Bilateral vocal fold paresis: the only presenting sign of anti-MUSK antibody myasthenia gravis.

A previously fit and well 53-year-old man was referred to the otolaryngology clinic with intermittent stridor and was found to have bilateral vocal fold paresis. Subsequent airway compromise necessitated emergency surgical tracheostomy. The man was discharged home with tracheostomy in situ and a diagnosis of idiopathic bilateral vocal cord palsy, as all primary investigations were negative. Neurological disease was suspected following readmission to hospital several weeks later with diplopia. Electromyography and serum antibody testing confirmed a diagnosis of anti-muscle-specific tyrosine kinase antibody positive myasthenia gravis (MuSK-MG); a subset of MG where autoantibodies are directed against MuSK. Resolution of bilateral vocal fold paresis was found 8 months after a short course of immunoglobulin (intravenous immunoglobulin (IVIg)) and daily mycophenolate therapy was commenced. Multidisciplinary teamwork between ear, nose and throat surgeons, neurologists and speech therapists enabled successful decannulation of tracheostomy. The patient has recovered well and remains minimally symptomatic.

Effect of endogenous mutant and wild-type PINK1 on Parkin in fibroblasts from Parkinson disease patients.

Mutations in the PTEN-induced putative kinase 1 (PINK1), a mitochondrial serine-threonine kinase, and Parkin, an E3 ubiquitin ligase, are associated with autosomal-recessive forms of Parkinson disease (PD). Both are involved in the maintenance of mitochondrial integrity and protection from multiple stressors. Recently, Parkin was demonstrated to be recruited to impaired mitochondria in a PINK1-dependent manner, where it triggers mitophagy. Using primary human dermal fibroblasts originating from PD patients with various PINK1 mutations, we showed at the endogenous level that (i) PINK1 regulates the stress-induced decrease of endogenous Parkin; (ii) mitochondrially localized PINK1 mediates the stress-induced mitochondrial translocation of Parkin; (iii) endogenous PINK1 is stabilized on depolarized mitochondria; and (iv) mitochondrial accumulation of full-length PINK1 is sufficient but not necessary for the stress-induced loss of Parkin signal and its mitochondrial translocation. Furthermore, we showed that different stressors, depolarizing or non-depolarizing, led to the same effect on detectable Parkin levels and its mitochondrial targeting. Although this effect on Parkin was independent of the mitochondrial membrane potential, we demonstrate a differential effect of depolarizing versus non-depolarizing stressors on endogenous levels of PINK1. Our study shows the necessity to introduce an environmental factor, i.e. stress, to visualize the differences in the interaction of PINK1 and Parkin in mutants versus controls. Establishing human fibroblasts as a suitable model for studying this interaction, we extend data from animal and other cellular models and provide experimental evidence for the generally held notion of PD as a condition with a combined genetic and environmental etiology.

MDR1 variants and risk of Parkinson disease. Association with pesticide exposure?

The multidrug resistance protein 1 (MDR1 or ABCB1) gene encodes a P-glycoprotein that protects the brain against neurotoxicants. Certain MDR1 genetic variants are known to compromise the function of this transporter and may thus be associated with Parkinson disease (PD). We therefore conducted a large case-control study investigating the potential relationship between MDR1 variants and PD. We determined the frequency of three MDR1 variants in 599 European PD patients and controls and further stratified the population by ethnicity, age at onset, and exposure to pesticides. We detected no relevant association in either the entire sample, or when separately investigating by ethnic origin or age at onset. However, the distribution of c.3435C/T differed significantly between PD patients exposed to pesticides compared to those non-exposed (odds ratio=4.74; confidence interval=[1.009; 22.306]); p=0.047), suggesting that common MDR1 variants might influence the risk to develop PD in conjunction with exposure to pesticides.

Limbic and frontal cortical degeneration is associated with psychiatric symptoms in PINK1 mutation carriers.

