Daniela Corongiu

mtDNA nt13708A variant increases the risk of multiple sclerosis.

Background Mitochondrial DNA (mtDNA) polymorphism is a possible factor contributing to the maternal parent-of-origin effect in multiple sclerosis (MS) susceptibility. Methods and Findings In order to investigate the role of mtDNA variations in MS, we investigated six European MS case-control cohorts comprising >5,000 individuals. Three well matched cohorts were genotyped with seven common, potentially functional mtDNA single nucleotide polymorphisms (SNPs). A SNP, nt13708 G/A, was significantly associated with MS susceptibility in all three cohorts. The nt13708A allele was associated with an increased risk of MS (OR = 1.71, 95% CI 1.28–2.26, P = 0.0002). Subsequent sequencing of the mtDNA of 50 individuals revealed that the nt13708 itself, rather than SNPs linked to it, was responsible for the association. However, the association of nt13708 G/A with MS was not significant in MS cohorts which were not well case-control matched, indicating that the significance of association was affected by the population structure of controls. Conclusions Taken together, our finding identified the nt13708A variant as a susceptibility allele to MS, which could contribute to defining the role of the mitochondrial genome in MS pathogenesis.

Heat shock protein 27 R127W mutation: Evidence of a continuum between axonal Charcot-Marie-Tooth and distal hereditary motor neuropathy

Background Heat shock protein 27 (HSP27) mutations have been reported to cause both Charcot-Marie-Tooth disease (CMT) type 2F and distal hereditary motor neuropathy (dHMN) although never previously in a single family. Objective To analyse clinical and electrophysiological findings obtained in a single large Sardinian family bearing the HSP27 R127W mutation. Methods Twenty-one members of a five generation Sardinian family have been studied, including thirteen members affected by peroneal muscular atrophy and proved heterozygous for the known HSP27 R127W mutation. Twelve patients and eight unaffected relatives were subjected to clinical examination. A standardised electrophysiological study was performed in eleven patients and six unaffected relatives. Results Mean age at onset (±SD) was 31.2±7.2 years. Mean age at investigation was 45.2±12.9 years and mean disease duration at the time of investigation was 14±12.9 years. According to current criteria for CMT2 and dHMN, of the 10 patients who had undergone both clinical and neurophysiological examination, five were diagnosed as CMT2, two as dHMN and a further two patients were labelled as an intermediate type. Finally, due to the presence of spastic paraplegia, the index patient did not meet established criteria for the diagnosis of CMT or dHMN. Discussion Findings obtained in the present study, broadening the spectrum of clinical manifestations of disorders associated with HSP27 mutations, support the hypothesis of a continuum between CMT2 and dHMN forms and suggest a possible common spectrum between these entities and several forms of CMT plus pyramidal features (HMSN V), providing important implications for molecular genetic testing.

A novel Cx32 mutation causes X-linked Charcot-Marie-Tooth disease with brainstem involvement and brain magnetic resonance spectroscopy abnormalities

The objective of this study was to study genetic and phenotypic features of a family with X-linked Charcot-Marie-Tooth consisting of a healthy father, affected mother, two affected sons and one healthy one. A detailed electrophysiological and neuroimaging study, along with sequencing of the Cx32 gene, was performed in all family members. A novel Cx32 123 G>C mutation, determining an aminoacid variation (Glu41Asp), was found in the mother and the affected sons. An alteration in brainstem evoked potentials was found in the mother and one affected son. The affected son, who underwent magnetic resonance imaging, showed symmetrical hyperintensities in paratrigonal white matter, not found in his heterozygous mother, while both subjects exhibited alterations in brain metabolite ratios derived from localised proton-magnetic resonance spectroscopy. These data extend previous findings about central nervous system involvement in Cx32 mutated subjects and further support a functional role of the protein expression in oligodendrocytes.

Bipolar affective disorder preceding frontotemporal dementia in a patient with C9ORF72 mutation: is there a genetic link between these two disorders?

