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1.
Cell ; 187(14): 3585-3601.e22, 2024 Jul 11.
Artículo en Inglés | MEDLINE | ID: mdl-38821050

RESUMEN

Dolichol is a lipid critical for N-glycosylation as a carrier for activated sugars and nascent oligosaccharides. It is commonly thought to be directly produced from polyprenol by the enzyme SRD5A3. Instead, we found that dolichol synthesis requires a three-step detour involving additional metabolites, where SRD5A3 catalyzes only the second reaction. The first and third steps are performed by DHRSX, whose gene resides on the pseudoautosomal regions of the X and Y chromosomes. Accordingly, we report a pseudoautosomal-recessive disease presenting as a congenital disorder of glycosylation in patients with missense variants in DHRSX (DHRSX-CDG). Of note, DHRSX has a unique dual substrate and cofactor specificity, allowing it to act as a NAD+-dependent dehydrogenase and as a NADPH-dependent reductase in two non-consecutive steps. Thus, our work reveals unexpected complexity in the terminal steps of dolichol biosynthesis. Furthermore, we provide insights into the mechanism by which dolichol metabolism defects contribute to disease.


Asunto(s)
Dolicoles , Dolicoles/metabolismo , Dolicoles/biosíntesis , Humanos , Glicosilación , 3-Oxo-5-alfa-Esteroide 4-Deshidrogenasa/metabolismo , 3-Oxo-5-alfa-Esteroide 4-Deshidrogenasa/genética , Proteínas de la Membrana/metabolismo , Proteínas de la Membrana/genética , Trastornos Congénitos de Glicosilación/metabolismo , Trastornos Congénitos de Glicosilación/genética , Masculino , Mutación Missense , Femenino
3.
J Lipid Res ; 65(6): 100557, 2024 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-38719152

RESUMEN

Dietary sphingomyelin (SM) has been reported to favorably modulate postprandial lipemia. Mechanisms underlying these beneficial effects on cardiovascular risk markers are not fully elucidated. Rodent studies showed that tritiated SM was hydrolyzed in the intestinal lumen into ceramides (Cer) and further to sphingosine (SPH) and fatty acids (FA) that were absorbed by the intestine. Our objective was to investigate the uptake and metabolism of SPH and/or tricosanoic acid (C23:0), the main FA of milk SM, as well as lipid secretion in Caco-2/TC7 cells cultured on semipermeable inserts. Mixed micelles (MM) consisting of different digested lipids and taurocholate were prepared without or with SPH, SPH and C23:0 (SPH+C23:0), or C23:0. Triglycerides (TG) were quantified in the basolateral medium, and sphingolipids were analyzed by tandem mass spectrometry. TG secretion increased 11-fold in all MM-incubated cells compared with lipid-free medium. Apical supply of SPH-enriched MM led to increased concentrations of total Cer in cells, and coaddition of C23:0 in SPH-enriched MM led to a preferential increase of C23:0 Cer and C23:0 SM. Complementary experiments using deuterated SPH demonstrated that SPH-d9 was partly converted to sphingosine-1-phosphate-d9, Cer-d9, and SM-d9 within cells incubated with SPH-enriched MM. A few Cer-d9 (2% of added SPH-d9) was recovered in the basolateral medium of (MM+SPH)-incubated cells, especially C23:0 Cer-d9 in (MM+SPH+C23:0)-enriched cells. In conclusion, present results indicate that MM enriched with (SPH+C23:0), such as found in postprandial micelles formed after milk SM ingestion, directly impacts sphingolipid endogenous metabolism in enterocytes, resulting in the secretion of TG-rich particles enriched with C23:0 Cer.