BACKGROUND: Mutations in the PINK1 gene can cause Parkinson's disease and are frequently associated with psychiatric symptoms that might even precede motor signs. METHODS: To determine whether specific gray matter degeneration of limbic and frontal structures might be liable to different psychiatric symptoms in PINK1 mutation carriers, observer-independent voxel-based morphometry was applied to high-resolution magnetic resonance images of 14 PINK1 mutation carriers from a large German family and 14 age- and gender-matched healthy control subjects. RESULTS: Psychiatric diagnoses in PINK1 mutation carriers comprised major depression without psychotic symptoms and schizophrenia-spectrum, panic, adjustment, and obsessive-compulsive personality disorders. As hypothesized, the categorical comparison between all PINK1 mutation carriers and control subjects demonstrated atrophy of limbic structures, especially the hippocampus and parahippocampus. More specifically, multiple regression analysis considering all psychiatric subscores simultaneously displayed different frontal (prefrontal, dorsolateral, and premotor cortex) and limbic (parahippocampus and cingulate) degeneration patterns. The duration of the psychiatric disease was also correlated with the extent of limbic and frontal gray matter volume decrease. CONCLUSIONS: Our results support the hypothesis that limbic and frontal gray matter alterations could explain various psychiatric symptoms observed in PINK1 mutation carriers. Factors determining individual susceptibility to degeneration of certain brain areas remain to be elucidated in future studies.

Phenotypic spectrum and sex effects in eleven myoclonus-dystonia families with epsilon-sarcoglycan mutations.

Myoclonus-dystonia (M-D) due to SGCE mutations is characterized by early onset myoclonic jerks, often associated with dystonia. Penetrance is influenced by parental sex, but other sex effects have not been established. In 42 affected individuals from 11 families with identified mutations, we found that sex was highly associated with age at onset regardless of mutation type; the median age onset for girls was 5 years versus 8 years for boys (P < 0.0097). We found no association between mutation type and phenotype.

Tumor therapy mediated by lentiviral expression of shBcl-2 and S-TRAIL.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) can selectively kill tumor cells and, in combination with other agents, could enhance tumor therapy. We explored the combined therapeutic effects of a secretable form of (S) TRAIL-induced apoptosis and the downregulation of Bcl-2 in human gliomas. We constructed a lentiviral delivery system: 1) for the expression of short hairpin (sh) RNA to downregulate Bcl-2 and for the expression of S-TRAIL to induce apoptosis in glioma cells; and 2) to follow delivery in vitro and the fate of tumors in real time in vivo. We demonstrate that lentiviral-mediated simultaneous downregulation of Bcl-2 and S-TRAIL-induced apoptosis leads to an increased expression of activated caspase-3 and caspase-7, thus resulting in accelerated S-TRAIL-mediated apoptosis in glioma cells in vitro. Using a highly malignant human glioma model expressing EGFRvIII and firefly luciferase, we show that the combined effect of Bcl-2 downregulation and S-TRAIL-induced apoptosis results in complete eradication of gliomas compared to S-TRAIL monotherapy. These results show that simultaneous triggering of TRAIL-mediated death receptor pathway and downregulation of Bcl-2 by shRNA leads to enhanced eradication of gliomas and serves as a template in developing and monitoring combination therapies for the treatment of drug-resistant cancers.

Intrafamilial phenotypic and genetic heterogeneity of dystonia.

Most cases of early-onset primary torsion dystonia are caused by the same 3-bp (GAG) deletion in the DYT1 gene. We describe a large Serbian family with significant intrafamilial variability of the DYT1 phenotype, from asymptomatic carrier status to late-onset focal, and generalized jerky dystonia. Seven mutation carriers (six proven by direct analysis and one by inferred haplotype) were identified, but only two of them were affected by dystonia (penetrance reduced to 29%). In addition, three GAG-deletion-negative family members also developed dystonia (two multifocal dystonia and one torticollis), suggesting that their involuntary movements are due to some other etiological factor(s) (i.e., another dystonia gene), or may be psychogenic.

Clinical spectrum of homozygous and heterozygous PINK1 mutations in a large German family with Parkinson disease: role of a single hit?

