Absence of linkage between the angiotensin converting enzyme locus and human essential hypertension.

The angiotensin converting enzyme (ACE) is a key component of the renin angiotensin system that contributes to the regulation of blood pressure (BP). Recent demonstration of linkage between the ACE locus and elevated BP in a rat model of hypertension has further emphasized ACE as a candidate gene in human hypertension. We report the localization of the ACE gene on the genetic map of chromosome 17, and identify an extremely polymorphic marker at the human growth hormone (hGH) locus which shows no recombination with ACE. We have found no evidence to support linkage between the ACE locus and hypertension, which suggests that mutations at the ACE locus do not commonly contribute to the pathogenesis of hypertension in our test population.     

A common polymorphism of the growth hormone receptor is associated with increased responsiveness to growth hormone

Growth hormone is used to increase height in short children who are not deficient in growth hormone, but its efficacy varies largely across individuals. The genetic factors responsible for this variation are entirely unknown. In two cohorts of short children treated with growth hormone, we found that an isoform of the growth hormone receptor gene that lacks exon 3 (d3-GHR) was associated with 1.7 to 2 times more growth acceleration induced by growth hormone than the full-length isoform (P < 0.0001). In transfection experiments, the transduction of growth hormone signaling through d3-GHR homo- or heterodimers was approximately 30% higher than through full-length GHR homodimers (P < 0.0001). One-half of Europeans are hetero- or homozygous with respect to the allele encoding the d3-GHR isoform, which is dominant over the full-length isoform. These observations suggest that the polymorphism in exon 3 of GHR is important in growth hormone pharmacogenetics.     

Les problèmes actuels du contrôle antidopage équin

Horse race as well as human dope testing are currently progressing and detection capabilities of laboratories must improve accordingly. Biomolecules such as Erythropoietin and Growth Hormone are up to date doping agents which break the integrity of racing and welfare of horses. New improvements of mass spectrometry allow detection of small concentrations of these peptides. Qualitative determinations can be performed when the structure of the hormone administered can be distinguished from the natural form. Selected indirect markers can also be used to detect abuse of growth hormone: these markers can be insulin growth factors (mainly IGF-1), binding proteins (IGFBP-3) and some bone markers such as osteocalcin. Encouraging results are already obtained but it is necessary to support and to encourage fundamental and applied research to maintain dope testing at the highest level.     

Techniques biologiques du diagnotic anténatal des infections virales et de la toxoplasmose

The main infections prenatally detected in fetuses are : cytomegalovirus parvovirus B19, rubella virus, and varicella-zoster infections. Today, prenatal diagnosis is currently performed after detection of maternal primary infection or because of abnormal ultrasound findings. Diagnosis of maternal primary infection is essentially based on the detection of specific IgM antibodies, but it was greatly improved by the use of complementary tests such as the measurement of the IgG avidity index.Prenatal diagnosis is based on the direct detection of the microorganism (CMV, toxoplasma), of its antigens (CMV) or of its genome. This direct detection can be done either on fetal blood or on amniotic fluid. Prenatal diagnosis can also be performed by detection of specific IgM in fetal blood (rubella). Non specific markers of fetal infection can also help in diagnosis. At the present time, prenatal diagnosis is essentially based on the detection of the genome in amniotic fluid.     

Neuroendocrine dysplasia in mice lacking protein tyrosine phosphatase sigma

Protein tyrosine phosphatase sigma (PTP-sigma, encoded by the Ptprs gene) is a member of the LAR subfamily of receptor-like protein tyrosine phosphatases that is highly expressed during mammalian embryonic development in the germinal cell layer lining the lateral ventricles of the developing brain, dorsal root ganglia, Rathke's pouch, olfactory epithelium, retina and developing lung and heart. On the basis of its expression and homology with the Drosophila melanogasterorthologues DPTP99 and DPTP100A (refs 5,6), which have roles in the targeting of axonal growth cones, we hypothesized that PTP-sigma may also have a modulating function in cell-cell interactions, as well as in axon guidance during mammalian embryogenesis. To investigate its function in vivo, we generated Ptprs-deficient mice. The resulting Ptprs-/-animals display retarded growth, increased neonatal mortality, hyposmia and hypofecundity. Anatomical and histological analyses showed a decrease in overall brain size with a severe depletion of luteinizing hormone-releasing hormone (LHRH)-immunoreactive cells in Ptprs-/- hypothalamus. Ptprs-/- mice have an enlarged intermediate pituitary lobe, but smaller anterior and posterior lobes. These results suggest that tyrosine phosphorylation-dependent signalling pathways regulated by PTP-sigma influence the proliferation and/or adhesiveness of various cell types in the developing hypothalamo-pituitary axis.     

