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ProfileXpert is a genomic facility from the Federative Institute of Neuroscience of Lyon, specialized in the analysis of transcriptome and genotyping using the DNA microarray technology. ProfileXpert was founded in 2003 to implement these technologies into the Federative Institute of Neuroscience of Lyon and in the Rhône-Alpes area. ProfileXpert has three objectives: to provide a global DNA microarray (transcriptome and genotyping) service, to reinforce teaching and training activity in genomic and finally, to develop research and development projects, focused on innovating micro- and nanogenomic tools and on the identification of clinical molecular markers. ProfileXpert is equipped with state-of-the-art required hardware and operates by highly-qualified staff. ProfileXpert is characterized by its capacities: to ensure two complementary approaches (transcriptome and genotyping); to propose a wide range of chips (expression, exons, promoters, mapping, tiling), applications (qualitative and quantitative expression, alternative splicing, loss of heterozygoty, copy number variation analysis) and genomes (human, mouse, rat, bovine, chicken, c-elegans…); to analyse micro-amount of nucleic acids compatible with clinical samples (biopsy, isolated cells, biological fluids…) and to ensure data analysis, by using data mining and data analysis softwares.

Mutations of the MEN1gene lead to the occurrence of multiple endocrine neoplasia type 1 (MEN1). To gain insights into the mechanisms of the tumorigenesis related to MEN1inactivation, we have used mice in which the Men1gene was specifically disrupted in pancreatic β-cells. In these mice, we observed full penetrance of insulinoma with defined histological characteristics of tumorigenesis. To identify the genetic factors taking part in the tumour development, we performed gene expression profiling analysis of these insulinomas at different stages. Here, we show that in late stage insulinomas, 56 genes are up-regulated and 194 are down-regulated more than fourfold compared with normal pancreatic islets. Clustering analysis reveals the deregulation of Hoxgene family and the genes involved in cell proliferation and cell cycle control. The altered expression of Igf2, Igfbp3and Igfbp6as well as cyclin A2, B2and D2are confirmed by quantitative RT-PCR, with the overexpression of all the three cyclins found in early stage insulinomas. Moreover, an increased proportion of cyclin A2- and D2-expressing cells and the overexpression of insulin-like growth factor 2 (IGF2) protein are detected in mouse Men1insulinomas by immunostaining. Interestingly, the analysis of DNA methylation patterns by quantitative serial pyrosequencing reveals that four specific CpGs in the intragenic differentially methylated region 2 (DMR2) region of the Igf2gene known to augment transcription through methylation are significantly hypermethylated in insulinomas of Men1β-cell mutant mice at 6 and 10 months of age, even before IGF2 overexpression can be detected. Thus, our data indicate the involvement of both genetic and epigenetic mechanisms in early tumorigenesis of β-cells related to MEN1inactivation.

The Na+-Ca2+exchanger (NCX) is well known to regulate intracellular Ca2+concentration in many excitable cells. To date, three members of the NCX exchanger gene family have been identified: NCX1, NCX2 and NCX3. In zona fasciculata of the bovine adrenal gland, a Na+-Ca2+exchanger has been recently found to display biochemical similarities with the NCX1 exchanger of renal basolateral cells. Although a Na+-Ca2+exchanger has already been characterized in zona medulla of adrenal gland, such an exchanger has never been studied in the adrenal cortex. Thus, we have used a reverse transcriptase-polymerase chain reaction (RT-PCR) assay with specific NCX primers on fasciculata cell cultures. The sequencing of the amplified fragment revealed, for the first time, the presence of a NACA3 isoform of the NCX1 exchanger in the adrenal cortex, similar to the one found in adrenal chromaffin cells and in renal cortex. This isoform is 92 and 94% identical in the portion compared with kidney Na+-Ca2+exchanger NACA3 of rat and pig, respectively.

Brown adipose tissue (BAT) is implicated in both cold-induced thermogenesis and regulation of energy expenditure and is mainly controlled by sympathetic innervation. To clarify the permissive and/or complementary roles of glucagon in cold-induced BAT activation, glucagon receptor gene expression and its modulation by sympathetic activity were investigated in rats. One pad of interscapular BAT was surgically denervated while the other pad was sham operated, then rats were either cold-exposed (CE) for 1 week at 4°C or kept near thermoneutrality (25°C, TN). Using a semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) assay, it was shown that cold exposure decreased (-44%) the relative abundance of BAT glucagon receptor mRNA, an effect which was prevented by unilateral surgical sympathectomy of BAT. The present results show a negative control by sympathetic nervous activity of glucagon receptor gene expression and/or of glucagon receptor mRNA stability in BAT of cold-exposed rats. The down-regulation of glucagon receptor expression during cold exposure does not support a major role of the peptide in the thermogenic control of BAT.