BACKGROUND: Although homozygous mutations in the PTEN-induced putative kinase 1 (PINK1) gene have been unequivocally associated with early-onset Parkinson disease (PD), the role of single heterozygous PINK1 mutations is less clear. OBJECTIVE: To investigate the role of homozygous and heterozygous PINK1 mutations in a large German pedigree (family W). DESIGN: Mutation analysis of PINK1 and results of standardized neurological and motor examination by 3 independent movement disorder specialists, including blinded video rating. SETTINGS: University of Lübeck. PARTICIPANTS: Twenty family members. MAIN OUTCOME MEASURES: The PINK1 genotype and PD status of all family members. RESULTS: The index patient of family W carried a homozygous nonsense mutation (c.1366C>T; p.Q456X) and presented with a phenotype closely resembling idiopathic PD but with an onset at 39 years of age. The family included a total of 4 affected homozygous members (age, 60-71 years; age at onset, 39-61 years), 6 members with slight or mild signs of PD (affected) and a heterozygous mutation (age, 31-49 years), and 5 unaffected heterozygous mutation carriers (age, 34-44 years). Although none of the heterozygous affected family members was aware of their signs (asymptomatic), the clinical findings were unequivocal and predominantly or exclusively present on their dominant right-hand side, eg, unilaterally reduced or absent arm swing and unilateral rigidity. The heterozygous members were all considerably younger than the affected homozygous mutation carriers. CONCLUSIONS: Heterozygous PINK1 mutations may predispose to PD, as was previously suggested by the presence of dopamine hypometabolism in asymptomatic mutation carriers. Long-term follow-up of our large family W provides an excellent opportunity to further evaluate the role of single heterozygous PINK1 mutations later in life, which will have major implications on genetic counseling.

Effects of genetic variations in the dystonia protein torsinA: identification of polymorphism at residue 216 as protein modifier.

Four naturally occurring sequence variations have been found in the coding region of the DYT1 gene encoding torsinA. One of these, a 3 bp (DeltaGAG) deletion, underlies dominantly inherited cases of early-onset torsion dystonia. Others, including a single nucleotide polymorphism that replaces aspartic acid (D) at residue 216 with histidine (H) in 12% of normal alleles and two other rare deletions, have not been clearly associated with disease. To gain insight into how these sequence variations affect torsinA, we used the structure of the related protein ClpB to provide a model of torsinA's AAA+ domain. Motifs important for ATP hydrolysis-sensor 1 and sensor 2-were identified, mutagenized and used to validate predictions of this model. Inspection revealed that the DeltaGAG deletion associated with dystonia removes one residue from an alpha-helix in the C-terminal portion of the AAA+ domain. The resulting distortion in torsinA structure may underlie this mutant's known tendency to produce ER-derived inclusions as well as its proposed loss of function. The D/H polymorphism at residue 216 falls in the N-terminal portion of the AAA+ domain near the sensor 1 motif. Surprisingly, cells expressing torsinA with the polymorphic histidine developed inclusions similar to those associated with DeltaGAG-torsinA, indicating that this change may also affect torsinA structure. Introducing H216 into DeltaGAG-torsinA reduced its tendency to form inclusions, suggesting that the two changes offset each other. Our findings point to a structural basis for the defects associated with the disease-linked DeltaGAG deletion in torsinA. They also suggest possible connections between the allelic polymorphism at residue 216 and the penetrance of DYT1 dystonia, as well as a possible role for this polymorphism in related disease states.

Co-occurrence of restless legs syndrome and Parkin mutations in two families.

Recent studies have suggested an association between restless legs syndrome (RLS) and Parkinson's disease (PD). We present a large multigenerational family and a smaller family with RLS. A Parkin mutation was found in 10 of 20 patients from both families with idiopathic RLS but was not considered causative. The clinical phenotype did not differ between RLS patients with and without a Parkin mutation. Inheritance of RLS was consistent with autosomal dominant transmission, and linkage analysis excluded all three known loci for RLS.

Frequency analysis and clinical characterization of different types of spinocerebellar ataxia in Serbian patients.

