A closer look at ARSA activity in a patient with metachromatic leukodystrophy.

Metachromatic leukodystrophy (MLD) is an autosomal recessive lysosomal storage disease mainly caused by a deficiency of arylsulfatase A activity. The typical clinical course of patients with the late infantile form includes a regression in motor skills with progression to dysphagia, seizures, hypotonia and death. We present a case of a 4-year-old female with rapidly progressive developmental regression with loss of motor milestones, spasticity and dysphagia. MRI showed volume loss and markedly abnormal deep white matter. Enzymatic testing in one laboratory showed arylsulfatase A activity in their normal range. However, extraction of urine showed a large increase in sulfatide excretion in a second laboratory. Measurement of arylsulfatase A in that laboratory showed a partial decrease in arylsulfatase A activity measured under typical conditions (about 37% of the normal mean). When the concentration of substrate in the assay was lowered to one quarter of that normally used, this individual had activity \u3c10% of controls. The patient was found to be homozygous for an unusual missense mutation in the arylsulfatase A gene confirming the diagnosis of MLD. This case illustrates the importance of careful biochemical and molecular testing for MLD if there is suspicion of this diagnosis

Kinetika i aktivnost leukocitne arilsulfataze A u osoba s dijagnozom cerebralne paralize

Activity and kinetics of arylsulfatase A (ASA, EC 3.1.6.8) were analyzed in leukocyte homogenates derived from patients suffering from cerebral palsy. Lower ASA activity was found in the patients\u27 leukocytes than in controls, as determined by spectrophotometry using chromogenic substrate p-nitrocatechol sulfate (p-NCS). Kinetic parameters, Km and vmax, for leukocyte ASA were determined from the dependence of initial reaction velocities on the p-NCS concentrations. A slight difference in Km values was found for leukocyte enzyme in cerebral palsy (0.26 mmol L-1) compared to the control (0.21 mmol L-1), whereas max-1 value for leukocyte ASA in disease reached only 58% of the control value. In addition, the presence of the most common mutations associated with ASA pseudodeficiency (N350S, 1524+95 A>G) and metachromatic leukodystrophy (P426L) was detected in all investigated patients. Changes in activity and kinetic parameters of leukocyte ASA in cerebral palsy are most probably related to the decrease of enzyme concentration; the detected mutations might at least partially contribute to the observed changes.Analizirane su aktivnost i kinetika arilsulfataze A (ASA, EC 3.1.6.8) u leukocitnim homogenatima osoba oboljelih od cerebralne paralize. Spektrofotometrijskim određivanjem aktivnosti ASA prema kromogenom supstratu p-nitrokatehol sulfatu (p-NCS) utvrđene su manje aktivnosti enzima u leukocitima oboljelih osoba. Kinetički parametri, Km i vmax, leukocitne ASA određeni su iz ovisnosti početne brzine reakcije o koncentraciji p-NCS. Utvrđena je manja razlika između Km vrijednosti enzima zdravih (0,21 mmol L-1) i oboljelih osobal (0.26 mmol L-1), dok je vrijednost vmax enzima iznosila 58% vrijednosti vmax enzima zdravih osoba. Također je u svih ispitanika s dijagnozom cerebralne paralize utvrđeno prisustvo najčešći mutacija povezanih s ASA pseudodeficijencijom (N350S, 1524+95 A>G) i metakromatskom leukodistrofijom (P426L). promjene aktivnosti i kinetičkih parametara leukocitne ASA u cerebralnoj parealizi najvjerojatnije su posljedica snižene koncentracije enzima; moguće je da nađene mutacije barem djelomično doprinose zapaženim promjenama

Kinetika i aktivnost leukocitne arilsulfataze A u osoba s dijagnozom cerebralne paralize