Asunto(s)
Ceramidas , Absorción Intestinal , Esfingosina , Esfingosina/análogos & derivados , Esfingosina/metabolismo , Humanos , Ceramidas/metabolismo , Células CACO-2 , Micelas , Triglicéridos/metabolismo , Marcaje Isotópico , Animales
4.
Genet Med ; 25(11): 100944, 2023 11.
Artículo en Inglés | MEDLINE | ID: mdl-37493040

RESUMEN

PURPOSE: Zellweger spectrum disorders (ZSDs) are known as autosomal recessive disorders caused by defective peroxisome biogenesis due to bi-allelic pathogenic variants in any of at least 13 different PEX genes. Here, we report 2 unrelated patients who present with an autosomal dominant ZSD. METHODS: We performed biochemical and genetic studies in blood and skin fibroblasts of the patients and demonstrated the pathogenicity of the identified PEX14 variants by functional cell studies. RESULTS: We identified 2 different single heterozygous de novo variants in the PEX14 genes of 2 patients diagnosed with ZSD. Both variants cause messenger RNA mis-splicing, leading to stable expression of similar C-terminally truncated PEX14 proteins. Functional studies indicated that the truncated PEX14 proteins lost their function in peroxisomal matrix protein import and cause increased degradation of peroxisomes, ie, pexophagy, thus exerting a dominant-negative effect on peroxisome functioning. Inhibition of pexophagy by different autophagy inhibitors or genetic knockdown of the peroxisomal autophagy receptor NBR1 resulted in restoration of peroxisomal functions in the patients' fibroblasts. CONCLUSION: Our finding of an autosomal dominant ZSD expands the genetic repertoire of ZSDs. Our study underscores that single heterozygous variants should not be ignored as possible genetic cause of diseases with an established autosomal recessive mode of inheritance.


Asunto(s)
Síndrome de Zellweger , Humanos , Alelos , Peroxisomas/genética , Peroxisomas/metabolismo , Transporte de Proteínas/fisiología , Proteínas/genética , Síndrome de Zellweger/genética
5.
Genet Med ; 24(2): 492-498, 2022 02.
Artículo en Inglés | MEDLINE | ID: mdl-34906476

RESUMEN

PURPOSE: Biallelic loss-of-function variants in ST3GAL5 cause GM3 synthase deficiency (GM3SD) responsible for Amish infantile epilepsy syndrome. All Amish patients carry the homozygous p.(Arg288Ter) variant arising from a founder effect. To date only 10 patients from 4 non-Amish families have been reported. Thus, the phenotypical spectrum of GM3SD due to other variants and other genetic backgrounds is still poorly known. METHODS: We collected clinical and molecular data from 16 non-Amish patients with pathogenic ST3GAL5 variants resulting in GM3SD. RESULTS: We identified 12 families originating from Reunion Island, Ivory Coast, Italy, and Algeria and carrying 6 ST3GAL5 variants, 5 of which were novel. Genealogical investigations and/or haplotype analyses showed that 3 of these variants were founder alleles. Glycosphingolipids quantification in patients' plasma confirmed the pathogenicity of 4 novel variants. All patients (N = 16), aged 2 to 12 years, had severe to profound intellectual disability, 14 of 16 had a hyperkinetic movement disorder, 11 of 16 had epilepsy and 9 of 16 had microcephaly. Other main features were progressive skin pigmentation anomalies, optic atrophy or pale papillae, and hearing loss. CONCLUSION: The phenotype of non-Amish patients with GM3SD is similar to the Amish infantile epilepsy syndrome, which suggests that GM3SD is associated with a narrow and severe clinical spectrum.


Asunto(s)
Epilepsia , Epilepsia/complicaciones , Epilepsia/genética , Homocigoto , Humanos , Sialiltransferasas/deficiencia , Sialiltransferasas/genética
6.
J Med Genet ; 57(1): 11-17, 2020 01.
Artículo en Inglés | MEDLINE | ID: mdl-31391289

