Raji P. Grewal

Large expansion of the ATTCT pentanucleotide repeat in spinocerebellar ataxia type 10

Spinocerebellar ataxia type 10 (SCA10; MIM 603516; refs 1,2) is an autosomal dominant disorder characterized by cerebellar ataxia and seizures. The gene SCA10 maps to a 3.8-cM interval on human chromosome 22q13–qter (refs 1,2). Because several other SCA subtypes show trinucleotide repeat expansions, we examined microsatellites in this region. We found an expansion of a pentanucleotide (ATTCT) repeat in intron 9 of SCA10 in all patients in five Mexican SCA10 families. There was an inverse correlation between the expansion size, up to 22.5 kb larger than the normal allele, and the age of onset (r2=0.34, P=0.018). Analysis of 562 chromosomes from unaffected individuals of various ethnic origins (including 242 chromosomes from Mexican persons) showed a range of 10 to 22 ATTCT repeats with no evidence of expansions. Our data indicate that the new SCA10 intronic ATTCT pentanucleotide repeat in SCA10 patients is unstable and represents the largest microsatellite expansion found so far in the human genome.

The observed human sperm mutation frequency cannot explain the achondroplasia paternal age effect

The lifelong spermatogonial stem cell divisions unique to male germ cell production are thought to contribute to a higher mutation frequency in males. The fact that certain de novo human genetic conditions (e.g., achondroplasia) increase in incidence with the age of the father is consistent with this idea. Although it is assumed that the paternal age effect is the result of an increasing frequency of mutant sperm as a man grows older, no direct molecular measurement of the germ-line mutation frequency has been made to confirm this hypothesis. Using sperm DNA from donors of different ages, we determined the frequency of the nucleotide substitution in the fibroblast growth factor receptor 3 (FGFR3) gene that causes achondroplasia. Surprisingly, the magnitude of the increase in mutation frequency with age appears insufficient to explain why older fathers have a greater chance of having a child with this condition. A number of alternatives may explain this discrepancy, including selection for sperm that carry the mutation or an age-dependent increase in premutagenic lesions that remain unrepaired in sperm and are inefficiently detected by the PCR assay.

Mitochondrial DNA deletion mutation levels are elevated in ALS brains.

This study was performed to explore the potential role of mitochondrial DNA mutations in the neurodegenerative process in amyotrophic lateral sclerosis (ALS). Using a semiquantitative assay, a common mitochondrial DNA deletion mutation (mt DNA4977) was assayed in brain tissue obtained from six sporadic ALS patients and compared to four controls. In each brain, levels of this mutation were measured in a brain region affected by neurodegeneration, the motor cortex (Brodmann area 4), and compared to the temporal cortex (Brodmann area 17). In the ALS but not control brains, levels of mt DNA4977 were an average of more than 30-fold (range 15–250) higher in Brodmann area 4. These results support and extend those of previous studies implying that mitochondria may participate in the neurodegenerative process in ALS.

Clinical and genetic analysis of a distinct autosomal dominant spinocerebellar ataxia

Objective: To characterize a distinct form of spinocerebellar ataxia (SCA) clinically and genetically. Background: The SCAs are a genetically heterogeneous group of neurodegenerative disorders affecting the cerebellum and its connections. The mutations for SCA1, 2, 3, 6, and 7 have been identified and shown to be due to expansion of a CAG repeat in the coding region of these genes. Two additional SCA loci on chromosomes 16 and 11 have been designated SCA4 and SCA5. However, up to 20% of individuals with autosomal dominant forms of ataxias cannot be designed any of these genotypes, implying the presence of other unidentified genes that may be involved in the development of ataxia. Methods: We ascertained and clinically characterized a six-generation pedigree segregating an autosomal dominant trait for SCA. We performed direct mutation analysis and linkage analysis for all known SCA loci. Results: The mutation analysis excludes SCA1, 2, 3, 6, and 7, and genetic linkage analysis excludes SCA4 and SCA5 (multipoint location scores < -2 across the candidate region). Clinical analysis of individuals in this family shows that all affected members have dysarthria, gait and limb ataxia, and nystagmus. No individuals have major brainstem or long-tract findings. Analysis of age at disease onset through multiple generations suggests anticipation. Conclusion: This pedigree represents a genetically distinct form of SCA with a phenotype characterized by predominantly cerebellar symptoms and signs.

C1qB and clusterin mRNA increase in association with neurodegeneration in sporadic amyotrophic lateral sclerosis.

We analyzed postmortem tissues of sporadic amyotrophic lateral sclerosis (SALS) for mRNA levels of two inflammatory proteins, complement C1qB and clusterin (apoJ). By Northern blot hybridization, SALS was associated with increased mRNA for C1qB and clusterin in the motor cortex (Brodmann area A4), but not in superior temporal cortex (A17), relative to neurologically normal controls. By in situ hybridization, SALS spinal cords showed increased C1qB and clusterin mRNA in areas undergoing neurodegeneration. This evidence implicates inflammatory mechanisms during neurodegenerative processes in SALS.

