Mapping of the hypokalaemic periodic paralysis (HypoPP) locus to chromosome 1q31-32 in three European families.

Hypokalaemic periodic paralysis (HypoPP) is an autosomal dominant muscle disease thought to arise from an abnormal function of ion channels. Performing a genome-wide search using polymorphic dinucleotide repeats, we have localized the HypoPP locus in three families of different geographic origin to chromosome 1q31-32, by linkage analysis. Using an intragenic microsatellite, we also demonstrate that the gene encoding the muscle DHP-sensitive calcium channel alpha 1 subunit (CACNL1A3) maps to the same region, sharing a 5 centiMorgan (cM) interval with the HypoPP locus. Moreover, CACNL1A3 co-segregates with HypoPP without recombinants in the two informative families, and is therefore a good candidate for the HypoPP gene.

Open-reading-frame sequence tags (OSTs) support the existence of at least 17,300 genes in C. elegans.

The genome sequences of Caenorhabditis elegans, Drosophila melanogaster and Arabidopsis thaliana have been predicted to contain 19,000, 13,600 and 25,500 genes, respectively. Before this information can be fully used for evolutionary and functional studies, several issues need to be addressed. First, the gene number estimates obtained in silico and not yet supported by any experimental data need to be verified. For example, it seems biologically paradoxical that C. elegans would have 50% more genes than Drosophilia. Second, intron/exon predictions need to be tested experimentally. Third, complete sets of open reading frames (ORFs), or "ORFeomes," need to be cloned into various expression vectors. To address these issues simultaneously, we have designed and applied to C. elegans the following strategy. Predicted ORFs are amplified by PCR from a highly representative cDNA library using ORF-specific primers, cloned by Gateway recombination cloning and then sequenced to generate ORF sequence tags (OSTs) as a way to verify identity and splicing. In a sample (n=1,222) of the nearly 10,000 genes predicted ab initio (that is, for which no expressed sequence tag (EST) is available so far), at least 70% were verified by OSTs. We also observed that 27% of these experimentally confirmed genes have a structure different from that predicted by GeneFinder. We now have experimental evidence that supports the existence of at least 17,300 genes in C. elegans. Hence we suggest that gene counts based primarily on ESTs may underestimate the number of genes in human and in other organisms.

Physicochemical properties of pH-controlled polyion complex (PIC) micelles of poly(acrylic acid)-based double hydrophilic block copolymers and various polyamines.

The physicochemical properties of polyion complex (PIC) micelles were investigated in order to characterize the cores constituted of electrostatic complexes of two oppositely charged polyelectrolytes. The pH-sensitive micelles were obtained with double hydrophilic block copolymers containing a poly(acrylic acid) block linked to a modified poly(ethylene oxide) block and various polyamines (polylysine, linear and branched polyethyleneimine, polyvinylpyridine, and polyallylamine). The pH range of micellization in which both components are ionized was determined for each polyamine. The resulting PIC micelles were characterized using dynamic light scattering and small-angle X-ray scattering experiments (SAXS). The PIC micelles presented a core-corona nanostructure with variable polymer density contrasts between the core and the corona, as revealed by the analysis of the SAXS curves. It was shown that PIC micelle cores constituted by polyacrylate chains and polyamines were more or less dense depending on the nature of the polyamine. It was also determined that the density of the cores of the PIC micelles depended strongly on the nature of the polyamine. These homogeneous cores were surrounded by a large hairy corona of hydrated polyethylene oxide block chains. Auramine O (AO) was successfully entrapped in the PIC micelles, and its fluorescence properties were used to get more insight on the core properties. Fluorescence data confirmed that the cores of such micelles are quite compact and that their microviscosity depended on the nature of the polyamine. The results obtained on these core-shell micelles allow contemplating a wide range of applications in which the AO probe would be replaced by various cationic drugs or other similarly charged species to form drug nanocarriers or new functional nanodevices.

Stepped horn actuated Kelvin probe.