RESUMEN

BACKGROUND: Proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitors are novel therapeutics for reducing low-density lipoprotein cholesterol (LDLc). While serious side-effects have not been observed in short-term clinical trials, there remain concerns that long-term PCSK9 inhibition may cause neurocognitive side-effects. METHODS AND RESULTS: An adult male with childhood-onset global developmental delay, cerebellar atrophy and severe hypolipidaemia underwent extensive biochemical and genetic investigations. Initial testing revealed low circulating PCSK9 levels and a common loss-of-function PCSK9 polymorphism, but these findings did not fully account for severe hypolipidaemia. Whole-exome sequencing was subsequently performed and identified two pathogenic phosphomannose mutase 2 (PMM2) variants (p.Arg141His and p.Pro69Ser) known to cause PMM2-associated congenital disorder of glycosylation (PMM2-CDG). A diagnosis of PMM2-CDG was consistent with the proband's neurological symptoms and severe hypolipidaemia. Given that PMM2-CDG is characterised by defective protein N-glycosylation and that PCSK9 is a negative regulator of LDLc, we postulated that loss of PCSK9 N-glycosylation mediates hypolipidaemia among patients with PMM2-CDG. First, in an independent cohort of patients with PMM2-CDG (N=8), we verified that circulating PCSK9 levels were significantly lower in patients than controls (p=0.0006). Second, we conducted in vitro experiments in hepatocyte-derived cells to evaluate the effects of PCSK9 N-glycosylation loss on LDL receptor (LDLR) activity. Experimental results suggest that defective PCSK9 N-glycosylation reduces the ability of circulating PCSK9 to degrade LDLR. CONCLUSION: Life-long exposure to genetically lower PCSK9 per se is unlikely to cause neurocognitive impairment. Both observational and experimental findings suggest that hypolipidaemia in PMM2-CDG may be partially mediated by loss of PCSK9 N-glycosylation and/or its regulators.


Asunto(s)
Trastornos Congénitos de Glicosilación/genética , Dislipidemias/metabolismo , Fosfotransferasas (Fosfomutasas)/deficiencia , Proproteína Convertasa 9/sangre , Receptores de LDL/metabolismo , Adulto , Estudios de Cohortes , Trastornos Congénitos de Glicosilación/complicaciones , Trastornos Congénitos de Glicosilación/metabolismo , Análisis Mutacional de ADN , Dislipidemias/etiología , Regulación de la Expresión Génica , Glicosilación , Células Hep G2 , Humanos , Mutación con Pérdida de Función , Masculino , Linaje , Fosfotransferasas (Fosfomutasas)/genética , Fosfotransferasas (Fosfomutasas)/metabolismo , Polimorfismo Genético , Proproteína Convertasa 9/genética , Proproteína Convertasa 9/metabolismo , Procesamiento Proteico-Postraduccional , Proteolisis , Receptores de LDL/genética , Secuenciación del Exoma
7.
Gut ; 69(3): 487-501, 2020 03.
Artículo en Inglés | MEDLINE | ID: mdl-31189655

RESUMEN

OBJECTIVE: To investigate whether milk polar lipids (PL) impact human intestinal lipid absorption, metabolism, microbiota and associated markers of cardiometabolic health. DESIGN: A double-blind, randomised controlled 4-week study involving 58 postmenopausal women was used to assess the chronic effects of milk PL consumption (0, 3 or 5 g-PL/day) on lipid metabolism and gut microbiota. The acute effects of milk PL on intestinal absorption and metabolism of cholesterol were assessed in a randomised controlled crossover study using tracers in ileostomy patients. RESULTS: Over 4 weeks, milk PL significantly reduced fasting and postprandial plasma concentrations of cholesterol and surrogate lipid markers of cardiovascular disease risk, including total/high-density lipoprotein-cholesterol and apolipoprotein (Apo)B/ApoA1 ratios. The highest PL dose preferentially induced a decreased number of intestine-derived chylomicron particles. Also, milk PL increased faecal loss of coprostanol, a gut-derived metabolite of cholesterol, but major bacterial populations and faecal short-chain fatty acids were not affected by milk PL, regardless of the dose. Acute ingestion of milk PL by ileostomy patients shows that milk PL decreased cholesterol absorption and increased cholesterol-ileal efflux, which can be explained by the observed co-excretion with milk sphingomyelin in the gut. CONCLUSION: The present data demonstrate for the first time in humans that milk PL can improve the cardiometabolic health by decreasing several lipid cardiovascular markers, notably through a reduced intestinal cholesterol absorption involving specific interactions in the gut, without disturbing the major bacterial phyla of gut microbiota. TRIAL REGISTRATION NUMBER: NCT02099032 and NCT02146339; Results.