The relative frequencies of different spinocerebellar ataxias (SCAs) vary widely among different ethnic groups, presumably due to a founder effect. We investigated the relative prevalence of SCA1-3, 6-8, 12, 17; dentate-rubro-pallidoluysian atrophy; and Friedreich's ataxia (FRDA) in Serbian patients with adult-onset (>20 years of age) hereditary and sporadic SCAs, and compared clinical features of patients with genetically confirmed SCAs. A total of 108 patients from 54 families (38 apparently dominant [ADCA] and 16 apparently recessive) with adult-onset hereditary ataxia and 75 apparently sporadic patients were assessed. Of 38 families with ADCA, 13 (34%) were positive for an expansion in an SCA1 and 5 families (13%) for an expansion in an SCA2 allele. In 20 families (53%), no expansions have been identified in any of the analyzed genes. Gaze palsy, spasticity, and hyperreflexia were significantly more common in SCA1, whereas slow saccades, hypotonia, hyporeflexia, and dystonia prevailed in SCA2 patients. Among the 16 families with an apparently recessive mode of ataxia inheritance, 4 (25%) were identified as having the FRDA mutation. Ataxia-causing mutations were identified in 8 (10.6%) of patients with apparently sporadic adult-onset ataxia.

RNAi blocks DYT1 mutant torsinA inclusions in neurons.

Early onset generalized dystonia is a dominantly inherited movement disorder caused by neuronal dysfunction without an apparent loss of neurons. The same single mutation (GAG deletion) causes most cases and results in loss of a glutamic acid (E) in the carboxy terminal region of torsinA (Delta302/303). To model the neuronal involvement, adult rat primary sensory dorsal root ganglion neurons in culture were infected with lentivirus vectors expressing human wild-type or mutant torsinA. Expression of the mutant protein resulted in formation of torsinA-positive perinuclear inclusions. When the cells were co-infected with lentivirus vectors expressing the mutant torsinA message and a shRNA selectively targeting this message, inclusion formation was blocked. Vector-delivered siRNAs have the potential to decrease the adverse effects of this mutant protein in neurons without affecting wild-type protein.

PINK1, Parkin, and DJ-1 mutations in Italian patients with early-onset parkinsonism.

Recessively inherited early-onset parkinsonism (EOP) has been associated with mutations in the Parkin, DJ-1, and PINK1 genes. We studied the prevalence of mutations in all three genes in 65 Italian patients (mean age of onset: 43.2+/-5.4 years, 62 sporadic, three familial), selected by age at onset equal or younger than 51 years. Clinical features were compatible with idiopathic Parkinson's disease in all cases. To detect small sequence alterations in Parkin, DJ-1, and PINK1, we performed a conventional mutational analysis (SSCP/dHPLC/sequencing) of all coding exons of these genes. To test for the presence of exon rearrangements in PINK1, we established a new quantitative duplex PCR assay. Gene dosage alterations in Parkin and DJ-1 were excluded using previously reported protocols. Five patients (8%; one woman/four men; mean age at onset: 38.2+/-9.7 (range 25-49) years) carried mutations in one of the genes studied: three cases had novel PINK1 mutations, one of which occurred twice (homozygous c.1602_1603insCAA; heterozygous c.1602_1603insCAA; heterozygous c.836G>A), and two patients had known Parkin mutations (heterozygous c.734A>T and c.924C>T; heterozygous c.924C>T). Family history was negative for all mutation carriers, but one with a history of tremor. Additionally, we detected one novel polymorphism (c.344A>T) and four novel PINK1 changes of unknown pathogenic significance (-21G/A; IVS1+97A/G; IVS3+38_40delTTT; c.852C>T), but no exon rearrangements. No mutations were found in the DJ-1 gene. The number of mutation carriers in both the Parkin and the PINK1 gene in our cohort is low but comparable, suggesting that PINK1 has to be considered in EOP.

Inhibition of N-linked glycosylation prevents inclusion formation by the dystonia-related mutant form of torsinA.