We have developed an original Kelvin probe system using an ultrasonic stepped horn sonotrode. This actuator is optimized in order to maximize the velocity of the tip end, and hence to increase the Kelvin current detected. Such development is essential to improve surface potential measurements at small spatial scale.

Regulation of membrane trafficking by a novel Cdc42-related protein in Caenorhabditis elegans epithelial cells.

Rho GTPases are mainly known for their implication in cytoskeleton remodeling. They have also been recently shown to regulate various aspects of membrane trafficking. Here, we report the identification and the characterization of a novel Caenorhabditis elegans Cdc42-related protein, CRP-1, that shows atypical enzymatic characteristics in vitro. Expression in mouse fibroblasts revealed that, in contrast with CDC-42, CRP-1 was unable to reorganize the actin cytoskeleton and mainly localized to trans-Golgi network and recycling endosomes. This subcellular localization, as well as its expression profile restricted to a subset of epithelial-like cells in C. elegans, suggested a potential function for this protein in polarized membrane trafficking. Consistent with this hypothesis, alteration of CRP-1 expression affected the apical trafficking of CHE-14 in vulval and rectal epithelial cells and sphingolipids (C(6)-NBD-ceramide) uptake and/or trafficking in intestinal cells. However, it did not affect basolateral trafficking of myotactin in the pharynx and the targeting of IFB-2 and AJM-1, two cytosolic apical markers of intestine epithelial cells. Hence, our data demonstrate a function for CRP-1 in the regulation of membrane trafficking in a subset of cells with epithelial characteristics.

Mutations in the muscle sodium channel gene (SCN4A) in 13 French families with hyperkalemic periodic paralysis and paramyotonia congenita: phenotype to genotype correlations and demonstration of the predominance of two mutations.

Hyperkalemic periodic paralysis (hyperPP), paramyotonia congenita (PC) and PC with myotonia permanens are closely related muscle disorders of genetic origin due to allelic mutations in the muscle sodium channel gene, SCN4A. Seven families of French origin with hyperPP were studied. Five of these had the Thr704Met mutation, but 2 families, genetically linked to SCN4A, failed to show any of the known mutations of SCN4A. Correlations between the phenotype and the genotype were made for patients with the Thr704Met mutation. All 12 patients over 30 years old with the Thr704Met mutation presented muscle weakness due to degeneration of muscle fibers in addition to periodic paralysis. Only approximately 12.5% of patients with the Thr704Met mutation presented with clinical myotonia and about 50% with hyperkalemia. One family with PC displayed the Gly1306Val mutation with a phenotype similar to the one already reported for this mutation. Five families with either PC or PC with myotonia permanens had the Thr1313Met mutation indicating that the severity of myotonia and its permanence were variable. Two mutations of SCN4A were found to be predominant in these 13 families: the Thr704Met and the Thr1313Met mutations. Only 2 families with the Thr704Met mutation and 3 families with the Thr1313Met shared the same SCN4A haplotype determined with intragenic dinucleotide repeats. Recurrent mutations of SCN4A may contribute to the predominance of these two mutations in the French population.

The type I interferon receptor: structure, function, and evolution of a family business.

Recent results indicate that coherent models of how multiple interferons (IFN) are recognized and signal selectively through a common receptor are now feasible. A proposal is made that the IFN receptor, with its subunits IFNAR-1 and IFNAR-2, presents two separate ligand binding sites, and this double structure is both necessary and sufficient to ensure that the different IFN are recognized and can act selectively. The key feature is the duplication of the extracellular domain of the IFNAR-1 subunit and the configurational geometry that this imposes on the intracellular domains of the receptor subunits and their associated tyrosine kinases.

IgG synthesis within the central nervous system. Comparison of three formulas.

