Mutations of EXOSC3/Rrp40p associated with neurological diseases impact ribosomal RNA processing functions of the exosome in S. cerevisiae.

The RNA exosome is a conserved multiprotein complex that achieves a large number of processive and degradative functions in eukaryotic cells. Recently, mutations have been mapped to the gene encoding one of the subunits of the exosome, EXOSC3 (yeast Rrp40p), which results in pontocerebellar hypoplasia with motor neuron degeneration in human patients. However, the molecular impact of these mutations in the pathology of these diseases is not well understood. To investigate the molecular consequences of mutations in EXOSC3 that lead to neurological diseases, we analyzed the effect of three of the mutations that affect conserved residues of EXOSC3/Rrp40p (G31A, G191C, and W238R; G8A, G148C, and W195R, respectively, in human and yeast) in S. cerevisiae We show that the severity of the phenotypes of these mutations in yeast correlate with that of the disease in human patients, with the W195R mutant showing the strongest growth and RNA processing phenotypes. Furthermore, we show that these mutations affect more severely pre-ribosomal RNA processing functions of the exosome rather than other nuclear processing or surveillance functions. These results suggest that delayed or defective pre-rRNA processing might be the primary defect responsible for the pathologies detected in patients with mutations affecting EXOSC3 function in residues conserved throughout eukaryotes.

Progression of subcortical atrophy and iron deposition in multiple system atrophy: a comparison between clinical subtypes.

Magnetic resonance imaging (MRI) can be useful not only for the diagnosis of multiple system atrophy (MSA) itself, but also to distinguish between different clinical subtypes. This study aimed to investigate whether there are differences in the progression of subcortical atrophy and iron deposition between two variants of MSA. Two serial MRIs at baseline and follow-up were analyzed in eight patients with the parkinsonian variant MSA (MSA-P), nine patients with cerebellar variant MSA (MSA-C), and fifteen patients with Parkinson's disease (PD). The R2* values and volumes were calculated for the selected subcortical structures (caudate nucleus, putamen, globus pallidus, and thalamus) using an automated region-based analysis. In both volume and R2*, a higher rate of progression was identified in MSA-P patients. Volumetric analysis showed significantly more rapid progression of putamen and caudate nucleus in MSA-P than in MSA-C. With regard to R2* changes, a significant increase at follow-up and a higher rate of progression were identified in the putamen of MSA-P group compared to MSA-C and PD groups. This longitudinal study revealed different progression rates of MRI markers between MSA-P and MSA-C. Iron-related degeneration in the putamen may be more specific for MSA-P.

AMPD2 regulates GTP synthesis and is mutated in a potentially treatable neurodegenerative brainstem disorder.

Purine biosynthesis and metabolism, conserved in all living organisms, is essential for cellular energy homeostasis and nucleic acid synthesis. The de novo synthesis of purine precursors is under tight negative feedback regulation mediated by adenosine and guanine nucleotides. We describe a distinct early-onset neurodegenerative condition resulting from mutations in the adenosine monophosphate deaminase 2 gene (AMPD2). Patients have characteristic brain imaging features of pontocerebellar hypoplasia (PCH) due to loss of brainstem and cerebellar parenchyma. We found that AMPD2 plays an evolutionary conserved role in the maintenance of cellular guanine nucleotide pools by regulating the feedback inhibition of adenosine derivatives on de novo purine synthesis. AMPD2 deficiency results in defective GTP-dependent initiation of protein translation, which can be rescued by administration of purine precursors. These data suggest AMPD2-related PCH as a potentially treatable early-onset neurodegenerative disease.

Novel mutations in SCO1 as a cause of fatal infantile encephalopathy and lactic acidosis.

Isolated cytochrome c oxidase (COX) deficiency is a common cause of mitochondrial disease, yet its genetic basis remains unresolved in many patients. Here, we identified novel compound heterozygous mutations in SCO1 (p.M294V, p.Val93*) in one such patient with fatal encephalopathy. The patient lacked the severe hepatopathy (p.P174L) or hypertrophic cardiomyopathy (p.G132S) observed in previously reported SCO1 cases, so we investigated whether allele-specific defects in SCO1 function might underlie the genotype-phenotype relationships. Fibroblasts expressing p.M294V had a relatively modest decrease in COX activity compared with those expressing p.P174L, whereas both SCO1 lines had marked copper deficiencies. Overexpression of known pathogenic variants in SCO1 fibroblasts showed that p.G132S exacerbated the COX deficiency, whereas COX activity was partially or fully restored by p.P174L and p.M294V, respectively. These data suggest that the clinical phenotypes in SCO1 patients might reflect the residual capacity of the pathogenic alleles to perform one or both functions of SCO1.