Reliable identification of large numbers of candidate SNPs from public EST data

High-resolution genetic analysis of the human genome promises to provide insight into common disease susceptibility. To perform such analysis will require a collection of high-throughput, high-density analysis reagents. We have developed a polymorphism detection system that uses public-domain sequence data. This detection system is called the single nucleotide polymorphism pipeline (SNPpipeline). The analytic core of the SNPpipeline is composed of three components: PHRED, PHRAP and DEMIGLACE. PHRED and PHRAP are components of a sequence analysis suite developed to perform the semi-automated analysis required for large-scale genomes (provided courtesy of P. Green). Using these informatics tools, which examine redundant raw expressed sequence tag (EST) data, we have identified more than 3,000 candidate single-nucleotide polymorphisms (SNPs). Empiric validation studies of a set of 192 candidates indicate that 82% identify variation in a sample of ten Centre d'Etudes Polymorphism Humain (CEPH) individuals. Our results suggest that existing sequence resources may serve as a valuable source for identifying genetic variation.     

Érythropoïétine et dopage

Human erythropoietin (Epo) is a 30,4 kDa glycoprotein. It is composed of 165 amino acids and 4 carbohydrate side chain. Tissue hypoxia is the main stimulus for epo synthesis. Epo maintains red cell production by inhibiting apoptosis of erythrocyte progenotors, and by stimulating their proliferation and differentiation into normoblasts. Since the end eighties recombinant DNA technology has led to the large scale production of human erythropoietin for use as a antianaemic drug in patients with chronic renal failure. Quickly some athlete have used this hormone to increase their aerobic power and consequently their results specially in endurance sports. After several years of research now it is possible to detect it. The firts control took place during Sydney Olympic Games.     

Cryptorchidism in mice mutant for Insl3.

Impaired testicular descent (cryptorchidism) is one of the most frequent congenital abnormalities in humans, involving 2% of male births. Cryptorchidism can result in infertility and increases risk for development of germ-cell tumours. Testicular descent from abdomen to scrotum occurs in two distinct phases: the trans-abdominal phase and the inguino-scrotal phase. Currently, little is known about the factors that regulate the trans-abdominal phase of testicular descent. Leydig insulin-like hormone (Insl3) is a member of the insulin hormone superfamily expressed in the developing testis. We show here that mice mutant for Insl3 are viable, but exhibit bilateral cryptorchidism due to developmental abnormalities of the gubernaculum, resulting in abnormal spermatogenesis and infertility. Female homozygotes have impaired fertility associated with deregulation of the oestrus cycle. These findings reveal roles for Insl3 in the development of the urogenital tract and in female fertility. Insl3 may act as a hormone to regulate the growth and differentiation of the gubernaculum, thereby mediating intra-abdominal testicular descent.     

Conduite à tenir devant une sérologie d'hépatite C positive

The diagnosis of hepatitis C can be evoked in three settings : symptoms of liver disease, elevated serum alanine aminotransferase activity, and positive anti-HCV detection on systematic screening. The diagnosis is based on anti-hepatitis C virus (HCV) antibody detection by means of enzyme immuno-assays. HCV RNA detection should be restricted to the settings in which knowing the replicative status of the virus is needed, and to the assessment of the virological response to antiviral therapy. Pretreatment determination of viral load and HCV genotype should be used in future to tailor interferon plus ribavirin combination therapy duration.     

