Oral ribose supplementation in dystroglycanopathy: A single case study.

Three forms of muscular dystrophy-dystroglycanopathies are linked to the ribitol pathway. These include mutations in the isoprenoid synthase domain-containing protein (ISPD), fukutin-related protein (FKRP), and fukutin (FKTN) genes. The aforementioned enzymes are required for generation of the ribitol phosphate linkage in the O-glycan of alpha-dystroglycan. Mild cases of dystroglycanopathy present with slowly progressive muscle weakness, while in severe cases the eyes and brain are also involved. Previous research showed that ribose increased the intracellular concentrations of cytidine diphosphate-ribitol (CDP-ribitol) and had a therapeutic effect. Here, we report the safety and effects of oral ribose supplementation during 6 months in a patient with limb girdle muscular dystrophy type 2I (LGMD2I) due to a homozygous FKRP mutation. Ribose was well tolerated in doses of 9 g or 18 g/day. Supplementation with 18 g of ribose resulted in a decrease of creatine kinase levels of 70%. Moreover, metabolomics showed a significant increase in CDP-ribitol levels with 18 g of ribose supplementation (p < 0.001). Although objective improvement in clinical and patient-reported outcome measures was not observed, the patient reported subjective improvement of muscle strength, fatigue, and pain. This case study indicates that ribose supplementation in patients with dystroglycanopathy is safe and highlights the importance for future studies regarding its potential effects.

Examining the Distribution and Impact of Single-Nucleotide Polymorphisms in the Capsular Locus of Streptococcus pneumoniae Serotype 19A.

Streptococcus pneumoniae serotype 19A prevalence has increased after the implementation of the PCV7 and PCV10 vaccines. In this study, we have provided, with high accuracy, the genetic diversity of the 19A serotype in a cohort of Dutch invasive pneumococcal disease patients and asymptomatic carriers obtained in the period from 2004 to 2016. The whole genomes of the 338 pneumococcal isolates in this cohort were sequenced and their capsule (cps) loci compared to examine their diversity and determine the impact on the production of capsular polysaccharide (CPS) sugar precursors and CPS shedding. We discovered 79 types with a unique cps locus sequence. Most variation was observed in the rmlB and rmlD genes of the TDP-Rha synthesis pathway and in the wzg gene, which is of unknown function. Interestingly, gene variation in the cps locus was conserved in multiple alleles. Using RmlB and RmlD protein models, we predict that enzymatic function is not affected by the single-nucleotide polymorphisms as identified. To determine if RmlB and RmlD function was affected, we analyzed nucleotide sugar levels using ultrahigh-performance liquid chromatography-mass spectrometry (UHPLC-MS). CPS precursors differed between 19A cps locus subtypes, including TDP-Rha, but no clear correlation was observed. Also, significant differences in multiple nucleotide sugar levels were observed between phylogenetically branched groups. Because of indications of a role for Wzg in capsule shedding, we analyzed if this was affected. No clear indication of a direct role in shedding was found. We thus describe genotypic variety in rmlB, rmlD, and wzg in serotype 19A in the Netherlands, for which we have not discovered an associated phenotype.

Mimicking Behçet's disease: GM-CSF gain of function mutation in a family suffering from a Behçet's disease-like disorder marked by extreme pathergy.

Behçet's disease (BD) is an inflammatory disease mainly affecting men along the ancient Silk Route. In the present study we describe a Dutch family suffering from BD-like disease with extreme pathergic responses, but without systemic inflammation. Genetic assessment revealed a combination of the human leukocyte antigen (HLA)-B*51 risk-allele together with a rare heterozygous variant in the CSF2 gene (c.130A>C, p.N44H) encoding for granulocyte-macrophage colony-stimulating factor (GM-CSF) found by whole exome sequencing. We utilized an over-expression vector system in a human hepatocyte cell line to produce the aberrant variant of GM-CSF. Biological activity of the protein was measured by signal transducer and activator of transcription 5 (STAT-5) phosphorylation, a downstream molecule of the GM-CSF receptor, in wild-type peripheral mononuclear cells (PBMCs) using flow cytometry. Increased STAT-5 phosphorylation was observed in response to mutated GM-CSF when compared to the wild-type or recombinant protein. CSF2 p.N44H results in disruption of one of the protein's two N-glycosylation sites. Enzymatically deglycosylated wild-type GM-CSF also enhanced STAT-5 phosphorylation. The patient responded well to anti-tumor necrosis factor (TNF)-α treatment, which may be linked to the capacity of TNF-α to induce GM-CSF in phorbol 12-myristate 13-acetate (PMA)-treated PBMCs, while GM-CSF itself only induced dose-dependent interleukin (IL)-1Ra production. The identified CSF2 pathway could provide novel insights into the pathergic response of BD-like disease and offer new opportunities for personalized treatment.