Corticobasal degeneration with olivopontocerebellar atrophy and TDP-43 pathology: an unusual clinicopathologic variant of CBD.

Corticobasal degeneration (CBD) is a disorder affecting cognition and movement due to a progressive neurodegeneration associated with distinctive neuropathologic features, including abnormal phosphorylated tau protein in neurons and glia in cortex, basal ganglia, diencephalon, and brainstem, as well as ballooned neurons and astrocytic plaques. We identified three cases of CBD with olivopontocerebellar atrophy (CBD-OPCA) that did not have α-synuclein-positive glial cytoplasmic inclusions of multiple system atrophy (MSA). Two patients had clinical features suggestive of progressive supranuclear palsy (PSP), and the third case had cerebellar ataxia thought to be due to idiopathic OPCA. Neuropathologic features of CBD-OPCA are compared to typical CBD, as well as MSA and PSP. CBD-OPCA and MSA had marked neuronal loss in pontine nuclei, inferior olivary nucleus, and Purkinje cell layer. Neuronal loss and grumose degeneration in the cerebellar dentate nucleus were comparable in CBD-OPCA and PSP. Image analysis of tau pathology showed greater infratentorial tau burden, especially in pontine base, in CBD-OPCA compared with typical CBD. In addition, CBD-OPCA had TDP-43 immunoreactive neuronal and glial cytoplasmic inclusions and threads throughout the basal ganglia and in olivopontocerebellar system. CBD-OPCA met neuropathologic research diagnostic criteria for CBD and shared tau biochemical characteristics with typical CBD. These results suggest that CBD-OPCA is a distinct clinicopathologic variant of CBD with olivopontocerebellar TDP-43 pathology.

Pontocerebellar hypoplasia type 6 caused by mutations in RARS2: definition of the clinical spectrum and molecular findings in five patients.

Recessive mutations in the mitochondrial arginyl-transfer RNA synthetase (RARS2) gene have been associated with early onset encephalopathy with signs of oxidative phosphorylation defects classified as pontocerebellar hypoplasia 6. We describe clinical, neuroimaging and molecular features on five patients from three unrelated families who displayed mutations in RARS2. All patients rapidly developed a neonatal or early-infantile epileptic encephalopathy with intractable seizures. The long-term follow-up revealed a virtual absence of psychomotor development, progressive microcephaly, and feeding difficulties. Mitochondrial respiratory chain enzymes in muscle and fibroblasts were normal in two. Blood and CSF lactate was abnormally elevated in all five patients at early stages while appearing only occasionally abnormal with the progression of the disease. Cerebellar vermis hypoplasia with normal aspect of the cerebral and cerebellar hemispheres appeared within the first months of life at brain MRI. In three patients follow-up neuroimaging revealed a progressive pontocerebellar and cerebral cortical atrophy. Molecular investigations of RARS2 disclosed the c.25A>G/p.I9V and the c.1586+3A>T in family A, the c.734G>A/p.R245Q and the c.1406G>A/p.R469H in family B, and the c.721T>A/p.W241R and c.35A>G/p.Q12R in family C. Functional complementation studies in Saccharomyces cerevisiae showed that mutation MSR1-R531H (equivalent to human p.R469H) abolished respiration whereas the MSR1-R306Q strain (corresponding to p.R245Q) displayed a reduced growth on non-fermentable YPG medium. Although mutations functionally disrupted yeast we found a relatively well preserved arginine aminoacylation of mitochondrial tRNA. Clinical and neuroimaging findings are important clues to raise suspicion and to reach diagnostic accuracy for RARS2 mutations considering that biochemical abnormalities may be absent in muscle biopsy.

Changes of dopamine receptors in mice with olivocerebellar degeneration.