IgG synthesis within the central nervous system is the expression of a local pathologic immune reaction. Its detection is of great importance in a large number of neurologic diseases, especially multiple sclerosis (MS). The results obtained from the study of 115 neurologic patients (52 patients with MS and 63 patients with other neurologic diseases) using three different mathematic formulas (Tourtellotte's formula, the index of Tibbling and coworkers, and Schuller and Sagar's formula) are presented and statistically analyzed. Despite their obvious differences, the results from these three formulas are in accordance in 76% of the patients observed. The clinical correlations of these formulas are discussed; none of them allows a specific diagnosis of MS. The formula of Schuller and Sagar seems the most sensitive, detecting local central nervous system IgG synthesis in 79% of patients with MS, compared with 54% of patients with MS utilizing the index of Tibbling and coworkers and 50% of patients with MS utilizing Tourtellotte's formula. No correlation was established between serum and cerebrospinal fluid albumin concentrations, nor between serum albumin and IgG levels in cerebrospinal fluid. Thus, no evidence exists to support the inclusion of variations in serum albumin levels in the calculation of intrathecal IgG synthesis. Furthermore, Schuller and Sagar's formula allows classification of neuroimmunologic diseases and the evaluation of antibody-specific activity (ASA) in locally produced IgG. The comparison of intrathecal IgG ASA with serum IgG ASA, which reflects local and general immunity, respectively, seems potentially important.

Familial factors influence disability in MS multiplex families. French Multiple Sclerosis Genetics Group.

BACKGROUND: Both genetic and environmental factors play a role in the pathophysiology of MS and may influence the clinical expression of the disease. OBJECTIVE: To determine the contribution of familial factors to the clinical expression of MS. METHODS: The French Multiple Sclerosis Genetics Group identified 87 sibling pairs. For each patient, sex, age at onset, duration of the disease, and disease course from onset were recorded. Disability was determined by the progression index (PI), defined as the ratio of the Expanded Disability Status Scale (EDSS) score disease duration when the latter exceeded 5 years. Statistical analyses were performed either with a group of patients (clinical features, relation between human leukocyte antigen and clinical features) or with a group of sibpairs (concordance for clinical features). RESULTS: The mean age at onset was 29.6 years, the ratio of women to men was 59:28, and the mean PI was 0.27. There was no correlation for disease course and age at onset between sibs with MS. In contrast, we observed a weak but significant correlation of the PI in MS sibpairs (r = 0.234, p = 0.03). CONCLUSION: This study revealed a concordance in MS sibling pairs for the disease severity, supporting the hypothesis that the degree of disability might be partly influenced by familial factors (environmental or genetic).

A systematic study of oligodendrocyte growth factors as candidates for genetic susceptibility to MS. French Multiple Sclerosis Genetics Group.

OBJECTIVE: To test 23 genes coding for growth factors and their receptors as candidates for MS genetic susceptibility in 84 multiplex families of French origin by linkage analysis. BACKGROUND: Epidemiologic studies have indicated that genetic susceptibility in MS exists. To identify MS susceptibility genes, association and linkage studies were performed with candidate genes suggested by the pathology of MS. The most consistent result was genetic association and linkage of MS to human leukocyte antigen (HLA) DR15. Recent advances in the knowledge of MS pathology have suggested that the oligodendrocyte, the myelin-forming cell in the CNS, and its growth factors might play a crucial role in MS. METHODS: Fifty-two polymorphic markers within or flanking 23 candidate genes were used. Data were analyzed with the maximum likelihood score (MLS) approach. We also searched for a genetic interaction with HLA. RESULTS: Negative results were obtained for all candidate genes. The lower limits of the relative risk (Xs) possibly excluded for any candidate gene ranged from 1.3 to 2.8. Positive MLS values (up to 0.93) were observed for transforming growth factor beta 3 (TGFbeta3) in HLA DR15-associated families, suggesting a possible role for this growth factor in interaction with HLA. CONCLUSIONS: Oligodendrocyte growth factors do not play a significant role in MS genetic susceptibility, at least in the tested sample. TGFbeta3, the only gene highlighted by this study, deserves further analysis.

Determination of RNA antibodies in serum and unconcentrated CSF by counterimmunoelectrophoresis.

