SEL1L expression in non-small cell lung cancer.

SEL1L gene product plays a role in cell transformation and tumor progression in human breast, pancreas, esophageal, and prostate cancer. SEL1L expression was evaluated in a series of 76 surgically resected non-small cell lung carcinomas to investigate its clinical significance. SEL1L is scarcely detectable in normal lung, whereas in the initial stages of cell transformation, it becomes consistently expressed with evident staining in bronchial squamous metaplasia and in associated dysplastic changes. SEL1L immunoreactivity can be detected both in the cytoplasm and less commonly in the nuclei; the subcellular location correlates with tumor histotype, with cytoplasmic immunoreactivity being most prevalent in squamous cell carcinomas (P = .0005) and nuclear immunoreactivity being associated with adenocarcinomas (P = .02). Nuclear import and export signals are present in the SEL1L coding sequence, justifying the different subcellular location of the protein. SEL1L immunoreactivity was inversely correlated with tumor grade (P = .05); when considering only the adenocarcinomas, a stronger association was found (P = .006). SEL1L messenger RNA and protein evaluation in lung cancer cell lines confirmed the expression of the gene and the dual subcellular location of the protein in lung tumors. The data here reported suggest that, in non-small cell lung carcinoma, SEL1L may be an indicator of cell transformation, thus having important biologic and clinical implications.

A novel polymorphism in SEL1L confers susceptibility to Alzheimer's disease.

Alzheimer's disease (AD) is considered to be a conformational disease arising from the accumulation of misfolded and unfolded proteins in the endoplasmic reticulum (ER). SEL1L is a component of the ER stress degradation system, which serves to remove unfolded proteins by retrograde degradation using the ubiquitin-proteosome system. In order to identify genetic variations possibly involved in the disease, we analysed the entire SEL1L gene sequence in Italian sporadic AD patients. Here we report on the identification of a new polymorphism within the SEL1L intron 3 (IVS3-88 A>G), which contains potential binding sites for transcription factors involved in ER-induced stress. Our statistical analysis shows a possible role of the novel polymorphism as independent susceptibility factor of Alzheimer's dementia.

RNA-mediated interference indicates that SEL1L plays a role in pancreatic beta-cell growth.

The specificity of SEL1L expression and promoter activity for the pancreatic cell population, its chromosomal location, as well as its similarities to the yeast Hrd3p protein, a component of HRD complex which is responsible for endoplasmic reticulum (ER)-associated degradation of numerous ER-resident proteins, prompted us to study its effects on beta cell function. In this study we show that lowering SEL1L expression, by using the short interfering RNAs technology as well as antisense transfection, resulted in severe perturbation of betaTC-3 growth and metabolic activity. We hypothesize that SEL1L may exert its function by protecting the cells from ER stress and could counteract immune responses.

A whole genome screen for linkage disequilibrium in multiple sclerosis performed in a continental Italian population.

We have systematically screened the genome for evidence of linkage disequilibrium (LD) with multiple sclerosis (MS) by typing 6000 microsatellite markers in case-control and family based (AFBAC) cohorts from the Italian population. DNA pooling was used to reduce the genotyping effort involved. Four DNA pools were considered: cases (224 Italian MS patients), controls (231 healthy Italians), index (185 index cases from trio families) and parents (the 370 parents of the patient included in the Index pool), respectively. After refining analysis of the most promising 14 markers to emerge from this screening process, only marker D2S367 retained evidence for association.

High mutational burden in the mtDNA control region from aged muscles: a single-fiber study.

The ageing process is associated with the accumulation of somatic mutations of mitochondrial DNA (mtDNA). The aged human skeletal muscle tissue presents a mosaic of fibers when stained histochemically for cytochrome c oxidase (COX) activity with a proportion of COX negative fibers. Given the potential relevance of any alteration in the mtDNA control region for replication, we analysed the correlation between the presence of mutations and their degree of heteroplasmy and the COX phenotype in individual muscle fibers of aged healthy donors.A region of the mtDNA D-loop was cloned from single fiber-derived DNA and multiple clones were analysed. This strategy showed that a high level of mutational burden is present in all fibers and that several types of mtDNA rearrangements are detectable: recurrent (A189G, T408A and T414G) and rare point mutations, length variations affecting the homopolymeric tract and the (CA)(n) repeat and macrodeletions. The aggregate mutational load in the D-loop region correlated with the single fiber COX phenotype, suggesting that the cumulative burden of multiple, individually rare, mtDNA alterations might functionally impair the mitochondrial genetic machinery.

A collection of 33 novel human mtDNA homoplasmic variants.