Mutations in SUFU predispose to medulloblastoma

Enchondromas are common benign cartilage tumors of bone. They can occur as solitary lesions or as multiple lesions in enchondromatosis (Ollier and Maffucci diseases). Clinical problems caused by enchondromas include skeletal deformity and the potential for malignant change to chondrosarcoma. The extent of skeletal involvement is variable in enchondromatosis and may include dysplasia that is not directly attributable to enchondromas. Enchondromatosis is rare, obvious inheritance of the condition is unusual and no candidate loci have been identified. Enchondromas are usually in close proximity to, or in continuity with, growth-plate cartilage. Consequently, they may result from abnormal regulation of proliferation and terminal differentiation of chondrocytes in the adjoining growth plate. In normal growth plates, differentiation of proliferative chondrocytes to post-mitotic hypertrophic chondrocytes is regulated in part by a tightly coupled signaling relay involving parathyroid hormone related protein (PTHrP) and Indian hedgehog (IHH). PTHrP delays the hypertrophic differentiation of proliferating chondrocytes, whereas IHH promotes chondrocyte proliferation. We identified a mutant PTH/PTHrP type I receptor (PTHR1) in human enchondromatosis that signals abnormally in vitro and causes enchondroma-like lesions in transgenic mice. The mutant receptor constitutively activates Hedgehog signaling, and excessive Hedgehog signaling is sufficient to cause formation of enchondroma-like lesions.     

Différenciation lymphoïde T méthodes d'exploration des lymphocytes T et applications aux lymphoproliférations T/NK

T-cells are generated in thymus from hematopoietic precursor cells. T-cell differentiation includes T-cell receptor gene rearrangement, positive and negative selection processes, and eventual maturation steps. T-cells recently emigrated from thymus are integrated into the pool of naive cells, which have not yet encountered their antigen. Apart from cytological examination, blood investigation in search of T/NK lymphoma is based on (i) immunophenotypic characterization of T and NK subsets using flow cytometry, allowing to suspect the diagnosis, but globally less determining than in the setting of B-cell proliferations ; and on (ii) detection of a T-cell clone, most often realized through a molecular — based technique, involving PCR.However, due to rarity and great variety of different entities encompassed by the wide term of T/NK lymphoma, the diagnosis should be firmly established only when taking into account the whole clinical, morphological, immunophenotypic, and molecular data.     

Hépatite chronique virale C

The hepatitis C virus (HCV) is a common cause of cirrhosis and hepatocellular carcinoma, as well as the most common reason for liver transplantation. HCV transmission occurs through exposure to infected blood (blood transfusion before 1990), injection drug users. Publications have implied that up to 40% of HCV seropositives do not have recognized parenteral risk factors. After contamination, 50 à 80% of patients do not clear virus, but develop chronic hepatitis C. HCV is a RNA virus; the rate of viral is high (10¹² virious per day) and the turnover of virus is rapid, the half-life of 2 to 3 hours. The diagnosis of chronic hepatitis C is based on both anti-HCV detection using enzyme immunoassays (EIA) and HCV RNA detection using a polymerase chain reaction (PCR) method. The HCV genotype (numbered 1 to 6) should be determined before treatment. The chronic sequelae of hepatitis C include progressive hepatic fibrosis, cirrhosis and hepatocellular carcinoma. In the majority of patients with chronic hepatitis C, the progression of fibrosis is insidious and slow. The factors associated with rapid progression of fibrosis are excessive alcohol consumption, older age, viral co-infection, immune deficiency, overweight. Liver biopsy provides a unique source of information on fibrosis, but significant progress has been made in the non invasive assessment of hepatitis fibrosis. Treatment is recommanded for patients with on risk of developing cirrhosis, characterized by bridging fibrosis. Combinaison therapy (peg interferon and ribavirine) should be used in the treatment of hepatitis C. Choise of duration of therapy and dose of ribavirin be based on HCV genotype. Patients with genotype 1 achieve sustained virological response of 45 à 50%, compared with rates of 80% with genotypes 2 or 3.     

Approche moléculaire de la résistance à la méticilline de Staphylococcus aureus

Staphylococcus aureus is one of the most common species met in medical microbiology laboratories. It causes many infectious diseases, and some of them are severe. Today, it can resist to many antibiotics, that were initially fully active, such as G or A group penicillin, by producing a penicillinase. Hospital strains can also resist to M group penicillins (methicillin, oxacillin) by modifying the β-lactam target: penicillin binding protein (PBP). Indeed, these strains produce a new PBP, called PBP 2a, of which affinity for β-lactams is much lower than the one of common Staphylococcus PBP. This PBP2a is encoded by mecA gene, of which expression is under control of several regulation genes. The expression of methicillin-resistance can be homogeneous or heterogeneous. Some strains can resist to methicillin using other mechanisms (penicillinase overproduction, methicillin-specific hydrolytic enzyme, PBP modifications). Particular methods are needed for the phenotypic detection of methicillin-resistance: antibiograms on NaCl-supplemented agar or incubated at 30 °C. Molecular detection of mecA gene by PCR permits to detect that resistance on strains for which phenotypic detection has failed.     