Most cases of early-onset torsion dystonia are associated with a mutation in the DYT1 gene that results in the loss of a glutamic acid residue in the carboxy terminus of the encoded protein, torsinA. When overexpressed in cultured cells, wild-type torsinA distributes diffusely throughout the endoplasmic reticulum (ER), while the dystonia-related mutant, torsinADeltaE, accumulates within multilamellar membrane inclusions. Here we show that inclusion formation requires the addition of an N-linked oligosaccharide to one of two asparagine residues within the ATP-binding domain of the mutant protein. In the absence of this modification, overexpressed torsinADeltaE was localized diffusely throughout the cell in a reticular pattern resembling that of wild-type torsinA. In contrast, the localization of wild-type torsinA did not appear to vary with its glycosylation state. These results thus indicate that torsinADeltaE must achieve a specific conformation to induce formation of intracellular membrane inclusions.

Parkin gene alterations in hepatocellular carcinoma.

The Parkin gene is an extremely large gene (1.5 Mb) within the highly unstable FRA6E common fragile site (CFS) region, which is frequently altered in ovarian, breast, and hepatocellular carcinomas. Because Parkin/FRA6E has genomic similarities to FHIT/FRA3B and WWOX/FRA16D, two other large tumor-suppressor genes that are within CFS regions, we were interested in characterizing Parkin gene alterations and their possible association with cancer. After analyzing 50 cancer-derived cell lines including 11 hepatocellular carcinoma (HCC) cell lines, we found that one HCC cell line, PLC/PRF/5, had a detectable homozygous deletion encompassing exon 3. Using quantitative duplex PCR and fluorescence in situ hybridization analysis to characterize the copy number changes of Parkin exons in HCC cell lines, we found that 4 of 11 HCC cell lines had heterozygous deletions of Parkin exons and one, Hep3B, had an exon duplication. Parkin protein expression was significantly decreased or absent in all 11 HCC cell lines. Furthermore, more than 50% of HCC primary tumors had decreased Parkin expression compared to that in normal liver tissue. Parkin gene-transfected PLC5 and Hep3B cells grew more slowly than vector-only transfectants and also showed increased sensitivity to apoptosis induced by cell-cycle inhibitors. Therefore, we suggest that Parkin may be involved in tumor suppression and that the loss of Parkin contributes to the development of hepatocarcinoma.

Clinical and genetic features of myoclonus-dystonia in 3 cases: a video presentation.

Many cases of myoclonus-dystonia (M-D) are caused by mutations in the epsilon-sarcoglycan (SGCE) gene. We describe 3 children with a similar clinical picture of autosomal dominant M-D and an SGCE mutation in only one of them, suggesting that M-D is genetically heterogeneous.

Alterations in the common fragile site gene Parkin in ovarian and other cancers.

The cloning and characterization of the common fragile site (CFS) FRA6E (6q26) identified Parkin, the gene involved in the pathogenesis of many cases of juvenile, early-onset and, rarely, late-onset Parkinson's disease, as the third large gene to be localized within a large CFS. Initial analyses of Parkin indicated that in addition to playing a role in Parkinson's disease, it might also be involved in the development and/or progression of ovarian cancer. These analyses also indicated striking similarities among the large CFS-locus genes: fragile histidine triad gene (FHIT; 3p14.2), WW domain-containing oxidoreductase gene (WWOX; 16q23), and Parkin (6q26). Analyses of FHIT and WWOX in a variety of different cancer types have identified the presence of alternative transcripts with whole exon deletions. Interestingly, various whole exon duplications and deletions have been identified for Parkin in juvenile and early-onset Parkinson's patients. Therefore, we performed mutational/exon rearrangement analysis of Parkin in ovarian cancer cell lines and primary tumors. Four (66.7%) cell lines and four (18.2%) primary tumors were identified as being heterozygous for the duplication or deletion of a Parkin exon. Additionally, three of 23 (13.0%) nonovarian tumor-derived cell lines were also identified as having a duplication or deletion of one or more Parkin exons. Analysis of Parkin protein expression with antibodies revealed that most of the ovarian cancer cell lines and primary tumors had diminished or absent Parkin expression. While functional analyses have not yet been performed for Parkin, these data suggest that like FHIT and WWOX, Parkin may represent a tumor suppressor gene.

Transformation behavior, chemical composition, surface topography and bending properties of five selected 0.016" x 0.022" NiTi archwires.