Mitochondria are involved in cellular energy production via oxidative phosphorylation and this function may be damaged by any mutation in mitochondrial DNA (mtDNA). To identify novel mtDNA mutations, we have developed a program to systematically screen the entire mitochondrial genome in a large number of individuals with clinical and/or morphological features of mitochondrial dysfunction, but still no genetic diagnosis. The sequence-data were obtained with an automated rapid system, which gave us a series of information: in the eleven mitochondrial genomes analyzed we observed the presence of 33 differences from the revised Cambridge Reference Sequence (Andrews et al., 1999), but they were all homoplasmic in the patients' tissues analyzed (skeletal muscle and blood), suggesting that they are unlikely to be primarily pathogenic though they may be co-responsible in the determination of the disease. This work can therefore help complete the already ample mtDNA polymorphism existent database.

The Interleukin 1-beta exonic (+3953) polymorphism does not alter in vitro protein secretion.

Several studies have described polymorphisms in the genes encoding interleukin-1alpha (IL-1alpha) and IL-1beta cytokines and their implication in the onset of Alzheimer disease or in linkage disequilibrium with an as yet to be identified second locus on chromosome 2. However, these results have been associated with the sporadic forms of Alzheimer disease. Here we present data on the effects of the interleukin 1-beta exonic polymorphism (+3953) using a series of cohort healthy control samples and in vitro protein secretion assays.

Cross-species conservation of SEL1L, a human pancreas-specific expressing gene.

SEL1L is a recently cloned and organ-specific expressing human gene whose function is still at an embryonic stage but displays several interesting characteristics, among which a remarkable cross-species conservation. During evolution, the gene structural complexity increased, suggesting a diversification of its function; however, several amino acid motifs remain perfectly conserved from the bacteria to the human protein. SEL1L is the human ortholog of the C. elegans gene sel-1; the latter is implicated in the negative regulation of LIN-12/GLP-1/Notch receptor proteins. These receptor proteins play fundamental roles in signal transduction pathways and are key players in cell fate determination during the development of various organs. Studies in model organisms, such as C. elegans, helped to illuminate fundamental mechanisms involved in normal cellular functions and human diseases. This paper describes the conserved nature of SEL1L across a wide range of species suggesting, that the encoded protein most likely exerts a very important biological function; it may belong to a subclass of genes considered to be "essential."

Cloning and functional analysis of the allelic polymorphism in the transcription regulatory region of interleukin-1 alpha.

A strong correlation between the TT(-889) genotype over CC(-889) of the interleukin-1 alpha (IL-1 alpha) transcription regulatory region and age of Alzheimer's disease onset has recently been reported. To determine the functional effects of the genetic variants on plasma protein levels, we cloned the promoter region and determined its activity. The TT genotype significantly increased the transcriptional activity of the IL-1 alpha gene with respect to the CC genotype. A slight increase of the IL-1 alpha mRNA and protein levels was also observed in the plasma.

Identification of single nucleotide variations in the coding and regulatory regions of the myelin-associated glycoprotein gene and study of their association with multiple sclerosis.

The myelin-associated glycoprotein (MAG) gene is an appealing candidate in the 19q13 Multiple Sclerosis (MS) candidate region. Using denaturing high performance liquid chromatography (DHPLC), we identified 14 single nucleotide polymorphisms (SNPs) in MAG coding and regulatory regions, and we tested their possible association with MS in Italian patient and control DNA pools. Eight variations had a frequency <0.05, i.e. below the detection limit in the pools. Of these, Arg537Cys was further studied with individually genotyped individuals and was detected in 1/189 patients and 0/85 controls. The frequency of the six remaining SNPs were not significantly different in pools including a total of 1266 patient and 1612 control chromosomes. Considering the statistical power of the experimental design, these results exclude the MAG gene as an MS susceptibility factor with an odds ratio (OR) equal or higher than 1.3.

Low-flow ischemia and hypoxia stimulate apoptosis in perfused hearts independently of reperfusion.

Post-ischemic reperfusion leads to apoptosis-linked loss of myocytes in cultured cells and in vivo. We tested the hypothesis that apoptosis develops without reperfusion in Langendorff-perfused hearts exposed to either low-flow ischemia (LFI) or hypoxia (H). Rat hearts were perfused with amino-acid-enriched Krebs-Henseleit buffer and exposed for 6 h to LFI (flow=2 ml/min, PO(2)=500+/-50mmHg, mean+/-SD), H (10ml/min, 120+/-15mmHg), or control conditions (C, 10ml/min, 500+/-50mmHg). At selected times, DNA-fragmentation was measured by agarose-gel electrophoresis and in situ TUNEL assay. After 6 h, the ratio (TUNEL-positive)/(total nuclei) was 0.620+/-0.027, 0.615+/-0.005, 0.404+/-0.021 in LFI, H and C, respectively. The ratio was 0.813+/-0.021 in hearts exposed to 90 min global no-flow ischemia and reperfused (5 h). To assess the role of membrane-diffusible factors, separate experiments were performed recirculating the medium and exposing hearts to LFI or H as above. The degree of apoptosis was the same in both the recirculating and non-recirculating modes. Thus, apoptosis develops by similar extents and in a time-dependent fashion in crystalloid-perfused rat hearts during LFI or H at the same oxygen shortage (flow.PO(2)), even without the reperfusion.