A quantitative counterimmunoelectrophoresis technique has been applied to evaluation of RNA antibodies in serum and unconcentrated CSF. Serum anti-RNA antibodies have been found at high dilutions in some patients with infectious diseases, subacute panencephalitis and multiple sclerosis. RNA antibodies in CSF are significantly linked to oligoclonal aspect, i.e. to a synthesis of IgG inside the CNS.

Determination of DNA antibodies in normal and pathological sera by a new counterimmunoelectrophoresis method.

A quantitative counter-immunoelectrophoresis technique has been applied to the evaluation of antibodies against native and single-stranded DNA. Anti-DNA antibodies have been found at high dilutions in patients with systematic lupus erythematosus, without correlation with the existence of renal lesions or with the degree of DNA binding assessed by Farr assay. Significant precipitates were also observed at significantly lower dilutions in other pathological situations and in normal subjects, posing the problem of the nature of the precipitates in these cases.

Exclusion of the candidate locus FSP1 in six families with late-onset autosomal dominant spastic paraplegia.

Hereditary spastic paraplegias are neurological hereditary conditions of unknown aetiology. In pure spastic paraplegia, most of the pedigrees display an autosomal dominant mode of inheritance. A gene for pure autosomal dominant spastic paraplegia (ADSP), termed FSP1, was mapped to chromosome 14q in a large pedigree with early-onset disease. This locus was tested by linkage analysis in six large French kindreds of ADSP with late-onset disease, using four microsatellites spanning a 9 cM interval including FSP1. FSP1 could be excluded in five of the six families, while no evidence for linkage was found in the remaining family. These results suggest that FSP1 is not involved in late onset ADSP, at least in the six families studied.

Cytokines in genetic susceptibility to multiple sclerosis: a candidate gene approach. French Multiple Sclerosis Genetics Group.

The immune system is involved in the pathophysiology of multiple sclerosis (MS) but the initiating antigen(s) is not yet identified. Since cytokines control both the intensity and the quality of the immune response they may be relevant candidates for the genetic susceptibility to MS. To analyze the contribution of type 1 and type 2 cytokine and cytokine receptor genes in the genetic susceptibility to MS, we have examined, in 116 French MS sibpairs, whether there is significant linkage between MS and 15 cytokine or cytokine receptor genes using 31 highly polymorphic genetic markers. The data were analyzed using the maximum likelihood score and the transmission disequilibrium approaches. None of the candidate genes tested was significantly linked to MS on the whole population. However, after stratification of the analysis on the basis of sharing (or not) of the HLA-DRB1*1501 allele, indication of linkage was found for the IL2-RB gene. These findings suggest that the IL2-RB locus contributes to the genetic susceptibility in a subgroup of MS patients.

Immunoglobulins and complement components in 37 patients infected by HIV-1 virus: comparison of general (systemic) and intrathecal immunity.

Intrathecal (IT) immunity was assessed by simultaneous analysis of paired cerebrospinal fluid (CSF) and sera of 37 patients infected by human immunodeficiency virus-1 (HIV-1). Only 8 of these 37 patients had no neurological or neuropsychiatric symptoms. There were 3 prominent abnormalities observed: (1) IT IgA production occurred in 15 patients, IT IgM production in 14 patients, and IT IgG production in 34 patients. (2) IT Anti-HIV-1 antibody specific activity (ASA) was higher than in serum in 33 of the 37 patients indicating that IT synthesis of antibody specific for HIV-1 occurs even in asymptomatic patients; IT anti-HIV-1 antibody synthesis was not correlated with clinical severity or neurological involvement. IT anti-herpes simplex ASA was also higher than serum ASA in 6 patients indicating a possible associated herpes simplex virus infection. (3) IT production of the complement component C4 was found frequently and was highly correlated with increased serum C4. IT C3 levels were decreased in 21 of 37 patients indicating that complement activation is a frequent accompaniment of the IT immune response in HIV-1-positive patients. These results indicate a unique and localized IT immune response which is different from the pattern observed in the systemic immune compartment in HIV-1-seropositive individuals and from the pattern common to the other CNS infectious diseases.