Activity and kinetics of arylsulfatase A (ASA, EC 3.1.6.8) were analyzed in leukocyte homogenates derived from patients suffering from cerebral palsy. Lower ASA activity was found in the patients\u27 leukocytes than in controls, as determined by spectrophotometry using chromogenic substrate p-nitrocatechol sulfate (p-NCS). Kinetic parameters, Km and vmax, for leukocyte ASA were determined from the dependence of initial reaction velocities on the p-NCS concentrations. A slight difference in Km values was found for leukocyte enzyme in cerebral palsy (0.26 mmol L-1) compared to the control (0.21 mmol L-1), whereas max-1 value for leukocyte ASA in disease reached only 58% of the control value. In addition, the presence of the most common mutations associated with ASA pseudodeficiency (N350S, 1524+95 A>G) and metachromatic leukodystrophy (P426L) was detected in all investigated patients. Changes in activity and kinetic parameters of leukocyte ASA in cerebral palsy are most probably related to the decrease of enzyme concentration; the detected mutations might at least partially contribute to the observed changes.Analizirane su aktivnost i kinetika arilsulfataze A (ASA, EC 3.1.6.8) u leukocitnim homogenatima osoba oboljelih od cerebralne paralize. Spektrofotometrijskim određivanjem aktivnosti ASA prema kromogenom supstratu p-nitrokatehol sulfatu (p-NCS) utvrđene su manje aktivnosti enzima u leukocitima oboljelih osoba. Kinetički parametri, Km i vmax, leukocitne ASA određeni su iz ovisnosti početne brzine reakcije o koncentraciji p-NCS. Utvrđena je manja razlika između Km vrijednosti enzima zdravih (0,21 mmol L-1) i oboljelih osobal (0.26 mmol L-1), dok je vrijednost vmax enzima iznosila 58% vrijednosti vmax enzima zdravih osoba. Također je u svih ispitanika s dijagnozom cerebralne paralize utvrđeno prisustvo najčešći mutacija povezanih s ASA pseudodeficijencijom (N350S, 1524+95 A>G) i metakromatskom leukodistrofijom (P426L). promjene aktivnosti i kinetičkih parametara leukocitne ASA u cerebralnoj parealizi najvjerojatnije su posljedica snižene koncentracije enzima; moguće je da nađene mutacije barem djelomično doprinose zapaženim promjenama

Mammalian sulfo-conjugate metabolism

Sulfoconjugates occur ubiquitously as sulfopolysaccharides, sulfolipids and sulfoproteins. A variety of sulfotransferases catalyze the sulfation process with 3'- phosphoadenosine 5'-phosphosulfate as the sulfate donor. Sulfatases that catalyze the desulfation of different sulfoconjugates are known to be deficient in a number of genetic storage disorders

Arylsulfatase a Gene Polymorphisms in Relapsing Remitting Multiple Sclerosis: Genotype-Phenotype Correlation and Estimation of Disease Progression

Arylsulfatase A (ASA) is a lysosomal enzyme involved in catabolism of cerebroside-sulfate, major lipid constituent of oligodendrocyte membranes. Various polymorphisms in ASA gene have been described, leading to different levels of enzyme deficiency. Progressive demyelination occurs in metachromatic leukodystrophy (MLD), while the condition of ASA-pseudodeficiency (ASA-PD) is suggested to contribute to complex pathogenesis of multiple sclerosis (MS). This work presents usefulness of genotype-phenotype correlation in estimation of disease severity and progression. The presence of two most common mutations associated with ASA-PD was analyzed in 56 patients with diagnosis of relapsing-remitting multiple sclerosis, by polymerase chain reaction restriction fragment length polymorphism method. In MS patients confirmed as ASA-PD mutations carriers, arylsulfatase activity was determined in leukocyte homogenates by spectrophotometry. To determine whether there is a difference between disability level and/or disease progression in patients with or without mutations we have estimated disability level using Expanded disability status scale (EDSS) and disease progression using Multiple sclerosis severity score (MSSS). Correlation of genotypes and disease progression was statistically analyzed by Kruskal-Wallis test. Patients showing higher MSSS score and found to be carriers of both analyzed ASA-PD mutations were additionally examined using conventional magnetic resonance (MR) techniques. The presence of either one or both mutations was determined in 13 patients. Lower ASA activities were observed in allMS patients carrying the mutations. Nine of the mutations carriers had mild disability (EDSS=0–4.0), 1 had moderate disability (EDSS=4.5–5.5), and 3 had severe disability (EDSS6.0). On the other hand, only 3 MS patients who were mutation carriers showed MSSS values lower than 5.000 while in other MS patients-mutation carriers the MSSS values ranged from 5.267 to 9.453. Comparison of MR findings between MS patients, mutations carrier vs. non-carrier, matched for sex, age and disease duration, showed that the total number of lesions and the number of hypointense lesions on T1-weighted images was greater in MS patient carrying the ASA-PD mutations. Our results on genotype-phenotype correlation analysis indicate a possible contribution of detected arylsulfatase A gene polymorphisms to the clinical severity of multiple sclerosis, estimated by EDSS, MSSS and MR findings. The MSSS proved to be more appropriate indicator of disease progression and should be more frequently used in clinical practice especially for comparison of disease progression in different groups of patients and identification of factors that may influence disease progression such as the presence of gene polymorphisms

The clinical features and diagnosis of Metachromatic leukodystrophy: A case series of Iranian Pediatric patients