Asunto(s)
Enfermedades Cardiovasculares/sangre , Metabolismo de los Lípidos/efectos de los fármacos , Lípidos/farmacología , Sobrepeso/metabolismo , Esfingomielinas/metabolismo , Animales , Apolipoproteína A-I/sangre , Apolipoproteína B-100/sangre , Colestanol/metabolismo , Colesterol/metabolismo , HDL-Colesterol/sangre , Estudios Cruzados , Suplementos Dietéticos , Método Doble Ciego , Emulsionantes/farmacología , Heces/química , Femenino , Microbioma Gastrointestinal/efectos de los fármacos , Humanos , Ileostomía , Absorción Intestinal/efectos de los fármacos , Lípidos/administración & dosificación , Lípidos/análisis , Persona de Mediana Edad , Leche/química , Posmenopausia , Factores de Riesgo
8.
Adv Exp Med Biol ; 1299: 71-80, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-33417208

RESUMEN

Zellweger syndrome disorders (ZSD) is the principal group of peroxisomal disorders characterized by a defect of peroxisome biogenesis due to mutations in one of the 13 PEX genes. The clinical spectrum is very large with a continuum from antenatal forms to adult presentation. Whereas biochemical profile in body fluids is classically used for their diagnosis, the revolution of high-throughput sequencing has extended the knowledge about these disorders. The aim of this review is to offer a large panorama on molecular basis, clinical presentation and treatment of ZSD, and to update the diagnosis strategy of these disorders in the era of next-generation sequencing (NGS).


Asunto(s)
Enfermedades del Recién Nacido , Síndrome de Zellweger , Adulto , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Recién Nacido , Enfermedades del Recién Nacido/diagnóstico , Enfermedades del Recién Nacido/genética , Enfermedades del Recién Nacido/patología , Mutación , Peroxisomas/metabolismo , Peroxisomas/patología , Síndrome de Zellweger/diagnóstico , Síndrome de Zellweger/genética , Síndrome de Zellweger/patología
9.
Clin Immunol ; 202: 33-39, 2019 05.
Artículo en Inglés | MEDLINE | ID: mdl-30946917

RESUMEN

PURPOSE: Severe combined immunodeficiency (SCID) refers to a group of genetic disorders characterized by greatly compromised cellular and humoral immunity. Children with SCID are asymptomatic at birth, but they die from infections within the first months of life if not treated. Quantification of T-cell receptor excision circles is an extremely sensitive screening method for detecting newborns who may have SCID.The goal of the DEPISTREC study was to evaluate the feasibility of nationwide newborn screening for severe T-cell lymphopenia in France as well as its economic and clinical utility. METHODS: The test universally used for neonatal screening for SCID was the quantification of TRECs on Guthrie cards. We compared a group of 190,517 babies from 48 maternities across the country who underwent newborn SCID screening with a control group of 1.4 million babies out of whom 28 were diagnosed with SCID without such screening during the course of the study. RESULTS: Within the screening group, 62 babies were found to be lymphopenic, including three with SCID. The cost of screening ranged from 4.7€ to €8.15 per newborn. The average 18-month cost was €257,574 vs €204,697 in the control group. CONCLUSIONS: In this large-scale study, we demonstrate that routine SCID screening is feasible and effective. This screening offers the additional benefit of aiding in the diagnosis of non-SCID lymphopenia. Economic evaluation allowed us to calculate the cost per test. Newborn screening may also prevent death by SCID before any curative treatment can be administered. The difference in cost between screened and control children could not be ascertained because of the very low numbers and death of one of the children tested.


Asunto(s)
Linfopenia/diagnóstico , Tamizaje Neonatal/economía , Inmunodeficiencia Combinada Grave/diagnóstico , Costos y Análisis de Costo , Pruebas con Sangre Seca/economía , Femenino , Francia , Humanos , Lactante , Recién Nacido , Recuento de Linfocitos , Linfopenia/economía , Masculino , Receptores de Antígenos de Linfocitos T/inmunología , Inmunodeficiencia Combinada Grave/economía , Linfocitos T/inmunología
10.
Am J Hum Genet ; 98(2): 310-21, 2016 Feb 04.
Artículo en Inglés | MEDLINE | ID: mdl-26833332