The FTD includes three main clinical syndromes: the behavioural variant (bvFTD), semantic dementia and progressive non-fluent aphasia [1]. For the BvFTD, which is characterized by a rich constellation of psychiatric and behavioural symptoms, the differential diagnosis with psychiatric disorders like schizophrenia and affective disorders, may be challenging [2, 3]. Recently a hexanucleotide repeat expansion in a noncoding region of the C9ORF72 gene has been identified as the cause of chromosome 9p21-linked ALS, FTD and ALS–FTD [4, 5]. We report the case of a man who developed from the age of 42 years an affective disorder characterized by repeated manic and hypomanic episodes. The manic phases were characterized by euphoria, racing thoughts, logorrhea, very high self-esteem, artistic hyperproductivity, little need for sleep. The patient fulfilled the DSM IV criteria for bipolar affective disorder (BPAD) type I [6] and was put on a long term lithium therapy with a good clinical response; during manic phases haloperidol was administered up to the acute episode resolution. The patient worked as a teacher until the age of 57 years. A patient’s maternal uncle developed a parkinsonism at the age of 60. The patient aged 64 presented to our neurological unit because he developed euphoria, impulsivity, logorrhea, sexual disinhibition with his wife, delusional fixed ideas with repetitive behaviours. He was obsessed by the idea of cleaning or putting petrol in his car and spent all day watching sport TV shows. As many of these symptoms were present in the past BPAD episodes, the patient underwent a new psychiatric examination; low doses of haloperidol were started without any improvement, so the therapy was stopped. Later a progressive cognitive impairment appeared. Neuropsychological assessment showed marked attention and executive functions troubles, working memory impairment, anomia and verbal fluency dysfunction. Neurologic examination revealed mild parkinsonism with postural tremor, camptocormia, diffuse bradykinesia and rigidity. Extrapyramidal symptoms arose when the patient was not under antipsychotic treatment. Mutations of TARDBP and PGRN genes were excluded. The patient has been found positive for a GGGGCC hexanucleotide repeat expansion in the first intron of C9ORF72 gene ([70 repeats). Brain MRI showed bilateral frontotemporal atrophy, prominent in frontal areas (Fig. 1). The orbitofrontal cortex was more involved than the dorsolateral prefrontal cortex; moreover there was a significant involvement of the fronto-insula and anterior cingulate gyrus. Perfusion SPECT demonstrated reduction of uptake in the left frontotemporal and right frontoparietal regions (Fig. 1). Structural and functional neuroimaging findings G. Floris G. Borghero A. Cannas F. D. Stefano F. Marrosu Department of Neurology, Azienda Universitaria-Ospedaliera of Cagliari and University of Cagliari, Cagliari, Italy

Genetic loci linked to Type 1 Diabetes and Multiple Sclerosis families in Sardinia

BackgroundThe Mediterranean island of Sardinia has a strikingly high incidence of the autoimmune disorders Type 1 Diabetes (T1D) and Multiple Sclerosis (MS). Furthermore, the two diseases tend to be co-inherited in the same individuals and in the same families. These observations suggest that some unknown autoimmunity variant with relevant effect size could be fairly common in this founder population and could be detected using linkage analysis.MethodsTo search for T1D and MS loci as well as any that predispose to both diseases, we performed a whole genome linkage scan, sequentially genotyping 593 microsatellite marker loci in 954 individuals distributed in 175 Sardinian families. In total, 413 patients were studied; 285 with T1D, 116 with MS and 12 with both disorders. Model-free linkage analysis was performed on the genotyped samples using the Kong and Cox logarithm of odds (LOD) score statistic.ResultsIn T1D, aside from the HLA locus, we found four regions showing a lod-score ≥1; 1p31.1, 6q26, 10q21.2 and 22q11.22. In MS we found three regions showing a lod-score ≥1; 1q42.2, 18p11.21 and 20p12.3. In the combined T1D-MS scan for shared autoimmunity loci, four regions showed a LOD >1, including 6q26, 10q21.2, 20p12.3 and 22q11.22. When we typed more markers in these intervals we obtained suggestive evidence of linkage in the T1D scan at 10q21.2 (LOD = 2.1), in the MS scan at 1q42.2 (LOD = 2.5) and at 18p11.22 (LOD = 2.6). When all T1D and MS families were analysed jointly we obtained suggestive evidence in two regions: at 10q21.1 (LOD score = 2.3) and at 20p12.3 (LOD score = 2.5).ConclusionThis suggestive evidence of linkage with T1D, MS and both diseases indicates critical chromosome intervals to be followed up in downstream association studies.

Vitamin D Responsive Elements within the HLA-DRB1 Promoter Region in Sardinian Multiple Sclerosis Associated Alleles

Vitamin D response elements (VDREs) have been found in the promoter region of the MS-associated allele HLA-DRB1*15∶01, suggesting that with low vitamin D availability VDREs are incapable of inducing *15∶01 expression allowing in early life autoreactive T-cells to escape central thymic deletion. The Italian island of Sardinia exhibits a very high frequency of MS and high solar radiation exposure. We test the contribution of VDREs analysing the promoter region of the MS-associated DRB1 *04∶05, *03∶01, *13∶01 and *15∶01 and non-MS-associated *16∶01, *01, *11, *07∶01 alleles in a cohort of Sardinians (44 MS patients and 112 healthy subjects). Sequencing of the DRB1 promoter region revealed a homozygous canonical VDRE in all *15∶01, *16∶01, *11 and in 45/73 *03∶01 and in heterozygous state in 28/73 *03∶01 and all *01 alleles. A new mutated homozygous VDRE was found in all *13∶03, *04∶05 and *07∶01 alleles. Functionality of mutated and canonical VDREs was assessed for its potential to modulate levels of DRB1 gene expression using an in vitro transactivation assay after stimulation with active vitamin D metabolite. Vitamin D failed to increase promoter activity of the *04∶05 and *03∶01 alleles carrying the new mutated VDRE, while the *16∶01 and *03∶01 alleles carrying the canonical VDRE sequence showed significantly increased transcriptional activity. The ability of VDR to bind the mutant VDRE in the DRB1 promoter was evaluated by EMSA. Efficient binding of VDR to the VDRE sequence found in the *16∶01 and in the *15∶01 allele reduced electrophoretic mobility when either an anti-VDR or an anti-RXR monoclonal antibody was added. Conversely, the Sardinian mutated VDRE sample showed very low affinity for the RXR/VDR heterodimer. These data seem to exclude a role of VDREs in the promoter region of the DRB1 gene in susceptibility to MS carried by DRB1* alleles in Sardinian patients.