Diagnostic des pathologies liées au virus Epstein-Barr

Epstein-Barr virus (EBV), a member of the Herpesviridae familly, infects most human (95 % of the world population). In immunocompetent hosts primary infection results in acute infectious mononucleosis. In immunocompromised hosts, EBV is associated with lymphoproliferative disorders which occur with an incidence 30 to 50 higher than in the immunocompetent host. The laboratory diagnosis of an EBV infection is based on two techniques : demonstration of the virus by viral antigen or viral DNA detection and serologic responses. Direct culture of the virus on B lymphocytes is time consuming and is not of general use.     

Diagnostic des légionelloses

Legionellosis has three distinct forms: Legionnaire's disease, the more severe form of infection wich includes pneumonia, extrapulmonar legionellosis forms, and Pontiac fever, a milder illness. About 15% to 20% of known cases of Legionnaire's disease have been fatal. Delay in instituting appropriate therapy for Legionella pneumonia significantly increases mortality underlining the need for a rapid diagnosis. The distinction between Legionnaire's disease from other types of pneumonia by symptoms alone is not easily done. The diagnosis of legionellosis requires specialized laboratory tests not routinely performed on patients with fever and pneumonia. The definitive method is culture of Legionella but the growth of the organism is fastidious. Direct fluorescent-antibody staining is a rapid diagnostic test with a low sensitivity. The Legionella urinary antigen assay is a rapid test which detects only antigens of L. pneumophila serogroupe 1 in urine. Serologic tests are useful and necessite to evidence the increase of the antibody titers to Legionella in two blood samples obtained 3 to 6 weeks apart. It is a tardif diagnosis. Assays based on the polymerase chain reaction (PCR) have been used to detect Legionella in urine samples, bronchoalveolar-lavage fluid and serum. The primary advantage of this technique is the ability to detect Legionella rapidly and species other than L. pneumophila.     

Automating sequence-based detection and genotyping of SNPs from diploid samples

The detection of sequence variation, for which DNA sequencing has emerged as the most sensitive and automated approach, forms the basis of all genetic analysis. Here we describe and illustrate an algorithm that accurately detects and genotypes SNPs from fluorescence-based sequence data. Because the algorithm focuses particularly on detecting SNPs through the identification of heterozygous individuals, it is especially well suited to the detection of SNPs in diploid samples obtained after DNA amplification. It is substantially more accurate than existing approaches and, notably, provides a useful quantitative measure of its confidence in each potential SNP detected and in each genotype called. Calls assigned the highest confidence are sufficiently reliable to remove the need for manual review in several contexts. For example, for sequence data from 47-90 individuals sequenced on both the forward and reverse strands, the highest-confidence calls from our algorithm detected 93% of all SNPs and 100% of high-frequency SNPs, with no false positive SNPs identified and 99.9% genotyping accuracy. This algorithm is implemented in a software package, PolyPhred version 5.0, which is freely available for academic use.     

Dosage du cobalt par spectrométrie d'absorption atomique et application à l'analyse de la vitamine B12

We purpose in this study to describe a method for determination of vitamin B12 based in the speciation of the metal element: cobalt. First, a method for determination of cobalt by atomic absorption spectrometry with graphite furnace atomization was evaluated.Calibration was prepared with aqueous solution is linear from 10 to 150 μg/L (correlation coefficient = 0,98); the limit of detection is about 6,8 μg/L. The precision (repeatability: 1,2 à 6,9 %) and accuracy (108 %) were acceptable. This procedure was studied in serum with the same protocol. Secondly, the method was applicated to the determination of vitamin B12 in pharmaceutical injection and in serum. Recoveries ranged from 98,5 to 100,9 % for drugs. Comparison with results obtained with the microparticule enzymatic method (MEIA Abbott®) for serums were less satisfactory.