L. Espinosa, C. Chouabe, A. Morales, J. Lachuer, B. Georges, M. Fatemi, C. Terrenoire, Y. Tourneur and R. Bonvallet. Increased Sodium–Calcium Exchange Current in Right Ventricular Cell Hypertrophy Induced by Simulated High Altitude in Adult Rats. Journal of Molecular and Cellular Cardiology(2000) 32, 639–653. Ventricular hypertrophy is associated with an increase in action potential (AP) duration which is potentially arrhythmogenic. The implication of the Na–Ca exchange current (INa–Ca) in the lengthening of the AP is controversial. The role of this current in the increased duration of the low plateau of the AP in hypertrophied adult rat ventricular myocytes by simulated chronic high-altitude exposure (≈4500 m) was evaluated. Electrophysiological experiments were carried out on isolated right ventricular myocytes from exposed and control rats with the perforated patch or the conventional whole-cell technique in current or in voltage clamp condition. With the two techniques, a significant increase of the low plateau duration was observed in hypertrophied myocytes as compared to controls. The low plateau in hypertrophied myocytes was depressed when Na was replaced by Li and was no longer recorded when intracellular Ca was buffered with EGTA. Inward tail currents, evoked either on repolarization to −80 mV following a depolarizing pulse to +10 mV or by interrupted AP technique, were greater in hypertrophied than in control myocytes and were abolished when Na was replaced by Li or when intracellular Ca was buffered with EGTA, indicating an increased Na–Ca exchange activity. The Li-sensitive current-voltage curves, obtained by a voltage clamp ramp protocol with an intracellular calcium buffered solution, were not significantly different in both hypertrophied and control myocytes, suggesting no modification in the density of the Na–Ca exchange protein. This was corroborated by the lack of difference in NCX1 mRNA levels between right ventricles from control and exposed rats. We conclude that increased duration of the low plateau of rat ventricular AP in altitude cardiac hypertrophy may be attributed to an increase of the inward INa–Ca. This augmented INa–Camay result from a modification in the intracellular Ca homeostasis.

The aim of the present study was to explore in male rats the role of the catecholaminergic innervation of the hypothalamus in corticotropic and adrenal responses to different kinds of stress conditions. For this purpose, 6-hydroxydopamine (3 µg in 0.2 µl saline) was stereotaxically and bilaterally infused at two levels of the main noradrenergic ascending brain stem bundle (NAB-X). The efficiency of catecholaminergic denervation of the hypothalamus was checked by measuring noradrenaline concentrations in paraventricular nuclei punches by HPLC and was confirmed by a 86% fall in noradrenaline levels of NAB-X rats killed after the stress experiments. Seven days after lesioning the NAB, sham operated controls and NAB-X lesioned animals were divided into 4 groups and submitted to 4 different stressors, i.e.: 2 min ether vapors (n = 5), 1 h immobilization (n = 7), i.v. histamine (2 mg/kg; n = 7) or i.v. insuline (10 I.U./kg; n = 8) injections. ACTH and corticosterone were measured in blood samples sequentially taken from a chronic carotid cannula, before stress and at short intervals over the 2 following hours. In comparison to the respective control groups, NAB-X dramatically reduced the ACTH response to ether (-78%) and to restraint (-53%) stress whereas the corticosterone response was affected to a lesser extent. In contrast, NAB-X slightly altered these responses in the histamine-treated group, although, surprisingly, the ACTH response tended to decrease and that of corticosterone to increase. Finally, NAB-X provoked a biphasic response to insulineinduced hypoglycemia, with a very early (5 min) rise in ACTH and corticosterone in comparison to the control group, followed by a trend to low hormonal levels up to 120 min. These results strongly suggest a differential involvement of the hypothalamic noradrenergic innervation upon the hypothalamic-pituitary-adrenal axis according to the nature of stress conditions.

Summary: We present the nucleotide sequence of a new Alu family member that lies between the delta- and beta-globin genes in gorilla DNA. The sequence exhibits 91% similarity with a consensus sequence of the Alu family. It is flanked by a perfect repetition of a 16-nucleotide target sequence and terminates with 24 adenylic residues. As this sequence is absent at this locus in other primate DNAs, its insertion occurred less than 8 million years ago, thus supporting the idea that Alu sequences are still mobile elements in the hominoid genome.

L’évaluation préopératoire du risque de malignité des nodules thyroïdiens repose sur l’échographie et la cytoponction, au terme desquels 30 % sont classés indéterminés.