PURPOSE: The aim of this study was to characterize five selected commercial NiTi archwires in terms of their transformation behavior, chemical composition, surface topography and mechanical properties (at temperatures of 22 degrees C, 37 degrees C and 60 degrees C). MATERIAL AND METHODS: The rectangular orthodontic archwires investigated were Neo Sentalloy F80 (GAC, Central Islip, NY, USA), 35 degrees C Thermo-Active Copper NiTi (A-Company/Ormco, Glendora, CA, USA), Rematitan "Lite" (Dentaurum, Pforzheim, Germany), Titanol SE S (Forestadent, Pforzheim, Germany) and Titanal (Lancer, San Marcos, CA, USA) in size 0.016" x 0.022". The chemical composition and surface topography were analyzed by energy dispersive X-ray spectroscopy using an analytical scanning electron microscope (XL30, EDAX SUTW Saphire Detector; Philips, Eindhoven, Netherlands). The transition temperatures were measured by means of differential scanning calorimetry (DSC; Perkin-Elmer Pyris 1, Perkin-Elmer, Fremont, CA, USA) in a range of - 80 degrees C to + 80 degrees C. The mechanical properties and their dependence on temperature were determined by means of 3-point bending tests. The binary archwire materials were characterized by a two-phase structure (NiTi matrix and Ni3Ti4 precipitates). RESULTS: The SEM analyses revealed abradant residues in virtually all archwires, while DSC revealed complex transformation properties. In addition to the martensitic and austenitic transformations, an R-phase transformation was also detected. The bending tests showed pronounced loading and unloading plateaus. The martensitic archwires (Neo Sentalloy F80, 35 degrees C Thermo-Active Copper NiTi) were found to have a lower strength than the martensitic-austenitic (Rematitan "Lite") and the austenitic archwires (Titanol SE S, Titanal). With increasing temperature (in the range from 22 degrees C to 60 degrees C) a linear rise in the plateau forces was recorded. CONCLUSIONS: When assessing the quality of archwires, account should be taken of the surface quality, as it is this that determines corrosion resistance, biocompatibility and friction characteristics. The mechanical properties depend on the initial state; moderate plateau forces and plateau moments can only be achieved with martensitic archwires. In contrast to conventional steel alloys, the strength characteristics are heavily dependent on temperature and need to be known if NiTi archwires are to be used to optimal effect. In addition, the superelastic plateau is used only partially, if at all, when minimum leveling is required.

Evidence that paternal expression of the epsilon-sarcoglycan gene accounts for reduced penetrance in myoclonus-dystonia.

Myoclonus-dystonia (M-D) is a movement disorder characterized by rapid muscle contractions and sustained twisting and repetitive movements and has recently been associated with mutations in the epsilon-sarcoglycan gene (SGCE). The mode of inheritance is autosomal dominant with reduced penetrance upon maternal transmission, suggesting a putative maternal imprinting mechanism. We present an apparently sporadic M-D case and two patients from an M-D family with seemingly autosomal recessive inheritance. In both families, we detected an SGCE mutation that was inherited from the patients' clinically unaffected fathers in an autosomal dominant fashion. Whereas, in the first family, RNA expression studies revealed expression of only the mutated allele in affected individuals and expression of the normal allele exclusively in unaffected mutation carriers, the affected individual of the second family expressed both alleles. In addition, we identified differentially methylated regions in the promoter region of the SGCE gene as a characteristic feature of imprinted genes. Using a rare polymorphism in the promoter region in a family unaffected with M-D as a marker, we demonstrated methylation of the maternal allele, in keeping with maternal imprinting of the SGCE gene. Loss of imprinting in the patient with M-D who had biallelic expression of the SGCE gene was associated with partial loss of methylation at several CpG dinucleotides.

Epsilon-sarcoglycan mutations found in combination with other dystonia gene mutations.

Myoclonus-dystonia is a movement disorder associated with mutations in the epsilon-sarcoglycan gene (SGCE) in most families and in the DRD2 and DYT1 genes in two single families. In both of the latter families, we also found a mutation of SGCE. The molecular mechanisms through which the detected mutations may contribute to myoclonus-dystonia remain to be determined.