How to Cite This Article: Jabbehdari S, Rahimian E, Jafari N, Sanii S, Khayatzadeh Kakhki S, Nejad Biglari H. The Clinical Features and Diagnosis of Metachromatic Leukodystrophy: A Case Series of Iranian Pediatric Patients. Iran J Child Neurol. Summer 2015;9(3):57-61.AbstractObjectiveMetachromatic leukodystrophy disorder (MLD) is one of the rare neurometabolicdiseases caused due to lack of saposin B and arylsulfatase A enzyme deficiency.Materials & MethodsEighteen patients diagnosed as metachromatic leukodystrophy in the NeurologyDepartment of Mofid Children’s Hospital in Tehran, Iran between 2010 and2014 were included in our study. The disorder was confirmed by clinical,EMG-NCV, arylsulfatase A enzyme checking and neuroimaging findings alongwith neurometabolic and genetic assessment from reference laboratory in Iran.We assessed age, gender, past medical history, developmental status, clinicalmanifestations, and neuroimaging findings of 18 patients with metachromaticleukodystrophy.ResultsFrom 18 patients, 80% were offspring from consanguineous marriages. A familyhistory of metachromatic leukodystrophy disease was positive for four patients.Twelve patients had late infantile form of this disorder and six patients had juvenile form. A history of tonic type seizure was positive in 20% of the patients and tonic spasm was confirmed with clinical information. Electromyographgraphy (EMG) in 96% of patients was abnormal with demyelinating sensorimotor neuropathy pattern. MRI in all patients showed the leukodystrophic pattern as arcuate fibers sparing and subcortical rim in white matter and periventricular involvement. Our diagnosis was confirmed by EMG-NCV findings with sensorimotor neuropathy pattern and the assessment of arylsulfatase A enzyme function. ConclusionMLD is an inheritance metabolic disorder, which was confirmed by theassessment of arylsulfatase A enzyme function, peripheral blood leukocyte thatassessed in a referral laboratory in Iran

Brain MRI and biological diagnosis in five Tunisians MLD patients

Metachromatic leukodystrophy (MLD) is a recessive autosomal disease which is characterized by an accumulation of sulfatides in the central and peripheral nervous system. It is due to the enzyme deficiency of the sulfatide sulfatase i.e. arylsulfatase A (ASA). we studied 5/200 cases of MLD and clearly distinguished three clinical forms. One of them presented the juvenile form; two presented the late infantile form; and two other presented the adult form. The Magnetic Resonance Imaging (MRI) of these patients showed a diffuse, bilateral and symmetrical demyelination. The biochemical diagnosis of MLD patients evidencing the low activity of ASA and sulfatide accumulation

Accumulation of lysosulfatide in the brain of arylsulfatase A-deficient mice

Lysosomal storage diseases are a group of disorders where accumulation of catabolites is manifested in the lysosomes of different cell types. In metachromatic leukodystrophy (Arylsulfatase A [EC.3.1.6.8] deficiency) storage of the glycosphingolipid sulfatide in the brain leads to demyelination, resulting in neuromotor co-ordination deficits and regression. In a mouse model for metachromatic leukodystrophy, the ASA null mutant mouse, the accumulation of sulfatide in correlation to phenotype has been thoroughly investigated. Another lipid species reported to accumulate in patients with metachromatic leukodystrophy is the sulfatide related lipid lysosulfatide. Lysosulfatide was shown to be a cytotoxic compound in cell culture experiments and thus suggested to be involved in the pathology of metachromatic leukodystrophy. In this study, we further investigated the developmental profile of lysosulfatide in the brain of ASA null mutant mice by using high performance liquid chromatography. Lysosulfatide could be detected in the brain of normal mice (ASA +/+) from 1.8 months up to 23.1 months of age. From the age of 8.8 months the lysosulfatide levels remained constant at 1 pmol/mg wet tissue. The developmental change (< 20 months) of brain lysosulfatide showed an accumulation in ASA null mutant mice at ages above one month compared to its normal counterpart (ASA +/+). Thus, the ASA null mutant mouse might be a suitable model to further investigate the role of lysosulfatide in the pathogenesis of metachromatic leukodystrophy

MOLECULAR TROJAN HORSES IN THE TREATMENT OF METACHROMATIC LEUKODYSTROPHY

Metachromatic leukodystrophy (MLD) is a lysosomal storage disease caused by a mutation in the enzyme, arylsulfatase A (ASA) and is characterized by progressive and fatal demyelination in the central and peripheral nervous systems. To date, no effective treatment exists and development is hampered by the blood-brain barrier (BBB). I have successfully generated fusion proteins using an ApoE fragment as a molecular Trojan horse fused to the full-length human ASA. These proteins were expressed in eukaryotic cells transduced with lentiviral expression vectors and have been assessed for in vitro enzymatic activity and kinetic properties. Although preliminary studies show that these constructs did not cross an in vitro BBB model under these experimental conditions, there is evidence to suggest that the ApoE fragment facilitates the uptake of fusion proteins into the endothelial cells of the BBB. Recommendations for further study are encouraged to reach the goal to find treatment for ML