Prosthetic gait of unilateral lower-limb amputees with current and novel prostheses: A pilot study.

BACKGROUND: Novel lower-limb prostheses aim to improve the quality of locomotion of individuals with an amputation. This study evaluates the biomechanics of a novel bionic foot during walking. METHODS: Able-bodied individuals (n = 7) and individuals with a transfemoral (n = 6) or transtibial amputation (n = 6) were included. Able-bodied individuals conducted one experimental trial, whereas individuals with transtibial and transfemoral amputations conducted a familiarization (with current prosthesis) and two experimental trials using a passive and bionic prosthesis. Each trial consisted of 3 bouts of 2 min of treadmill walking at different speeds. Biomechanical data were gathered using a force platform and motion capture system and analysed using Statistical Parametric Mapping and (non)-parametric tests. FINDINGS: Conventional prosthetic feet alter gait patterns and induce locomotion difficulties. While walking at a normal speed with the passive prosthesis, transtibial amputees display reduced maximum heel forces, increased ankle and trunk angular velocities at midstance, and increased knee angle during stance and swing phases on their effected side (P ≤ 0.026). Improved lower-limb kinematics was demonstrated during slow and normal speed walking with the bionic prosthesis; however, dynamic trunk stability was negatively impacted during this condition. The bionic prosthesis did not benefit transfemoral amputees at any walking speed. INTERPRETATION: Transtibial amputees can better approximate typical movement patterns at slow and normal walking speeds using the novel bionic prosthesis; however the same benefit was not observed in transfemoral amputees.

Dynamic Performance of an Oil Starved Squeeze Film Damper Combined With a Cylindrical Roller Bearing.

Squeeze film dampers (SFDs) are widely used to dissipate mechanical energy caused by rotor vibrations as well as to improve overall stability of the rotor system. Especially turbomachine rotors, supported on little damped rolling element bearings (REBs), are primarily sensitive to unbalance excitation and thus high amplitude vibrations. To ensure safe operation, potential failure modes, such as an oil starved damper state, need to be well examined prior to the introduction in the ultimate industrial application. Hence, the aim of this research project is to evaluate the performance of the rotor support for a complete oil starvation of the SFD. An academic rotor dynamic test bench has been developed and briefly presented. Experimental testing has been conducted for two static radial load cases resembling the full load and idle condition of a certain turbomachine. Evidently, the measurement results exposed severe vibration problems. Even a split first whirl mode arises due to a pronounced anisotropic bearing stiffness. Moreover, for the least radially loaded bearing, highly nonlinear behavior emerged at elevated unbalance excitation. Consequently, the rollers start to rattle which will have a negative effect on the overall bearing lifetime. To explain the nature of the nonlinear behavior, advanced quasi-static bearing simulations are exploited. A number of possible solutions are proposed in order to help mitigate the vibration issues.

Relative quantification of plasma N-glycans in type II congenital disorder of glycosylation patients by mass spectrometry.

BACKGROUND: Type II Congenital Disorders of Glycosylation (CDG-II) are a group of diseases with challenging diagnostics characterized by defects in the processing of glycans in the Golgi apparatus. Mass Spectrometry (MS) has been a valuable tool in the definition of CDG-II subtypes. While some CDG-II subtypes are associated with specific N-glycan structures, others only produce changes in relative levels, reinforcing the demand for quantification methods. METHODS: Plasma samples from control individuals were pooled, derivatized with deuterated iodomethane (I-CD3), and used as internal standards for controls and patients whose glycans were derivatized with iodomethane (I-CH3), followed by MALDI MS, LC-MS and -MS/MS analyses. RESULTS: Total N-glycans from fifteen CDG-II patients were evaluated, and 4 cases with molecular diagnosis were considered in detail: 2ATP6V0A2-CDG siblings, and 2 MAN1B1-CDG patients, one of them carrying a previously undescribed p.Gly536Val mutation. CONCLUSIONS: Our methodology offers a feasible alternative to the current methods for CDG-II diagnosis by MS, which quantify glycan structures as fractions of the total summed signal across a mass spectrum, a strategy that lowers the variability of minor components. Moreover, given its sensitivity for less concentrated yet biologically relevant structures, it might assist the uncovering of novel diagnostic glycans in other CDG-II subtypes.

A homozygous DPM3 mutation in a patient with alpha-dystroglycan-related limb girdle muscular dystrophy.