Lurcher mutants are mice with functional mutation in the 82 glutamate receptor (GluRdelta2) that is predominantly expressed in cerebellar Purkinje cells and plays a crucial role in cerebellar functions. These mice display ataxia and impaired motor-related learning tasks. In order to elucidate the role of dopaminergic receptor system in coping with mutation in delta2 glutamate receptor the behavioral effect (spatial learning) of D1 dopamine receptor activation and inhibition and changes in D1-like and D2-like dopamine receptors in striatum, hippocampus and cerebellum in C57BI/7 and C3H Lurcher mutants and wild type mice were studied. We have found that Lurcher mutants were worse in the spatial learning but mice of both types reacted similarly to D1 dopamine receptor agonist (without effect) and antagonist (worsening). Moreover, Lurchers revealed substantial higher density of both D1-like and D2-like dopamine receptors in hippocampus in C57BI/7 strain, while in C3H strain only D1-like dopamine receptors were higher. In C57BI/7 strain, D-like dopamine receptors were lower in cerebellum; D2-like dopamine receptors were not affected. In the striatum, the receptor densities were similar to the wild type counterparts. Our results suggest specific participation of dopamine receptor system in coping with olivocerebellar degeneration.

Selective loss of expression of glutamate GluR2/R3 receptor subunits in cerebellar tissue from a patient with olivopontocerebellar atrophy.

Expression of the mRNAs encoding the astrocytic (EAAT1, EAAT2) and neuronal (EAAT3, EAAT4) excitatory amino acid transporters and the AMPA-type glutamate receptor subunits GluR2 and GluR3 was investigated in postmortem cerebellar extracts from a patient with olivopontocerebellar atrophy (OPCA) and in material from three age-matched controls. Decreased expression in the steady state level of EAAT4 mRNA in the OPCA sample was correlated with the selective loss of Purkinje cells. Neuropathological evaluation revealed reactive gliosis and concomitantly increased expression of the mRNA encoding astrocytic glial fibrillary acidic protein (GFAP). Expression of the mRNAs encoding the AMPA receptor subunits GluR2 and GluR3 subunits was found to be decreased in OPCA suggesting that excitotoxic mechanism could play a role in the pathogenesis of the selective neuronal cell death in this disorder.

Oxidative injury is present in Purkinje cells in patients with olivopontocerebellar atrophy.

To verify the presence of lipid peroxidation products in spinocerebellar degeneration (SCD), the cerebella from eight patients with olivopontocerebellar atrophy (OPCA) and six non-OPCA patients were immunohistochemically investigated with 4-hydroxy-2-nonenal (HNE) antibody. On average, 84.6% of Purkinje cells were positively or strongly positively immunostained in OPCA patients while only 15.5% were positive in non-OPCA patients. Other cells in the molecular and granular layers showed no obvious immunoreactivity. These data suggest that a lipid peroxidation product is present in Purkinje cells of OPCA patients and that oxidative stress may play an important role in the degeneration process of SCD.

Preclinical impairment of the striatal dopamine transporter system in sporadic olivopontocerebellar atrophy: studied with [(123)I]beta-CIT and SPECT.

We investigated whether the dopamine (DA) transporter system is impaired in sporadic olivopontocerebellar atrophy (sOPCA) patients without clinical parkinsonism using the DA transporter radiotracer [(123)I]beta-CIT [2beta-carboxymethoxy-3beta-(4-iodophenyl)tropane] and single-photon emission computed tomography (SPECT). SPECT scans were acquired in 9 patients with sOPCA, 7 multiple system atrophy (MSA) patients with parkinsonism (MSA-P), and 7 age-matched healthy controls 20-24 h after the intravenous injection of [(123)I]beta-CIT. [(123)I]beta-CIT-specific binding in the striatum was determined as the radioactivity ratio of the striatum to the occipital cortex (specific binding ratio, SBR). In patients with sOPCA and MSA-P, SBRs in the right and left striatum and the mean SBR were significantly lower than those in controls (p < 0.05). The mean SBRs in patients with sOPCA and MSA-P were reduced to 69.0 and 60.7% of the control mean, respectively. However, there was no significant difference in SBRs between sOPCA and MSA-P patients. In sOPCA patients, the mean SBR was significantly correlated with the score of the clinical cerebellar function scale (r = -0.670, p = 0.024). These results indicate that even in the absence of clinical parkinsonism, the striatal dopaminergic system may be impaired in sOPCA. The DA transporter loss in sOPCA serves as another clue for sOPCA being a part of the spectrum of MSA.