Scavenger receptor mRNAs in rat brain microglia are induced by kainic acid lesioning and by cytokines.

THE expression and localization of two distinct mRNAs from the macrophage scavenger receptor gene family were studied in rat brain cells in vivo and in vitro. In general, brains of control male rats showed low level signals by in situ hybridization for the macrophage scavenger receptor (MSR) and murine adherent macrophage (MAMA) receptor. In contrast, the reticular thalamic nucleus had a subpopulation of intensely labeled cells. Kainic acid (KA) treatment induced MSR and MAMA mRNA levels on different schedules in brain regions that are susceptible to KA, including hippocampal areas CA1 and CA3. The combination of immunocytochemistry and in situ hybridization localized theMSR and MAMA mRNA to microglia of KA-treated rats. Northern blot hybridization detected both MSR and MAMA mRNAs in primary cultures of mixed glia that contained microglia. Both MSR and MAMA mRNA were induced by treatment of primary mixedglia with lipopolysaccharide and interferon-γ, but not TGFβ1. MSR, but not MAMA, mRNA levels were increased after treatment with interleukin-1α.These results demonstrate the differential regulation of scavenger receptor mRNAs in microglia that is consistent with distinct roles for scavenger receptors in responses to neurodegeneration.

The spinal cord in lightning injury: A report of two cases

We present 2 patients in whom the predominant neurological complication following lightning strike was spinal cord injury. One patient, who was followed for 5 years, showed clinical, electrophysiologic and MRI evidence of cervical spinal cord injury. This patient had significant recovery, which in part, may be related to early and intense rehabilitation. The second patient presented with the symptoms and signs of spinal shock which then evolved into a myelopathy. Follow-up several months later showed almost complete resolution of symptoms. These patients demonstrate that the prognosis of spinal cord injury complicating a lightning strike may not be uniformly poor.

Neurodegeneration in Xeroderma Pigmentosum: a trinucleotide repeat mutation analysis

Xeroderma Pigmentosum (XP) is a rare autosomal recessive disorder caused by defects in DNA repair. In some forms, it is clinically and pathologically characterized by neurological involvement and premature neuronal death. This study explores the hypothesis that defects in DNA repair in XP may contribute to neurological involvement by destabilizing trinucleotide repeats during replication causing expansion mutations into disease producing ranges. Trinucleotide repeat instability in each of the genes causing Machado-Joseph Disease, myotonic dystrophy, Kennedys Disease and Huntingtons Disease was analyzed by performing single genome PCR. The results of trinucleotide repeat analysis of 360 single genomes from three different forms of XP showed that the size of the repeats were in the normal range and that there was no mitotic instability. These results suggest that in XP, trinucleotide repeat expansion mutations are not involved in the pathophysiology of neurodegeneration.

Genetic mapping and haplotype analysis of oculopharyngeal muscular dystrophy.

Oculopharyngeal muscular dystrophy (OPMD) is an autosomal dominant muscular dystrophy characterized by late onset ptosis, proximal muscle weakness and swallowing difficulties. This disease has been recently linked to chromosome 14q11.2-q13 in French-Canadian pedigrees. We studied three unrelated American families with OPMD of Hispanic descent and our results indicate that in this ethnic group, this disease also maps to chromosome 14q11.2-q13 (marker MYH7.24; Zmax = 3.98; &thetas;max = 0). These results represent an independent demonstration of disease linkage in a second distinct ethnic group. Furthermore, our analysis demonstrates a unique haplotype that is shared by affected individuals from all three families suggesting a founder effect for OPMD in this population. Meiotic recombinants and radiation hybrid mapping permit the narrowing of the critical region to 1 Mb which will facilitate positional cloning of the OPMD disease gene.

The mutation properties of spinal and bulbar muscular atrophy disease alleles.

ABSTRACTWe studied the gene for the trinucleotide repeat disorder X-linked spinal and bulbar muscular atrophy (SBMA) to quantify the spectrum of mutations and gain insight into genetic anticipation. This analysis was performed using single sperm typing from an affected individual. This method allows the quantification of large numbers of meioses and therefore provides accurate information about genetic instability of the CAG repeat expansions which cause SBMA. Among 198 X chromosome-containing sperm cells, 20% had a CAG repeat number equal to the donors somatic DNA of 49 CAG repeats, 56% were expansions, and 24% contractions. Most of the expansions (84%) and contractions (94%) were between 1 and 3 CAG repeats. These results are consistent with those obtained from one previously studied SBMA patient and reveal greater CAG repeat instability in sperm than in somatic tissue. Our results indicate that in SBMA, in contrast to sperm typing analysis of Huntingtons disease, there is relative stability of the CAG repeat number during paternal transmissions and that the spectrum of mutations is narrow. These results are in agreement with the limited available clinical data and suggest that anticipation may not be a significant feature of this disease.