RESUMEN

Disorders of Golgi homeostasis form an emerging group of genetic defects. The highly heterogeneous clinical spectrum is not explained by our current understanding of the underlying cell-biological processes in the Golgi. Therefore, uncovering genetic defects and annotating gene function are challenging. Exome sequencing in a family with three siblings affected by abnormal Golgi glycosylation revealed a homozygous missense mutation, c.92T>C (p.Leu31Ser), in coiled-coil domain containing 115 (CCDC115), the function of which is unknown. The same mutation was identified in three unrelated families, and in one family it was compound heterozygous in combination with a heterozygous deletion of CCDC115. An additional homozygous missense mutation, c.31G>T (p.Asp11Tyr), was found in a family with two affected siblings. All individuals displayed a storage-disease-like phenotype involving hepatosplenomegaly, which regressed with age, highly elevated bone-derived alkaline phosphatase, elevated aminotransferases, and elevated cholesterol, in combination with abnormal copper metabolism and neurological symptoms. Two individuals died of liver failure, and one individual was successfully treated by liver transplantation. Abnormal N- and mucin type O-glycosylation was found on serum proteins, and reduced metabolic labeling of sialic acids was found in fibroblasts, which was restored after complementation with wild-type CCDC115. PSI-BLAST homology detection revealed reciprocal homology with Vma22p, the yeast V-ATPase assembly factor located in the endoplasmic reticulum (ER). Human CCDC115 mainly localized to the ERGIC and to COPI vesicles, but not to the ER. These data, in combination with the phenotypic spectrum, which is distinct from that associated with defects in V-ATPase core subunits, suggest a more general role for CCDC115 in Golgi trafficking. Our study reveals CCDC115 deficiency as a disorder of Golgi homeostasis that can be readily identified via screening for abnormal glycosylation in plasma.


Asunto(s)
Aparato de Golgi/genética , Homeostasis , Proteínas del Tejido Nervioso/deficiencia , Proteínas del Tejido Nervioso/genética , Secuencia de Aminoácidos , Niño , Preescolar , Clonación Molecular , Retículo Endoplásmico/metabolismo , Exoma , Femenino , Fibroblastos/citología , Glicosilación , Aparato de Golgi/metabolismo , Células HeLa , Heterocigoto , Humanos , Lactante , Masculino , Datos de Secuencia Molecular , Linaje , Fenotipo , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción
13.
J Clin Immunol ; 38(7): 778-786, 2018 10.
Artículo en Inglés | MEDLINE | ID: mdl-30251145

RESUMEN

Severe combined immunodeficiency (SCID) is characterized by a major T cell deficiency. Infants with SCID are asymptomatic at birth but die from infections in the first year of life if not treated. Survival rates are better for early treatment. SCID therefore meets criteria for newborn screening (NBS). T cell receptor excision circle (TREC) quantification is a reliable marker of T cell deficiency and can be performed using Guthrie cards. The DEPISTREC project was designed to study the feasibility, clinical utility, and cost-effectiveness of generalized SCID screening in France. About 200,000 babies from all over the country were screened at birth with a commercial kit. We determined assay performance and proposed a cutoff for classification of results. Our findings suggest that, given clearly established validation rules and decision-making procedures, the TREC assay is a suitably specific and sensitive method for high-throughput SCID screening. Clinical Trials: NCT02244450.


Asunto(s)
Inmunodeficiencia Combinada Grave/diagnóstico , Bioensayo , Biomarcadores , Toma de Decisiones Clínicas , Análisis Costo-Beneficio , Manejo de la Enfermedad , Francia/epidemiología , Humanos , Lactante , Recién Nacido , Tamizaje Neonatal , Vigilancia en Salud Pública , Juego de Reactivos para Diagnóstico , Receptores de Antígenos de Linfocitos T/metabolismo , Reproducibilidad de los Resultados , Inmunodeficiencia Combinada Grave/epidemiología , Inmunodeficiencia Combinada Grave/inmunología , Linfocitos T/inmunología , Linfocitos T/metabolismo
15.
Mol Genet Metab ; 123(4): 463-471, 2018 04.
Artículo en Inglés | MEDLINE | ID: mdl-29478817