Genetic analysis for five LRRK2 mutations in a Sardinian parkinsonian population: Importance of G2019S and R1441C mutations in sporadic Parkinson’s disease patients

Mutations in the LRRK2 gene are the most common known cause of familial and sporadic Parkinsons disease (PD). Few studies performed to date to assess frequency of these mutations are actually only representative of specific areas. Here we study the frequency and clinical phenotype of LRRK2 G2019S, I2020T and R1441C/G/H mutations in 356 Sardinian patients with idiopathic PD and 208 controls. Seventeen additional subjects, relatives of PD mutated probands, were enrolled. Eight patients were mutated in heterozygosis for LRRK2 gene (2.3%): six carried the G2019S (1.7%) and two the R1441C (0.6%) mutation. Three PD patients G2019S carriers (50%) were detected in two contiguous villages comprising 3921 inhabitants while the other three (50%) were identified in the remaining population of 796,079 inhabitants. Only one mutated proband had a family history of PD. LRRK2 G2019S and R1441C mutations associated with PD were not an uncommon mutation in a Sardinian population, especially in sporadic PD patients. The detection of the G2019S variant in ten unaffected relatives confirms a reduced penetrance of the underlying mutation and might explain its prevalence among patients with sporadic PD. These findings may provide new insights into the importance of studies of frequency of LRKK2 mutations in PD patients originating from small ethnically homogeneous populations.

Parkin Exon Rearrangements and Sequence Variants in LRRK2 Mutations Carriers: Analysis on a Possible Modifier Effect on LRRK2 Penetrance

Mutations in LRRK2 represent the most common causes of Parkinsons disease (PD) identified to date, but their penetrance is incomplete and probably due to the presence of other genetic or environmental factors required for development of the disease. We analyzed the presence of parkin sequence variants (mutations or polymorphisms) and exon rearrangements in LRRK2 mutations carriers (both PD patients and unaffected relatives) in order to detect a possible modifier effect on penetrance. Eight families with nine PD patients with heterozygous LRRK2 mutations (identified within 380 Sardinian PD patients screened for the presence of the five most common LRRK2 mutations) and sixteen additional relatives were genetically investigated for the presence of LRRK2 and parkin mutations. No evidence was found for the presence of pathological parkin mutations or exon rearrangements in patients or not affected family members. Three single-nucleotide polymorphisms (SNPs) were identified both in patients and unaffected relatives but did not significantly differ between the two groups. These data provide no support to the hypothesis whereby such parkin gene mutations may be commonly implicated in possible effect on penetrance in LRRK2 mutation carriers.

Constructional apraxia in frontotemporal dementia associated with the C9orf72 mutation: Broadening the clinical and neuropsychological phenotype

Abstract In our study we analysed clinical and neuropsychological data in a cohort of 57 Sardinian patients with FTD (55 apparently unrelated and two belonging to the same family), who underwent genetic screening for the C9orf72 mutation. Eight out of 56 patients were found positive for the C9orf72 mutation representing 14% of the entire cohort and 31.6% of the familial cases (6/19). C9orf72 mutated patients differed from the other FTD cases of the cohort for a younger age of onset, higher frequency of familial history for FTD and higher prevalence of delusional psychotic symptoms and hallucinations. In the neuropsychological assessment, C9orf72 mutated patients differed from non-mutated for the high frequency of visuospatial dysfunction regarding constructional apraxia (p = 0.02). In conclusion, our study confirms that Sardinian FTD patients have peculiar genetic characteristics and that C9orf72 mutated patients have a distinctive clinical and neuropsychological profile that could help differentiate them from other FTD patients. In our cohort we found that constructional apraxia, rarely reported in FTD, can properly discriminate between C9orf72 mutated and non-mutated patients and contribute to broaden the neuropsychological profile in frontotemporal dementia associated with this mutation.

Phenotypic variability related to C9orf72 mutation in a large Sardinian kindred

Abstract We investigated intrafamilial phenotypic variability in carriers of the C9orf72 mutation, analysing clinical, neuropsychological and imaging characteristics of various members from a large Sardinian kindred with FTD or ALS. We compared these with those of C9 + patients in our ALS and FTD cohorts. Results showed that three patients carried the C9orf72 mutation: two with ALS and one with FTD and Parkinsonism. C9 + patients in our bvFTD Sardinian cohort had a higher frequency of Parkinsonism than non-mutated patients (75% vs. 36.3%, p <0.02). Parkinsonism was present in 2.7% of our ALS cohort and 3.3% of the C9 + patients. The prevalence of Parkinsonism in C9 + patients in the bvFTD and ALS cohorts showed a statistically significant difference (p <0.006). In conclusion, Parkinsonism was frequently associated with FTD but not ALS in a large Sardinian family, a finding reflected in the wider C9orf72 associated Sardinian ALS and FTD populations.