Cerebellar activation during ataxic gait in olivopontocerebellar atrophy: a PET study.

OBJECTIVE: To investigate the possible abnormal regional brain metabolism during ataxic gait in olivopontocerebellar atrophy (OPCA), and to evaluate the response of the cerebellar subregions to instability during bipedal gait. MATERIAL AND METHODS: On 9 patients with OPCA in early phase and on 10 age-matched normal subjects, we performed positron emission tomography (PET) with 2-[18F]fluoro-2-deoxy-D-glucose (FDG) under two different conditions: supine resting and 30 min treadmill walking. RESULTS: Both in normals and in patients with OPCA, the FDG uptake in the walking state (Uwalk) was significantly greater than that in the resting state (Urest) in the pyramis, declive-folium-tuber and culmen of the cerebellar vermis, and in the thalamus. In the patients, the Uwalk was also significantly greater than the Urest in the posterior lobe of cerebellar hemisphere and in the pons and midbrain. In the pyramis, the activation ratio (= Uwalk/Urest) of the patients was significantly lower than that of the normals. CONCLUSIONS: We considered that these findings reflect the pathophysiology of ataxic gait in OPCA patients and the compensatory mechanism for the instability during ataxic gait.

A case of multiple system atrophy with hyperglycinaemia due to a selective deficiency of glycine transporter mRNA.

A patient presented with features of olivopontocerebellar atrophy and was found to have marked hyperglycinaemia. Severe atrophy of the cerebellum and brain stem was found at post-mortem, with numerous glial cytoplasmic inclusions (GCIs) in atrophic areas, characteristic of multiple system atrophy. In situ hybridization studies of the spinal cord demonstrated a selective reduction in expression of glycine transporter mRNA. We suggest that the resulting impairment of regulation of glycine concentrations at synaptic level resulted in excitotoxic damage to neurons.

Congenital olivopontocerebellar atrophy: report of two siblings with paleo- and neocerebellar atrophy.

We report two sisters with congenital olivopontocerebellar atrophy, including immunohistochemical studies of autopsy brain tissue. Both cases showed microcephaly with disproportionately marked hypoplasia of the posterior fossa structures including pons, inferior olivary nuclei, and cerebellum. Microscopically, the pons was atrophic with near total loss of pontine nuclei and transverse pontocerebellar tracts (inferior and middle cerebellar peduncles). The medulla showed absent inferior olivary and arcuate nuclei. The cerebellum showed hypoplasia with rudimentary dentate nuclei, profound loss of Purkinje cells and external granule cell layer, a sparse internal granule cell layer of the entire dorsal vermis and the dorsal portions of the lateral folia, as well as markedly reduced underlying axon fibers in the white matter with marked astrogliosis. These features were highlighted by immunohistochemical study using antibodies against Purkinje cell epitopes, synaptophysin, neurofilament, glial fibrillary acidic protein, and tuberin. The cerebral hemispheres were unremarkable. Our cases are characterized by a pattern of diffuse posterior cerebellar involvement that has rarely been described in previous reports. An autosomal recessive pattern of inheritance is suggested. The abnormalities may result from antenatal degeneration or atrophy of neurons in the involved sites rather than hypoplasia or developmental arrest starting in the second and third month of late embryonic life.

Loss of dopamine-D2 receptor binding sites in Parkinsonian plus syndromes.