RESUMEN

Creatine transporter is currently the focus of renewed interest with emerging roles in brain neurotransmission and physiology, and the bioenergetics of cancer metastases. We here report on amendments of a standard creatine uptake assay which might help clinical chemistry laboratories to extend their current range of measurements of creatine and metabolites in body fluids to functional enzyme explorations. In this respect, short incubation times and the use of a stable-isotope-labeled substrate (D3-creatine) preceded by a creatine wash-out step from cultured fibroblast cells by removal of fetal bovine serum (rich in creatine) from the incubation medium are recommended. Together, these measures decreased, by a first order of magnitude, creatine concentrations in the incubation medium at the start of creatine-uptake studies and allowed to functionally discriminate between 4 hemizygous male and 4 heterozygous female patients with X-linked SLC6A8 deficiency, and between this cohort of eight patients and controls. The functional assay corroborated genetic diagnosis of SLC6A8 deficiency. Gene anomalies in our small cohort included splicing site (c.912G > A [p.Ile260_Gln304del], c.778-2A > G and c.1495 + 2 T > G), substitution (c.407C > T) [p.Ala136Val] and deletion (c.635_636delAG [p.Glu212Valfs*84] and c.1324delC [p.Gln442Lysfs*21]) variants with reduced creatine transporter function validating their pathogenicity, including that of a previously unreported c.1324delC variant. The present assay adaptations provide an easy, reliable and discriminative manner for exploring creatine transporter activity and disease variations. It might apply to drug testing or other evaluations in the genetic and metabolic horizons covered by the emerging functions of creatine and its transporter, in a way, however, requiring and completed by additional studies on female patients and blood-brain barrier permeability properties of selected compounds. As a whole, the proposed assay of creatine transporter positively adds to currently existing measurements of this transporter activity, and determining on a large scale the extent of its exact suitability to detect female patients should condition in the future its transfer in clinical practice.


Asunto(s)
Encefalopatías Metabólicas Innatas/metabolismo , Creatina/deficiencia , Fibroblastos/metabolismo , Discapacidad Intelectual Ligada al Cromosoma X/metabolismo , Mutación , Proteínas del Tejido Nervioso/deficiencia , Proteínas del Tejido Nervioso/genética , Proteínas de Transporte de Neurotransmisores en la Membrana Plasmática/deficiencia , Adolescente , Encefalopatías Metabólicas Innatas/genética , Encefalopatías Metabólicas Innatas/patología , Estudios de Casos y Controles , Niño , Preescolar , Estudios de Cohortes , Creatina/genética , Creatina/metabolismo , Femenino , Fibroblastos/patología , Estudios de Seguimiento , Humanos , Lactante , Masculino , Discapacidad Intelectual Ligada al Cromosoma X/genética , Discapacidad Intelectual Ligada al Cromosoma X/patología , Proteínas de Transporte de Neurotransmisores en la Membrana Plasmática/genética , Proteínas de Transporte de Neurotransmisores en la Membrana Plasmática/metabolismo , Pronóstico
16.
Electrophoresis ; 39(24): 3123-3132, 2018 12.
Artículo en Inglés | MEDLINE | ID: mdl-29869806

RESUMEN

Congenital disorders of glycosylation (CDG) are rare autosomal genetic diseases affecting the glycosylation of proteins and lipids. Since CDG-related clinical symptoms are classically extremely variable and nonspecific, a combination of electrophoretic, mass spectrometric, and gene sequencing techniques is often mandatory for obtaining a definitive CDG diagnosis, as well as identifying causative gene mutations and deciphering the underlying biochemical mechanisms. Here, we illustrate the potential of integrating data from capillary electrophoresis of transferrin, two-dimensional electrophoresis of N- and O-glycoproteins, mass spectrometry analyses of total serum N-linked glycans and mucin core1 O-glycosylated apolipoprotein C-III for the determination of various culprit CDG gene mutations. "Step-by-step" diagnosis pathways of four particular and new CDG cases, including MGAT2-CDG, ATP6V0A2-CDG, SLC35A2-CDG, and SLC35A3-CDG, are described as illustrative examples.


Asunto(s)
Trastornos Congénitos de Glicosilación , Electroforesis/métodos , Espectrometría de Masas/métodos , Análisis de Secuencia de ADN/métodos , Adolescente , Niño , Preescolar , Trastornos Congénitos de Glicosilación/sangre , Trastornos Congénitos de Glicosilación/diagnóstico , Femenino , Glicómica , Glicoproteínas/sangre , Glicoproteínas/química , Humanos , Lactante , Masculino , Polisacáridos/análisis , Polisacáridos/química
17.
J Inherit Metab Dis ; 41(3): 457-477, 2018 05.
Artículo en Inglés | MEDLINE | ID: mdl-29556840

