Multiple Sclerosis Severity Score: using disability and disease duration to rate disease severity.

BACKGROUND: There is no consensus method for determining progression of disability in patients with multiple sclerosis (MS) when each patient has had only a single assessment in the course of the disease. METHODS: Using data from two large longitudinal databases, the authors tested whether cross-sectional disability assessments are representative of disease severity as a whole. An algorithm, the Multiple Sclerosis Severity Score (MSSS), which relates scores on the Expanded Disability Status Scale (EDSS) to the distribution of disability in patients with comparable disease durations, was devised and then applied to a collection of 9,892 patients from 11 countries to create the Global MSSS. In order to compare different methods of detecting such effects the authors simulated the effects of a genetic factor on disability. RESULTS: Cross-sectional EDSS measurements made after the first year were representative of overall disease severity. The MSSS was more powerful than the other methods the authors tested for detecting different rates of disease progression. CONCLUSION: The Multiple Sclerosis Severity Score (MSSS) is a powerful method for comparing disease progression using single assessment data. The Global MSSS can be used as a reference table for future disability comparisons. While useful for comparing groups of patients, disease fluctuation precludes its use as a predictor of future disability in an individual.

Neurophysiological findings in SPG4 patients differ from other types of spastic paraplegia.

The authors examined 12 families with autosomal dominant hereditary spastic paraplegia for phenotypic characteristics predicting the underlying genotype. They found no clinical differences between patients with or without mutations in the spastin gene (SPG4). Motor evoked potentials and nerve conduction studies were almost normal in those with SPG4. In contrast, non-SPG4 families had prolonged central motor conduction times or marked peripheral neuropathy, or both.

Mutation analysis and association studies of nuclear factor-kappaB1 in sporadic Parkinson's disease patients.

Biochemical and morphological studies revealed that oxidative stress and apoptosis play a role in neurodegeneration in Parkinson's disease (PD). Reactive oxygen species may be directly involved in apoptosis or via upregulation of toxic cytokines, i.e. tumor necrosis factor alpha (TNFalpha). We recently demonstrated that the TNFalpha pathway contributes to the pathogenesis of sporadic PD using a genetic approach. These signalling pathways converge to the transcription factor nuclear factor kappaB (NF-kappaB), which has been found activated in affected neurons in PD. We performed a detailed mutation analysis of the p50 subunit of NF-kappaB (NFKB1 gene) in 96 sporadic PD patients. Previously, positive association was demonstrated in this cohort to chromosome 4q21-23 containing the NFKB1 gene. We identified three base exchanges not affecting the amino acid sequence, which were found at similar frequencies in controls. Our study does not support a genetically definable role of NFKB1 in the pathogenesis of sporadic PD.

Mutation analysis of the spastin gene (SPG4) in patients in Germany with autosomal dominant hereditary spastic paraplegia.

Hereditary spastic paraplegias (HSP) comprise a genetically and clinically heterogeneous group of neurodegenerative disorders characterized by progressive spasticity and hyperreflexia of the lower limbs. Autosomal dominant hereditary spastic paraplegia 4 linked to chromosome 2p (SPG4) is the most common form of autosomal dominant hereditary spastic paraplegia. It is caused by mutations in the SPG4 gene encoding spastin, a member of the AAA protein family of ATPases. In this study the spastin gene of HSP patients from 161 apparently unrelated families in Germany was analyzed. The authors identified mutations in 27 out of the 161 HSP families; 23 of these mutations have not been described before and only one mutation was found in two families. Among the detected mutations are 14 frameshift, four nonsense, and four missense mutations, one large deletion spanning several exons, as well as four mutations that affect splicing. Most of the novel mutations are located in the conserved AAA cassette-encoding region of the spastin gene. The relative frequency of spastin gene mutations in an unselected group of German HSP patients is approximately 17%. Frameshift mutations account for the majority of SPG4 mutations in this population. The proportion of splice mutations is considerably lower than reported elsewhere.

Inhibitors in the NFkappaB cascade comprise prime candidate genes predisposing to multiple sclerosis, especially in selected combinations.

Multiple sclerosis (MS) is an autoimmune disease displaying different clinical courses. In this multifactorial disease complex environmental as well as genetic predisposition factors contribute to the disease manifestation. Following the candidate gene approach we analysed several genes of the NFkappaB cascade, which are prime candidates for MS because of their involvement in almost all immunological reactions. MS association was excluded for the NFKB1 and NFKB3 genes, which show remarkably low degrees of polymorphism. The genes of NFkappaB inhibitors exhibit more sequence variations. In the IKBL gene a predisposing allele was identified (13.1% vs 7.5% in the control group, P < 0.001). This difference in the allelic distribution was even increased in the group of MS patients with a relapsing remitting course of the disease (14.9%, P < 0.0001). A protecting allele was found in the NFKBIA promotor for the patients with primary progressive MS (15.4% vs 28.4% in the control group, P < 0.01). Given predisposing alleles increase MS risk dramatically in certain combinations.

PCR/SSCP detects reliably and efficiently DNA sequence variations in large scale screening projects.