UNLABELLED: This study analyzed temporal changes of striatal dopamine-D2 receptor binding during the course of different extrapyramidal movement disorders using 123I-iodobenzamide (IBZM) SPECT. METHODS: Eighteen patients (9 with Parkinson's disease, 9 with parkinsonian plus syndrome) were followed for 11-53 mo. Dopamine-D2 receptor binding was assessed using 123I-IBZM SPECT at the beginning and at the end of the follow-up period. SPECT data were acquired 120 min postinjection of 3-5 mCi 123I-IBZM. A semiautomated algorithm was applied to the raw data for semiquantitative evaluation of regional cerebral receptor binding. RESULTS: Intraobserver (r = 0.992) and interobserver (r = 0.930) variance was low for the semiautomated interpretation of the SPECT examination of the dopaminergic D2 receptor binding, reflecting a highly reproducible SPECT algorithm. Mean specific dopamine-D2 receptor binding was lower in patients with parkinsonian plus syndrome compared to patients with Parkinson's disease on the initial (p < 0.001) as well as the follow-up study (p < 0.001). In patients with Parkinson's disease, we observed an unaffected receptor binding compared to a reduced binding of radiotracer in patients with parkinsonian plus syndrome during the course of the disease (p < 0.001). CONCLUSION: During the follow-up, patients with Parkinson's disease showed a constant dopamine-D2 receptor binding. In contrast, patients with parkinsonian plus syndrome revealed a decline of the binding of dopamine-D2 receptor. These findings are in agreement with histopathological data that demonstrated a preserved dopamine-D2 receptor status in patients with Parkinson's disease and a decline of the dopamine-D2 receptors in patients with parkinsonian plus syndrome. SPECT examinations using 123I-IBZM are useful for assessing dynamic changes of dopamine-D2 receptors in extrapyramidal movement disorders. Semiquantitative SPECT evaluations may provide valuable information for clinical management and prognosis of the patient with extrapyramidal movement disorders.

Neuronal inclusions in the dentate fascia in patients with multiple system atrophy.

Ubiquitin-immunoreactive neuronal inclusions in the granular cells in the dentate fascia (UNIDs) of patients with multiple system atrophy (MSA) were examined for immunohistochemical and ultrastructural characterization especially in comparison with those which were recently reported for amyotrophic lateral sclerosis with dementia (ALS-D). Eight of 23 MSA patients had UNIDs which were also identified by Gallyas-Braak impregnation but immunonegative for other antibodies including against tau, neurofilaments, and alphaB crystallin. Ultrastructurally, loosely aggregated fibrils without limiting membrane located around the nucleus, which was confirmed by the results of ubiquitin-immunoelectron microscopy. The formation of UNIDs in MSA and ALS-D was suggested to be caused by different types of degeneration because UNIDs in MSA differ from these in ALS-D in terms of their stainability by Gallyas-Braak impregnation and ultrastructurally. In this study hippocampal involvement in MSA differing from ALS-D was clarified.

Positron emission tomography (PET) in Machado-Joseph disease.

Positron emission tomography studies on the regional cerebral glucose metabolism (rCMRglc) and 18F-fluorodopa (18F-Dopa) uptake were performed in 3 patients with Machado-Joseph disease (MJD), a dominantly inherited degenerative disease in the cerebellum, brainstem and basal ganglia. The rCMRglc in MJD was found to be significantly decreased in the cerebellum, brainstem, striatum and whole cerebral cortex in comparison to that in normal subjects. These results of rCMRglc were different from those for dominantly inherited olivopontocerebellar atrophy (dOPCA) or cerebellar cortical degeneration (CCD), however they were similar to those for sporadic olivopontocerebellar atrophy (sOPCA) and multiple system atrophy (MSA). The 18F-Dopa uptake in MJD was found to be significantly decreased in the putamen and relatively spared in the caudate, which was different from that of MSA. In addition, these results indicate that MJD showed a dysfunction, not only in the regions with apparent pathological involvement such as cerebellum, brainstem and nigro-striatal dopaminergic system, but also in the cerebral cortex and the striatum where no pathology could be observed using conventional morphological techniques.

Measurement of the dopaminergic degeneration in Parkinson's disease with [123I] beta-CIT and SPECT. Correlation with clinical findings and comparison with multiple system atrophy and progressive supranuclear palsy.