RESUMEN

Tandem mass spectrometry (MS/MS) is a highly sensitive and specific technique. Thanks to the development of triple quadrupole analyzers, it is becoming more widely used in laboratories working in the field of inborn errors of metabolism. We review here the state of the art of this technique applied to the diagnosis of lysosomal storage disorders (LSDs) and how MS/MS has changed the diagnostic rationale in recent years. This fine technology brings more sensitive, specific, and reliable methods than the previous biochemical ones for the analysis of urinary glycosaminoglycans, oligosaccharides, and sialic acid. In sphingolipidoses, the quantification of urinary sphingolipids (globotriaosylceramide, sulfatides) is possible. The measurement of new plasmatic biomarkers such as oxysterols, bile acids, and lysosphingolipids allows the screening of many sphingolipidoses and related disorders (Niemann-Pick type C), replacing tedious biochemical techniques. Applied to amniotic fluid, a more reliable prenatal diagnosis or screening of LSDs is now available for fetuses presenting with antenatal manifestations. Applied to enzyme measurements, it allows high throughput assays for the screening of large populations, even newborn screening. The advent of this new method can modify the diagnostic rationale behind LSDs.


Asunto(s)
Enfermedades por Almacenamiento Lisosomal/diagnóstico , Espectrometría de Masas en Tándem , Biomarcadores/análisis , Femenino , Humanos , Recién Nacido , Enfermedades por Almacenamiento Lisosomal/epidemiología , Tamizaje Neonatal/métodos , Valor Predictivo de las Pruebas , Embarazo , Diagnóstico Prenatal/métodos , Espectrometría de Masas en Tándem/métodos
18.
Ann Gen Psychiatry ; 16: 19, 2017.
Artículo en Inglés | MEDLINE | ID: mdl-28392828

RESUMEN

BACKGROUND: Wilson's disease (WD) is a rare autosomal-recessive, inherited disorder caused by a mutation in the copper-transporting gene ATP7B affecting the liver and nervous system. About 30% of patients with WD may initially present with psychiatric symptoms, and diagnosis can be difficult to establish. The objectives of the present preliminary study were [1] to evaluate the relevance of serum copper (Cu) and ceruloplasmin (Cp) measures in hospitalized patients with psychiatric disorders; and [2] to identify possible mutations in the ATP7B gene in patients with abnormal biological copper profile. METHODS: All psychiatric patients who participated in this study were hospitalized in Saint-Jean de Dieu Hospital (Lyon, France). Cp was measured by immunoturbidimetry and serum Cu by inductively coupled plasma-optical emission spectrometry. When Cp and serum Cu levels were inferior to, respectively, 0.18 g/L and 0.88 mg/L in combination with atypical psychiatric presentations, complete clinical examinations were performed by multidisciplinary physicians specialized in WD. In addition, mutation detection in the ATP7B gene was performed. RESULTS: A total of 269 patients completed the study. (1) 51 cases (19%) showed both decreased Cp and Cu concentrations. (2) Molecular genetic tests were performed in 29 patients, and one ATP7B mutation (heterozygous state) was found in four patients. We identified three different missense mutations: p.His1069Gln, c.3207C>A (exon 14), p.Pro1379Ser, c.4135C>T (exon 21) and p.Thr1434Met, c.4301C>T (exon 21). No pathogenic mutation on either ATP7B allele was detected. CONCLUSION: Results of Cp and/or serum Cu concentrations below the normal limits are common in patients with psychiatric disorders and nonrelevant and/or informative for the WD diagnosis. WD diagnosis is based on a combination of clinical and biological arguments. Psychiatric patients with suspicion of WD should be evaluated in a reference center. Trial registration CPP Lyon Sud-Est IVNo 10/044, CNIL No DR-2011-470, Afssaps No B100832-40 and CCTIRS No 10.612 bis, registered 8 June 2010.

