Use of a multiplex ligation-dependent probe amplification method for the detection of deletions/duplications in the GBA1 gene in Gaucher disease patients

Basgalupp, Suelen Porto; Siebert, Marina; Vairo, Filippo Pinto e; Chami, Anisse Marques; Pinto, Louise Lapagesse de Camargo; Carvalho, Gérson; Schwartz, Ida Vanessa Döederlein

doi:10.1016/j.bcmd.2016.10.013

Cited by 2 publications

(3 citation statements)

References 12 publications

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“…40,41 The occurrence of deletion and/or duplication of any region of GBA1 can be specifically addressed by multiplex ligation-dependent probe amplification. 40,41…”

Section: Diagnosismentioning

confidence: 99%

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Hampered Vitamin B12 Metabolism in Gaucher Disease?

Hannibal

Siebert

Basgalupp

et al. 2017

Journal of Inborn Errors of Metabolism and Screening

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Untreated vitamin B 12 deficiency manifests clinically with hematological abnormalities and combined degeneration of the spinal cord and polyneuropathy and biochemically with elevated homocysteine (Hcy) and methylmalonic acid (MMA). Vitamin B 12 metabolism involves various cellular compartments including the lysosome, and a disruption in the lysosomal and endocytic pathways induces functional deficiency of this micronutrient. Gaucher disease (GD) is characterized by dysfunctional lysosomal metabolism brought about by mutations in the enzyme beta-glucocerebrosidase (Online Mendelian Inheritance in Man (OMIM): 606463; Enzyme Commission (EC) 3.2.1.45, gene: GBA1). In this study, we collected and examined available literature on the associations between GD, the second most prevalent lysosomal storage disorder in humans, and hampered vitamin B 12 metabolism. Results from independent cohorts of patients show elevated circulating holotranscobalamin without changes in vitamin B 12 levels in serum. Gaucher disease patients under enzyme replacement therapy present normal levels of Hcy and MMA. Although within the normal range, a significant increase in Hcy and MMA with normal serum vitamin B 12 was documented in treated GD patients with polyneuropathy versus treated GD patients without polyneuropathy. Thus, a functional deficiency of vitamin B 12 caused by disrupted lysosomal metabolism in GD is a plausible mechanism, contributing to the neurological form of the disorder but this awaits confirmation. Observational studies suggest that an assessment of vitamin B 12 status prior to the initiation of enzyme replacement therapy may shed light on the role of vitamin B 12 in the pathogenesis and progression of GD.

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“…40,41 The occurrence of deletion and/or duplication of any region of GBA1 can be specifically addressed by multiplex ligation-dependent probe amplification. 40,41…”

Section: Diagnosismentioning

confidence: 99%

“…39 In order to describe a recombinant allele, a combination of direct sequencing along with an additional method, such as Southern blot or qPCR, is strongly recommended. 40,41 The occurrence of deletion and/or duplication of any region of GBA1 can be specifically addressed by multiplex ligation-dependent probe amplification.…”

Section: Diagnosismentioning

confidence: 99%

Hampered Vitamin B12 Metabolism in Gaucher Disease?

Hannibal

Siebert

Basgalupp

et al. 2017

Journal of Inborn Errors of Metabolism and Screening

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“…The standard method for variant analysis in GD is full-gene sequencing of GBA1. Complementary techniques such as Multiplex Ligation-dependent Probe Amplification (MLPA) can be used to identify deletions or duplications of any region of this gene [6].…”

Section: Introductionmentioning

confidence: 99%

Assessment of cellular cobalamin metabolism in Gaucher disease

et al. 2020

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Background: Gaucher disease (GD) is a lysosomal disorder caused by biallelic pathogenic mutations in the GBA1 gene that encodes beta-glucosidase (GCase), and more rarely, by a deficiency in the GCase activator, saposin C. Clinically, GD manifests with heterogeneous multiorgan involvement mainly affecting hematological, hepatic and neurological axes. This disorder is divided into three types, based on the absence (type I) or presence and severity (types II and III) of involvement of the central nervous system. At the cellular level, deficiency of GBA1 disturbs lysosomal storage with buildup of glucocerebroside. The consequences of disturbed lysosomal metabolism on biochemical pathways that require lysosomal processing are unknown. Abnormal systemic markers of cobalamin (Cbl, B 12 ) metabolism have been reported in patients with GD, suggesting impairments in lysosomal handling of Cbl or in its downstream utilization events. Methods: Cultured skin fibroblasts from control humans (n = 3), from patients with GD types I (n = 1), II (n = 1) and III (n = 1) and an asymptomatic carrier of GD were examined for their GCase enzymatic activity and lysosomal compartment intactness. Control human and GD fibroblasts were cultured in growth medium with and without 500 nM hydroxocobalamin supplementation. Cellular cobalamin status was examined via determination of metabolomic markers in cell lysate (intracellular) and conditioned culture medium (extracellular). The presence of transcobalamin (TC) in whole cell lysates was examined by Western blot. Results: Cultured skin fibroblasts from GD patients exhibited reduced GCase activity compared to healthy individuals and an asymptomatic carrier of GD, demonstrating a preserved disease phenotype in this cell type. The concentrations of total homocysteine (tHcy), methylmalonic acid (MMA), cysteine (Cys) and methionine (Met) in GD cells were comparable to control levels, except in one patient with GD III. The response of these metabolomic markers to supplementation with hydroxocobalamin (HOCbl) yielded variable results. The content of transcobalamin in whole cell lysates was comparable in control human and GD patients. Conclusions: Our results indicate that cobalamin transport and cellular processing pathways are overall protected from lysosomal storage damage in GD fibroblasts. Extending these studies to hepatocytes, macrophages and plasma will shed light on cell-and compartment-specific vitamin B 12 metabolism in Gaucher disease.

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Use of a multiplex ligation-dependent probe amplification method for the detection of deletions/duplications in the GBA1 gene in Gaucher disease patients

Cited by 2 publications

References 12 publications

Hampered Vitamin B12 Metabolism in Gaucher Disease?

Hampered Vitamin B12 Metabolism in Gaucher Disease?

Assessment of cellular cobalamin metabolism in Gaucher disease

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