Defects of O-linked glycosylation of alpha-dystroglycan cause a wide spectrum of muscular dystrophies ranging from severe congenital muscular dystrophy associated with abnormal brain and eye development to mild limb girdle muscular dystrophy. We report a female patient who developed isolated pelvic girdle muscle weakness and wasting, which became symptomatic at age 42. Exome sequencing uncovered a homozygous c.131T > G (p.Leu44Pro) substitution in DPM3, encoding dolichol-P-mannose (DPM) synthase subunit 3, leading to a 50% reduction of enzymatic activity. Decreased availability of DPM as an essential donor substrate for protein O-mannosyltransferase (POMT) 1 and 2 explains defective skeletal muscle alpha-dystroglycan O-glycosylation. Our findings show that DPM3 mutations may lead to an isolated and mild limb girdle muscular dystrophy phenotype without cardiomyopathy.

PGM1 deficiency: Substrate use during exercise and effect of treatment with galactose.

Mutations in PGM1 (phosphoglucomutase 1) cause Glycogen Storage Disease type XIV, which is also a congenital disorder of protein N-glycosylation. It presents throughout life as myopathy with additional systemic symptoms. We report the effect of oral galactose treatment during five months in a patient with biochemically and genetically confirmed PGM1 deficiency. The 12-minute-walking distance increased by 225 m (65%) and transferrin glycosylation was restored to near-normal levels. The exercise assessments showed a severe exercise intolerance due to a block in skeletal muscle glycogenolytic capacity and that galactose treatment tended to normalize skeletal muscle substrate use from fat to carbohydrates during exercise.

Design and energetic evaluation of a prosthetic knee joint actuator with a lockable parallel spring.

There are disadvantages to existing damping knee prostheses which cause an asymmetric gait and higher metabolic cost during level walking compared to non-amputees. Most existing active knee prostheses which could benefit the amputees use a significant amount of energy and require a considerable motor. In this work, a novel semi-active actuator with a lockable parallel spring for a prosthetic knee joint has been developed and tested. This actuator is able to provide an approximation of the behavior of a healthy knee during most of the gait cycle of level walking. This actuator is expanded with a series-elastic actuator to mimic the full gait cycle and enable its use in other functional tasks like stair climbing and sit-to-stance. The proposed novel actuator reduces the energy consumption for the same trajectory with respect to a compliant or directly-driven prosthetic active knee joint and improves the approximation of healthy knee behavior during level walking compared to passive or variable damping knee prostheses.

Hypoglycosylation is a common finding in antithrombin deficiency in the absence of a SERPINC1 gene defect.

UNLABELLED: Essentials We investigated the molecular base of antithrombin deficiency in cases without SERPINC1 defects. 27% of cases presented hypoglycosylation, transient in 62% and not restricted to antithrombin. Variations in genes involved in N-glycosylation underline this phenotype. These results support a new form of thrombophilia. Click here to listen to Dr Huntington's perspective on thrombin inhibition by the serpins SUMMARY: Background Since the discovery of antithrombin deficiency, 50 years ago, few new thrombophilic defects have been identified, all with weaker risk of thrombosis than antithrombin deficiency. Objective To identify new thrombophilic mechanisms. Patients/methods We studied 30 patients with antithrombin deficiency but no defects in the gene encoding this key anticoagulant (SERPINC1). Results A high proportion of these patients (8/30: 27%) had increased hypoglycosylated forms of antithrombin. All N-glycoproteins tested in these patients (α1-antitrypsin, FXI and transferrin) had electrophoretic, HPLC and Q-TOF patterns indistinguishable from those of the congenital disorders of glycosylation (rare recessive multisystem disorders). However, all except one had no mental disability. Moreover, intermittent antithrombin deficiency and hypoglycosylation was recorded in five out of these eight patients, all associated with moderate alcohol intake. Genetic analysis, including whole exome sequencing, revealed mutations in different genes involved in the N-glycosylation pathway. Conclusions Our study provides substantial and novel mechanistic insights into two disease processes, with potential implications for diagnosis and clinical care. An aberrant N-glycosylation causing a recessive or transient antithrombin deficiency is a new form of thrombophilia. Our data suggest that congenital disorders of glycosylation are probably underestimated, especially in cases with thrombosis as the main or only clinical manifestation.

Ankle-Knee prosthesis with powered ankle and energy transfer for CYBERLEGs α-prototype.