19.
J Inherit Metab Dis ; 39(5): 611-624, 2016 09.
Artículo en Inglés | MEDLINE | ID: mdl-27393412

RESUMEN

Inborn errors of metabolism (IEMs) that present with abnormal imaging findings in the second half of pregnancy are mainly lysosomal storage disorders (LSDs), cholesterol synthesis disorders (CSDs), glycogen storage disorder type IV (GSD IV), peroxisomal disorders, mitochondrial fatty acid oxidation defects (FAODs), organic acidurias, aminoacidopathies, congenital disorders of glycosylation (CDGs), and transaldolase deficiency. Their biological investigation requires fetal material. The supernatant of amniotic fluid (AF) is useful for the analysis of mucopolysaccharides, oligosaccharides, sialic acid, lysosphingolipids and some enzyme activities for LSDs, 7- and 8-dehydrocholesterol, desmosterol and lathosterol for CSDs, acylcarnitines for FAODs, organic acids for organic acidurias, and polyols for transaldolase deficiency. Cultured AF or fetal cells allow the measurement of enzyme activities for most IEMs, whole-cell assays, or metabolite measurements. The cultured cells or tissue samples taken after fetal death can be used for metabolic profiling, enzyme activities, and DNA extraction. Fetal blood can also be helpful. The identification of vacuolated cells orients toward an LSD, and plasma is useful for diagnosing peroxisomal disorders, FAODs, CSDs, some LSDs, and possibly CDGs and aminoacidopathies. We investigated AF of 1700 pregnancies after exclusion of frequent etiologies of nonimmune hydrops fetalis and identified 108 fetuses affected with LSDs (6.3 %), 29 of them with mucopolysaccharidosis type VII (MPS VII), and six with GSD IV (0.3 %). In the AF of 873 pregnancies, investigated because of intrauterine growth restriction and/or abnormal genitalia, we diagnosed 32 fetuses affected with Smith-Lemli-Opitz syndrome (3.7 %).


Asunto(s)
Enfermedades Metabólicas/diagnóstico , Errores Innatos del Metabolismo/diagnóstico , Líquido Amniótico/química , Animales , Femenino , Feto/anomalías , Humanos , Enfermedades Metabólicas/genética , Redes y Vías Metabólicas/genética , Errores Innatos del Metabolismo/genética , Embarazo , Diagnóstico Prenatal/métodos
20.
Brain ; 138(Pt 2): 336-55, 2015 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-25567323

RESUMEN

Severe progressive neurological paediatric disease mucopolysaccharidosis III type C is caused by mutations in the HGSNAT gene leading to deficiency of acetyl-CoA: α-glucosaminide N-acetyltransferase involved in the lysosomal catabolism of heparan sulphate. To understand the pathophysiology of the disease we generated a mouse model of mucopolysaccharidosis III type C by germline inactivation of the Hgsnat gene. At 6-8 months mice showed hyperactivity, and reduced anxiety. Cognitive memory decline was detected at 10 months and at 12-13 months mice showed signs of unbalanced hesitant walk and urinary retention. Lysosomal accumulation of heparan sulphate was observed in hepatocytes, splenic sinus endothelium, cerebral microglia, liver Kupffer cells, fibroblasts and pericytes. Starting from 5 months, brain neurons showed enlarged, structurally abnormal mitochondria, impaired mitochondrial energy metabolism, and storage of densely packed autofluorescent material, gangliosides, lysozyme, phosphorylated tau, and amyloid-ß. Taken together, our data demonstrate for the first time that deficiency of acetyl-CoA: α-glucosaminide N-acetyltransferase causes lysosomal accumulation of heparan sulphate in microglial cells followed by their activation and cytokine release. They also show mitochondrial dysfunction in the neurons and neuronal loss explaining why mucopolysaccharidosis III type C manifests primarily as a neurodegenerative disease.


Asunto(s)
Enfermedades Mitocondriales/patología , Mucopolisacaridosis III/patología , Neuritis/patología , Enfermedades Neurodegenerativas/patología , Acetiltransferasas/deficiencia , Acetiltransferasas/genética , Animales , Conducta Animal , Metabolismo Energético/fisiología , Gangliósidos/metabolismo , Glicosaminoglicanos/metabolismo , Ratones , Ratones Endogámicos C57BL , Proteínas Asociadas a Microtúbulos/metabolismo , Enfermedades Mitocondriales/etiología , Mucopolisacaridosis III/complicaciones , Mucopolisacaridosis III/psicología , Neuritis/etiología , Enfermedades Neurodegenerativas/etiología , Enfermedades Neurodegenerativas/psicología , Examen Neurológico , Deficiencias en la Proteostasis/patología
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