The cocaine derivative [123I] beta-CIT binds with high affinity to dopamine uptake sites in the striatum and can be used to visualize dopaminergic nerve terminals in vivo in the human brain with SPECT. It has been validated that the calculation of a simple ratio of specific/nondisplaceable binding during a period of binding-equilibrium in the striatum about 20 hrs after bolus injection of the tracer gives a strong and reliable index of the binding potential of dopamine uptake sites. Previous studies have shown that the dopaminergic deficit in patients with Parkinson's disease (PD) can clearly be visualized and quantified using this method. Our own results in a group of 113 patients with PD demonstrate a 45% loss of striatal [123I] beta-CIT binding in comparison to age corrected control values. Highly significant correlations of SPECT findings with clinical data obtained from the UPDRS rating scale such as akinesia, rigidity, axial symptoms and activities of daily living are demonstrated, while no correlation is found with tremor. The signal loss in a region comprising the whole striatum ranges from 35% in Hoehn/Yahr stage 1 to over 72% in stage V and is highly significantly correlated to the different stages of disease severity. A comparison of [123I] beta-CIT binding in the striatum contralaterally and ipsilaterally to the affected body side in 29 patients with hemiparkinson shows a loss of striatal binding of 41% contralaterally and 30% ipsilaterally. Results from subregional analyses in caudate and putamen show relative sparing of the caudate nucleus in PD. Data in 9 patients with multiple system atrophy (MSA) and 4 patients with progressive supranuclear palsy (PSP) are similar to the findings in PD although the differences between caudate and putamen are somewhat less marked. These data demonstrate that the dopaminergic nerve cell loss in PD and other disorders with a dopaminergic lesion can be quantified with [123I] beta-CIT and SPECT and that hopefully a preclinical or very early diagnosis is made possible. Such studies might also open the way for a better evaluation of neuroprotective strategies in PD. It does not seem to be possible however to differentiate PD and MSA or PSP with this method in individual cases.

Glucose metabolism in the cortical and subcortical brain structures in multiple system atrophy and Parkinson's disease: a positron emission tomographic study.

The brain glucose metabolism was studied by PET with 18F-FDG in 11 patients with multiple system atrophy (MSA) and 12 patients with idiopathic Parkinson's disease (PD). Seven of the 11 MSA patients were diagnosed as having olivopontocerebellar atrophy, two had striatonigral degeneration, while two demonstrated Shy-Drager syndrome. The glucose metabolic rates for each region in the PD patients showed no difference from the normal controls. The frontal, temporal and parietal cortical glucose metabolic rates and the caudate, the putaminal, the cerebellar and the brainstem glucose metabolic rates in the MSA patients decreased significantly from the controls. The atrophy of the cerebellum and the brainstem in the MSA patients were scored by MRI. The cerebellar and brainstem glucose metabolism in the MSA patients decreased as the atrophy score in such regions advanced in each group; however, some patients with no atrophy showed a decreased glucose metabolism. Although the cerebellar and the brainstem glucose metabolism decreased in all MSA patients, such a decrease was not observed in the SND patients. The decrease in the glucose metabolism for the non-cortical regions in the MSA patients seems to be due to a diffuse depletion of the neurons not restricted to the nigrostriatal neurons. Deafferentation to the cerebral cortices seems to result in a decreased cortical metabolism. The differences in the glucose metabolism between MSA and PD as assessed by PET may be caused by the pathophysiological differences between MSA and PD, and such differences therefore appear to be useful when making a differential diagnosis between MSA and PD. The relative sparing of the brainstem and cerebellar glucose metabolism is considered to be a feature of patients with SND.

Decreased striatal monoaminergic terminals in olivopontocerebellar atrophy and multiple system atrophy demonstrated with positron emission tomography.

We used [11C]dihydrotetrabenazine, a new ligand for the type 2 vesicular monoamine transporter (VMAT2), with positron emission tomography to study striatal monoaminergic presynaptic terminals in 4 patients with multiple system atrophy, 8 with sporadic olivopontocerebellar atrophy, and 9 normal control subjects. Specific binding in the striatum was significantly reduced in the multiple system atrophy patients as compared with the normal control group, with average reductions of 61% in the caudate nucleus (p = 0.002) and 58% in the putamen (p = 0.009). Smaller reductions were found in the sporadic olivopontocerebellar atrophy group, averaging 26% in the caudate nucleus (p = 0.05) and 24% in the putamen (p = 0.11). Mean blood-to-brain [11C]dihydrotetrabenazine transport (K1) was significantly different between groups only in the cerebellum, with values for the sporadic olivopontocerebellar atrophy group diminished compared with the normal control group. Cerebellar K1 was not significantly decreased in the multiple system atrophy group. The finding of reduced striatal VMAT2 in sporadic olivopontocerebellar atrophy patients suggests nigrostriatal pathology, indicating that some may later develop symptomatic extrapyramidal disease.