Conformational analysis of the amino termini (5 residues) of human glycophorin AM and AN: differentiation of the structural features of the TN and T antigenic determinants in relation to their specificity.

The N-terminus of glycophorin A, the main transmembrane erythrocyte glycoprotein responsible for the MN blood-group specificity, has been modelled. As the minimum size of the protein recognised by the antiglycophorin A antibodies is the N-terminal glycopentapeptide, attention was focused on the TN and T antigenic determinants of this size in order to determine wether differences in 3D structure exist and how a specific response with different antibodies is induced.

[The sterile woman, the physician and time. Number of spontaneous pregnancies in patients with functional sterility]. / La femme stérile, le médecin et le temps. Taux de grossesses spontanées chez des patientes atteintes de stérilité fonctionnelle.

We analysed a one to eight year follow-up of the women remaining infertile after a double-blind study of cyclofenil versus placebo to determine the frequency of pregnancy occurring independently of treatment. Pregnancy occurred in 64 women, 25 during treatment, 39 without treatment. We evaluated the effect of age, cause and duration of infertility on the pregnancy rate. The cumulative pregnancy rates at 36 months after the beginning of the study was 92 per cent for anovulatory cycles, 55 per cent for luteal insufficiency, 69 per cent for cervical mucus insufficiency and 49 per cent for idiopathic infertility. Of the 213 women entering the study, pregnancy occurred in 111 women, 47 in 3 cycles by a placebo effect, 39 without any treatment and 25 during a cycle with treatment. We conclude that the rate of treatment-independent pregnancy is high among infertile couples and that the two main therapeutic factors are time and the doctor-patient relationship.

[Sterility from hormonal causes and unexplained sterility. Treatment with cyclofenil. Double-blind controlled study]. / Stérilités de cause hormonale et stérilités inexpliquées. Traitement par le cyclofénil. Etude contrôlée à double insu.

We treated 213 women during 3 cycles by cyclofenil versus placebo in a double-blind study. These women had infertility caused by ovulatory deficiencies, a cervical factor or idiopathic infertility. Twenty-six of the 114 women receiving cyclofenil became pregnant and 21 of the 99 receiving the placebo. The cumulative pregnancy rates were identical in the 2 groups: 22 per cent in 3 cycles. The authors insist on the importance of a good doctor-patient relationship and the role of the placebo effect in any treatment of infertility.

[Breast cytology. Its place in the diagnosis of tumors]. / La cytologie mammaire. Sa place dans le diagnostic des lésions tumorales

Our work consists in the review of the experience in our hospital over 9 months and 110 cases in which tumours of the breast were seen. In most of them a clinical study, a radiological study, a thermographic study and a cytological and histological study was carried out. This shows that an examination of the ctyology is a very useful addition to the work-up to be carried out before therapy is started. The difficulty in cytology of the breast lies in the obviously serious problems in interpretation of the appearances, and explains why there are 2.7% of false negatives and 0.9% of false positives. All the same we think that we can find out how to lessen the number of these failures in diagnosis by a better knowledge of the traps to be found in breast cytology. Far from being a substitute for pathological anatomy, cytology is an indispensable complement to the study of the pathology of the breast.

[Peroperative use of Cell Saver in elective surgery on the abdominal aorta]. / Utilisation peropératoire du Cell Saver dans la chirurgie réglée de l'aorte abdominale.

In order to determine the impact of intraoperative autotransfusion on vascular surgical care, data related to 200 abdominal aortic surgical operations performed over a 20 months period were prospectively analysed. Volumes of blood salvaged and transfused during and after each operation were considered. One hundred and twenty one patients had an intraoperative autologous transfusion at a mean volume of 616 +/- 410 ml. Among them, 36 patients (43%) had only their own autotransfused blood and no other homologous blood components were required. Rapid autotransfusion was associated neither with significant haemolysis, nor with coagulopathy. Neither mortality nor morbidity was related to intraoperative autotransfusion. These data suggest that intraoperative autotransfusion is a safe replacement method in major vascular surgery. The procedure should be used in conjunction with preoperative donations when feasible.