Gene expression in patients with abdominal aortic aneurysm - more than immunological mechanisms involved.

Abdominal aortic aneurysm (AAA) is a serious condition of unclear pathogenesis and progression. Two samples were collected from 48 patients during AAA surgery. One sample was collected from the aneurysm, the other from the aneurysm proximal neck where the tissue did not exhibit any aneurysmal changes. Subsequently, gene expression profiles using microarrays (Illumina) were compared in RNA extracted from the samples. Overall, 2,185 genes were found to be upregulated and 2,100 downregulated; from which 158 genes had a different expression with FDR<0.05 (False Discovery Rate) and FC>/=2 (Fold Change). Of this number, 115 genes were over-expressed and 43 under-expressed. The analysis of the gene list based on their biological pathways revealed that the regulation of inflammation was mediated by chemokine and cytokine signaling pathways, the integrin signaling pathway, and T and B cell activation. Moreover, a change was identified in the expression of genes involved in both intercellular and intracellular signaling systems.

Absence of spinocerebellar ataxia type 3/Machado-Joseph disease within ataxic patients in the Czech population.

Although spinocerebellar ataxia type 3 (SCA3)/Machado-Joseph disease is the most common type of SCA worldwide, we did not identify any cases of the disease amongst SCA patients in the Czech population. It has been proposed that the prevalence of large normal alleles correlates with the frequency of various types of SCA. We have therefore attempted to resolve the absence of SCA3 in our population by investigating, within 204 normal chromosomes, the frequency and nature of CAG repeats as well as two intragenic polymorphisms. We found that large normal alleles with more than 33 CAG repeats were observed at a frequency of only 0.49%. Whereas most of the expanded alleles worldwide have the CA haplotype, this was the least common (5.4%) variant observed in our study, although it was associated with a larger mean CAG repeat length (26.9). We postulate that the absence of SCA3 in the Czech population might be explained by the lack of large normal alleles and consequently a relatively small reservoir for aberrant CAG expansions at the SCA3 locus.

Fluorescent multiplex PCR--fast method for autosomal dominant spinocerebellar ataxias screening.

Expansion of CAG trinucleotide repeats has been shown to cause a number of autosomal dominant spinocerebellar ataxias such as SCA1, SCA2, SCA3/MJD, SCA6 and SCA7. These disorders are characterized by a wide inter- and intrafamiliar variation in clinical features. The same mutation can result in different phenotypes and the very similar phenotypes can be caused by different mutations. Therefore it is necessary to investigate more SCA genes (according to prevalence) to identify the causal elongation. We developed a fast and efficient screening method based on touchdown multiplex PCR with fluorescent labelled primers for the most common types of SCAs (SCA 1, 2, 3 and 7). It has been reliable in 113 probands tested. Fragment analysis was performed by using 6% denaturing polyacrylamide gel and employing the automated DNA sequencer. This method considerably shortens the process of molecular genetic screening of SCAs and might be used as a tip for designing other SCA screening sets.

Can ataxin-2 be down-regulated by allele-specific de novo DNA methylation in SCA2 patients?

Spinocerebellar ataxia type 2 (SCA2) is caused by a CAG trinucleotide repeat expansion within the coding region of the ataxin-2 gene. Affected individuals typically have between 34 and 57 CAG repeats. Signs of the disorder generally begin in adulthood and include progressive ataxia, dysarthria, tremor, hyporeflexia, and slow saccades. As with other trinucleotide repeat disorders, SCA2 exhibits an inverse correlation between the size of the CAG repeat and the age at onset of clinically detectable disease, with neonatal cases of SCA2 being reported in individuals harboring over 200 CAG repeats. However, a wide range of age at onset is typically observed, especially in individuals with < 40 CAG repeats. CAG repeat number alone explains approximately 25-80% of the variability. In this paper, we hypothesize that the level of mutant ataxin-2 protein in affected cells contributes to these differences. One of the mechanisms that might influence this protein levels is de novo DNA methylation, which would specifically target the allele with the expanded CAG repeat leading to transcriptional silencing. Consequently, the symptoms of SCA2 would occur later in the patient's life history. Our postulations, as well as those previously reported to account for the phenotype of SCA2, are discussed.

Eps8, a substrate for the epidermal growth factor receptor kinase, enhances EGF-dependent mitogenic signals.

A method which allows direct cloning of intracellular substrates for receptor tyrosine kinases (RTKs) was developed. By applying this technique to the study of the epidermal growth factor receptor (EGFR) signaling pathway, we have isolated a cDNA, designated eps8, which predicts a approximately 92 kDa protein containing an SH3 domain. Eps8 also contains a putative nuclear targeting sequence. Antibodies specific to the eps8 gene product recognize a protein of M(r) 97 kDa and a minor 68 kDa component, which are closely related, as demonstrated by V8 proteolytic mapping. The product of the eps8 gene is tyrosine-phosphorylated in vivo following EGF stimulation of intact cells and associates with the EGFR, despite the lack of a functional SH2 domain. Several other RTKs are also able to phosphorylate p97eps8. Thus, the eps8 gene product represents a novel substrate for RTKs. Adoptive expression of the eps8 cDNA in fibroblastic or hematopoietic target cells expressing the EGFR resulted in increased mitogenic response to EGF, implicating the eps8 gene product in the control of mitogenic signals.