatélites , MutaçãoRESUMO
We present serial clinical, magnetic resonance imaging (MRI) and neurophysiological findings of a patient with multiple sulphatase deficiency (MSD), who was first admitted at the age of 9 months, because of psychomotor retardation. MRI demonstrated extensive diffuse symmetrical high signal in the deep white matter of both cerebral hemispheres, as well as of the subcortical white matter and the brainstem, while there was additional enlargement of sulci and subdural spaces and mild atrophy. Assay of arylsulphatase A activity in white blood cell homogenates at the age of 29 months disclosed a marked deficiency of the enzyme, compatible with the diagnosis of early-infantile metachromatic leukodystrophy. During the course of a later admission, the presence of ichthyosis pointed out to the possible diagnosis of MSD; further assays of sulphatases in plasma, leukocytes as well as in cultured fibroblasts, combined with an abnormal excretion of mucopolysaccharides and sulphatides in urine confirmed the diagnosis. Molecular analysis identified a homozygous disease-causing mutation (R349W) of the SUMF1 gene. Serial neurophysiological and MRI studies demonstrated the progressive nature of the disorder (regarding both central and peripheral nervous system), correlating with the clinical deterioration (spastic quadriplegia, optic atrophy and epilepsy) with subsequent death at the age of 4 years.
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Encéfalo/patologia , Erros de Diagnóstico , Imageamento por Ressonância Magnética , Doença da Deficiência de Múltiplas Sulfatases/diagnóstico , Sulfatases/análise , Humanos , Lactente , Recém-Nascido , Leucodistrofia Metacromática/patologia , Masculino , Doença da Deficiência de Múltiplas Sulfatases/genética , Doença da Deficiência de Múltiplas Sulfatases/fisiopatologia , Mutação , Oxirredutases atuantes sobre Doadores de Grupo Enxofre , Sulfatases/genéticaRESUMO
OBJECTIVES: Long-chain polyunsaturated fatty acids (LC-PUFA) are important for fetal and infant growth and development. The effects of prematurity and perinatal asphyxia on the levels of linoleic acid (LA) and arachidonic acid (AA) in plasma and red blood cell (RBC) membranes were investigated. METHODS: Fifty-five neonates were studied: 18 full term neonates with perinatal asphyxia (group A), nine preterm neonates (group B), and 28 healthy term neonates (group C). Non-esterified and total levels of LA and AA in plasma and RBC membranes were estimated using gas chromatography within the first day of life. Malondialdehyde (MDA) levels were measured using the thiobarbituric acid (TBA) reactivity method. RESULTS: Compared to group C, statistically significant lower levels of plasma free and total AA and free LA were observed in group A, whereas statistically significant higher levels of RBC total LA and AA were observed in RBC membranes of group B. A negative correlation between MDA and LC-PUFA levels was found. CONCLUSION: Perinatal asphyxia is associated with a reduction in LC-PUFA levels, most likely as a result of increased oxidative stress. Premature infants soon after birth have higher LC-PUFA levels than term neonates, probably reflecting the overall metabolic activity and/or intrauterine transport of LC-PUFA.