Restoring natural walking for amputees has been increasingly investigated because of demographic evolution, leading to increased number of amputations, and increasing demand for independence. The energetic disadvantages of passive pros-theses are clear, and active prostheses are limited in autonomy. This paper presents the simulation, design and development of an actuated knee-ankle prosthesis based on a variable stiffness actuator with energy transfer from the knee to the ankle. This approach allows a good approximation of the joint torques and the kinematics of the human gait cycle while maintaining compliant joints and reducing energy consumption during level walking. This first prototype consists of a passive knee and an active ankle, which are energetically coupled to reduce the power consumption.

Socio-emotional Problems in Children with CDG.

BACKGROUND: Congenital disorders of glycosylation (CDG) form a group of inherited metabolic diseases. Although the clinical presentation shows extreme variability, the nervous system is frequently affected. Several parents of our patients diagnosed with CDG reported behavioral problems, including mood swings, depressive behavior, and anxiety. This raised the question whether patients with CDG have an increased risk for socio-emotional problems. METHODS: We evaluated 18 children with confirmed CDG. The Child Behavior Checklist (CBCL) was used to screen for socio-emotional problems. To determine the disease progression and severity in CDG, the Nijmegen Paediatric CDG Rating Scale (NPCRS) was used. RESULTS were compared to "norm scores" and to children with mitochondrial disorders and children with other chronic metabolic disorders with multisystem involvement. RESULTS: RESULTS showed a high prevalence of socio-emotional problems in children with CDG. Mean total scores, scores on withdrawn/depressed behavior, social problems, and somatic complaints were significantly increased. More than two thirds of our CDG patients have abnormal scores on CBCL. The mean score on social problems was significantly higher compared to our two control groups of patients with other chronic metabolic disorders. CONCLUSIONS: Patients with CDG have an increased risk of developing socio-emotional problems. A standard screening for psychological problems is recommended for the early detection of psychological problems in CDG patients.

Thrombotic complications in patients with PMM2-CDG.

Many proteins regulating coagulation, including factor IX, factor XI, Antithrombin-III, Protein C and Protein S are deficient or decreased in activity in congenital disorders of glycosylation (CDG). Because of the imbalance of coagulation and anticoagulation factors, some patients develop acute vascular events, such as thrombosis. Identifying patients with increased risk for thrombotic events could prevent serious complications and even mortality. We performed a systematic review on patients diagnosed with the most common CDG form; PMM2-CDG, reported between 1990 and 2012 in medical literature. We also evaluated our PMM2-CDG patient-cohort of 15 patients. In total, based on the availability of comprehensive clinical descriptions, 100 patients were included in the study. Patients with and without thrombotic events were compared based on the alterations of the following glycosylated coagulation and anticoagulation factors: Antithrombin-III, Protein C, Protein S, factors IX and XI. We also assessed the global hemostasis, family history and provoking events. In the group of 100 PMM2-CDG patients 14 had suffered a venous or arterial thrombotic event. Low activity of several anticoagulation factors correlated with thrombotic events. Relatively high factor IX and XI activities were not associated with thrombosis. Prolonged PT and aPTT did not seem to protect against thrombosis in patients. Surgical procedures were frequently associated with thrombotic events. Based on the association of thrombosis and surgery in PMM2-CDG we advise to avoid elective surgical procedures in PMM2-CDG patients. Easily preventable risk factors like immobility should be treated with regular physiotherapy. We suggest a yearly follow-up for Antithrombin-III and Protein C levels and parent education for early thrombotic signs in CDG.

Acid sphingomyelinase (Asm) deficiency patients in The Netherlands and Belgium: disease spectrum and natural course in attenuated patients.

Niemann-Pick disease (NPD) is a neurovisceral lysosomal storage disorder caused by acid sphingomyelinase (ASM) deficiency, which can be categorized as either Niemann-Pick disease type A [NPD-A], with progressive neurological disease and death in early childhood, or as Niemann-Pick disease type B [NPD-B], with a more variable spectrum of manifestations. Enzyme replacement therapy (ERT) with recombinant sphingomyelinase is currently studied as potential treatment for NPD-B patients. The objective of this study is to characterize the clinical features of patients with ASM deficiency in the Netherlands and Belgium with focus on the natural disease course of NPD-B patients. Prospective and retrospective data on ASM deficient patients were collected in The Netherlands and part of Belgium. Patients with NPD-B that could be followed prospectively were evaluated every 6-12 months for pulmonary function tests, 6 minute walk test (6 MWT), imaging (bone marrow infiltration measured by QCSI, organ volumes by MRI and CT scan of the lungs) and biochemical markers. Twenty-five patients with ASM deficiency were identified (13 males, 12 females, median age 13years, range 1-59 years). Nine patients had died at the time of the study, including four NPD-A patients at the age of 1,1, 2, 3 and five NPDB patents at the age of 5, 6, 43, 56 and 60 years. There was a high prevalence of homozygosity and compound heterozygosity for the common p.Arg608del mutation in 43% and 19% of NPD-B patients, respectively. In NPD-B patients, thrombocytopenia was present in most, while anemia and leucopenia were less common (33% and 6 % respectively). HDL cholesterol was reduced in most patients. Pulmonary disease was severe in several patients. Follow-up up to 11 years revealed a gradual decrease in platelet count. Detailed investigations in 6 NPD-B patients with follow-up in 4 patients revealed remarkable stable disease parameters up to 6 years, with some decline in pulmonary function and 6 MWT. Bone marrow fat fractions were decreased, indicating the presence of storage macrophages. Lung involvement was not related to the extent of visceromegaly, cytopenia or bone marrow involvement. In conclusion, in NPD-B patients pulmonary disease is the most debilitating feature. Disease manifestations are mostly stable in attenuated patients. Bone marrow infiltration is a less prominent feature of the disease.