A simple and fast method with high reliability is necessary for the identification of mutations, polymorphisms and sequence variants (MPSV) within many genes and many samples, e.g. for clarifying the genetic background of individuals with multifactorial diseases. Here we review our experience with the polymerase chain reaction/single-strand conformation polymorphism (PCR/SSCP) analysis to identify MPSV in a number of genes thought to be involved in the pathogenesis of multifactorial neurological disorders, including autoimmune diseases like multiple sclerosis (MS) and neurodegenerative disorders like Parkinson s disease (PD). The method is based on the property of the DNA that the electrophoretic mobility of single stranded nucleic acids depends not only on their size but also on their sequence. The target sequences were amplified, digested into fragments ranging from 50-240 base pairs (bp), heat-denatured and analysed on native polyacrylamide (PAA) gels of different composition. The analysis of a great number of different PCR products demonstrates that the detection rate of MPSV depends on the fragment lengths, the temperature during electrophoresis and the composition of the gel. In general, the detection of MPSV is neither influenced by their location within the DNA fragment nor by the type of substitution, i.e., transitions or transversions. The standard PCR/SSCP system described here provides high reliability and detection rates. It allows the efficient analysis of a large number of DNA samples and many different genes.

The interferon gene cluster: a candidate region for MS predisposition? Multiple Sclerosis Study Group.

The clinical benefits of interferon (IFN) beta therapy in some multiple sclerosis (MS) patients are still unexplained, raising the question whether polymorphism within the IFNB gene itself would provide an explanation. Screening the IFNB gene by single strand conformation polymorphism (SSCP) analysis and sequencing, a single nucleotide polymorphism was identified. Both alleles were distributed with similar frequencies in MS patients and controls. Significant linkage disequilibrium (LD) between the IFN allele [153C] and allele [02] of the previously analyzed IFNA microsatellite (Epplen et al. Ann Neurol 1997; 41: 341-352) was observed in MS patients only, indicating a disease related haplotype. On the other hand an increased risk (RR = 12.41; Pc < 8 x 10(-5)) was observed for allele [07]. Hence the study was extended to neighbouring genes. Functionally relevant polymorphisms, i.e., premature stop codons in the IFNA10 [Cys20Stop] and IFNA17 [58Stop] genes and an aminoacid (aa) substitution [ile 184Arg] in the IFNA17 gene were analyzed. Patients carrying a non-functional IFNA17 allele bear an increased risk to develop MS (RR = 25.68; Pc < 0.06). In addition, LD analysis between IFNA10 [Cys20Stop], IFNA17 [58Stop] and the IFNA microsatellite alleles provides evidence for IFNA14, IFNA16 or IFNA5 as additional, most likely candidate genes. The present study excludes the IFNB gene as a candidate for MS predisposition but provides first evidence for predisposing IFNA genes.

Polymerase chain reaction-single strand conformation polymorphism or how to detect reliably and efficiently each sequence variation in many samples and many genes.

A simple and fast method with high reliability is necessary for the identification of mutations, polymorphisms and sequence variants (MPSV) within many genes and many samples, e.g. to clarify the genetic background of individuals with multifactorial diseases. We evaluated polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) analysis to identify MPSV in several genes, which are thought to be involved in the pathogenesis of multifactorial autoimmune diseases like multiple sclerosis. The method is based on the property, that the electrophoretic mobility of single-stranded nucleic acids depends not only on its size but also on its sequence. The target sequence was amplified, digested into fragments ranging from 50-200 bp, heat-denatured and analyzed on native gels. The analysis of 55 PCR systems, including a total of 145 fragments demonstrates, that the detection rate of MPSV depends primarily on the fragment lengths. Appropriate dilutions of samples enhances the proportion of ssDNA compared to dsDNA. Changing the gel conditions, glycerol concentrations and/or the addition of urea may increase fragment resolution in some cases. In general, the detection of MPSV is neither influenced by their location within the fragment nor by the type of substitution, i.e. transitions or transversions. The standard PCR-SSCP system described here provides high reliability and detection rates and allows the efficient analysis of many samples and many genes.

Differential stability of the (GAA)n tract in the Friedreich ataxia (STM7) gene.

Friedreich ataxia (FA) is an autosomal recessive, neurodegenerative disorder characterized by polypurine trinucleotide expansion. The (GAA)n motif is located in intron 18 of the STM7 gene (previously considered as intron 1 of the X25 gene) on chromosome 9q13. We studied the distribution profile of the polymorphic (GAA)n repetitive tract in 178 healthy individuals. The number of repeats of the trinucleotide block ranged from 7 to 29. In three individuals there were more than 29 repetitions of the GAA motif. While two of the individuals would be diagnosed as carriers of the FA mutation (GAA size > 90), the status of the third person, with a (GAA)58 tract, appears less clear at present. Thus an FA carrier rate of 1/60 to 1/90 can be assumed for the German population. In addition an intermediate-sized allele, (GAA)38 was identified in a mother with two affected children. The (GAA)38 allele appears to be expanded during transmission to at least (GAA)66 and (GAA) > 400 in her two FA-affected offspring. Therefore the shortest known STM7 allele conferring FA is (GAA)66. These novel facts have to be considered for differential diagnosis and definition of the FA carrier state.