Clinical and diagnostic approach in unsolved CDG patients with a type 2 transferrin pattern.

Dysmorphic features, multisystem disease, and central nervous system involvement are common symptoms in congenital disorders of glycosylation, including several recently discovered Golgi-related glycosylation defects. In search for discriminative features, we assessed eleven children suspected with a Golgi-related inborn error of glycosylation. We evaluated all genetically unsolved patients, diagnosed with a type 2 transferrin isofocusing pattern in the period of 1999-2009. By combining biochemical results with characteristic clinical symptoms, we used a diagnostic flow chart to approach the underlying defect in patients with congenital disorders of glycosylation-IIx. According to specific symptoms and laboratory results, we initiated additional, targeted biochemical and genetic studies. We found a distinctive spectrum of congenital disorders of glycosylation type 2-associated anomalies including sudden hearing loss, brain malformations, wrinkled skin, and epilepsy in combination with skeletal dysplasia, dilated cardiomyopathy, sudden cardiac arrest, abnormal copper and iron metabolism, and endocrine abnormalities in our patients. One patient with severe cortical malformations and mild skin abnormalities was diagnosed with a known genetic syndrome, due to an ATP6V0A2 defect. Here, we present unique congenital disorders of glycosylation type 2-associated anomalies, including both ATPase-related and unrelated cutis laxa and sensorineural hearing loss, a recently recognized symptom of congenital disorders of glycosylation. Based on our findings, we recommend clinicians to consider congenital disorders of glycosylation in patients with cardiac rhythm disorders, spondylodysplasia and biochemical abnormalities of the copper and iron metabolism even in absence of intellectual disability.

Adult metachromatic leukodystrophy treated by allo-SCT and a review of the literature.

Five patients with adult-onset metachromatic leukodystrophy (MLD) underwent allo-SCT. Conditioning was reduced in intensity and grafts were obtained from voluntary unrelated donors. All but one graft were depleted of T-lymphocytes. Patient age at transplantation varied from 18 to 29 (median, 27) years. Two patients rejected their graft and MLD progressed. The recipient of the unmanipulated graft converted to complete donor chimerism with normalization of arylsulphatase A (ARSA) levels. Despite ARSA normalization, he deteriorated. Another patient was a mixed chimera. Following escalated doses of donor lymphocyte infusions he converted to complete donor chimerism. His levels of ARSA correlated positively with the percentage of donor cells and MLD was not progressive. The fifth patient died after 35 days from complications associated with GVHD. We conclude that results of allo-SCT in symptomatic MLD patients are poor. However, allo-SCT may stop progression of MLD in selected patients.

New cases of adult-onset Sandhoff disease with a cerebellar or lower motor neuron phenotype.

Sandhoff disease is a lipid-storage disorder caused by a defect in ganglioside metabolism. It is caused by a lack of functional N-acetyl-beta-d-glucosaminidase A and B due to mutations in the HEXB gene. Typical, early-onset Sandhoff disease presents before 9 months of age with progressive psychomotor retardation and early death. A late-onset form of Sandhoff disease is rare, and its symptoms are heterogeneous. As drug trials that aim to intervene in the disease mechanism are emerging, the recognition and identification of Sandhoff disease patients-particularly those with atypical phenotypes-are becoming more important. The authors describe six new late-onset Sandhoff cases demonstrating cerebellar ataxia or lower motor neuron (LMN) involvement combined with, mostly subclinical, neuropathy. Two different mutations were found: IVS 12-26 G/A and c.1514G-->A. In patients with either progressive cerebellar ataxia or LMN disease in the setting of a possibly recessive disorder, Sandhoff disease should be suspected, even when the onset age is over 45 years.