RasGAP-derived fragment N increases the resistance of beta cells towards apoptosis in NOD mice and delays the progression from mild to overt diabetes.

The caspase-3-generated RasGAP N-terminal fragment (fragment N) inhibits apoptosis in a Ras-PI3K-Akt-dependent manner. Fragment N protects various cell types, including insulin-secreting cells, against different types of stresses. Whether fragment N exerts a protective role during the development of type 1 diabetes is however not known. Non-obese diabetic (NOD) mice represent a well-known model for spontaneous development of type 1 diabetes that shares similarities with the diseases encountered in humans. To assess the role of fragment N in type 1 diabetes development, a transgene encoding fragment N under the control of the rat insulin promoter (RIP) was back-crossed into the NOD background creating the NOD-RIPN strain. Despite a mosaic expression of fragment N in the beta cell population of NOD-RIPN mice, islets isolated from these mice were more resistant to apoptosis than control NOD islets. Islet lymphocytic infiltration and occurrence of a mild increase in glycemia developed with the same kinetics in both strains. However, the period of time separating the mild increase in glycemia and overt diabetes was significantly longer in NOD-RIPN mice compared to the control NOD mice. There was also a significant decrease in the number of apoptotic beta cells in situ at 16 weeks of age in the NOD-RIPN mice. Fragment N exerts therefore a protective effect on beta cells within the pro-diabetogenic NOD background and this prevents a fast progression from mild to overt diabetes.

SOCS-1 protein prevents Janus Kinase/STAT-dependent inhibition of beta cell insulin gene transcription and secretion in response to interferon-gamma.

In the pathogenesis of type I diabetes mellitus, activated leukocytes infiltrate pancreatic islets and induce beta cell dysfunction and destruction. Interferon (IFN)-gamma, tumor necrosis factor-alpha and interleukin (IL)-1 beta play important, although not completely defined, roles in these mechanisms. Here, using the highly differentiated beta Tc-Tet insulin-secreting cell line, we showed that IFN-gamma dose- and time-dependently suppressed insulin synthesis and glucose-stimulated secretion. As described previously IFN-gamma, in combination with IL-1 beta, also induces inducible NO synthase expression and apoptosis (Dupraz, P., Cottet, S., Hamburger, F., Dolci, W., Felley-Bosco, E., and Thorens, B. (2000) J. Biol. Chem. 275, 37672--37678). To assess the role of the Janus kinase/signal transducer and activator of transcription (STAT) pathway in IFN-gamma intracellular signaling, we stably overexpressed SOCS-1 (suppressor of cytokine signaling-1) in the beta cell line. We demonstrated that SOCS-1 suppressed cytokine-induced STAT-1 phosphorylation and increased cellular accumulation. This was accompanied by a suppression of the effect of IFN-gamma on: (i) reduction in insulin promoter-luciferase reporter gene transcription, (ii) decrease in insulin mRNA and peptide content, and (iii) suppression of glucose-stimulated insulin secretion. Furthermore, SOCS-1 also suppressed the cellular effects that require the combined presence of IL-1 beta and IFN-gamma: induction of nitric oxide production and apoptosis. Together our data demonstrate that IFN-gamma is responsible for the cytokine-induced defect in insulin gene expression and secretion and that this effect can be completely blocked by constitutive inhibition of the Janus kinase/STAT pathway.

Sensitivity of predictive species distribution models to change in grain size

Predictive species distribution modelling (SDM) has become an essential tool in biodiversity conservation and management. The choice of grain size (resolution) of environmental layers used in modelling is one important factor that may affect predictions. We applied 10 distinct modelling techniques to presence-only data for 50 species in five different regions, to test whether: (1) a 10-fold coarsening of resolution affects predictive performance of SDMs, and (2) any observed effects are dependent on the type of region, modelling technique, or species considered. Results show that a 10 times change in grain size does not severely affect predictions from species distribution models. The overall trend is towards degradation of model performance, but improvement can also be observed. Changing grain size does not equally affect models across regions, techniques, and species types. The strongest effect is on regions and species types, with tree species in the data sets (regions) with highest locational accuracy being most affected. Changing grain size had little influence on the ranking of techniques: boosted regression trees remain best at both resolutions. The number of occurrences used for model training had an important effect, with larger sample sizes resulting in better models, which tended to be more sensitive to grain. Effect of grain change was only noticeable for models reaching sufficient performance and/or with initial data that have an intrinsic error smaller than the coarser grain size.

Human invariant V alpha 24-J alpha Q TCR supports the development of CD1d-dependent NK1.1+ and NK1.1- T cells in transgenic mice.

A sizable fraction of T cells expressing the NK cell marker NK1.1 (NKT cells) bear a very conserved TCR, characterized by homologous invariant (inv.) TCR V alpha 24-J alpha Q and V alpha 14-J alpha 18 rearrangements in humans and mice, respectively, and are thus defined as inv. NKT cells. Because human inv. NKT cells recognize mouse CD1d in vitro, we wondered whether a human inv. V alpha 24 TCR could be selected in vivo by mouse ligands presented by CD1d, thereby supporting the development of inv. NKT cells in mice. Therefore, we generated transgenic (Tg) mice expressing the human inv. V alpha 24-J alpha Q TCR chain in all T cells. The expression of the human inv. V alpha 24 TCR in TCR C alpha(-/-) mice indeed rescues the development of inv. NKT cells, which home preferentially to the liver and respond to the CD1d-restricted ligand alpha-galactosylceramide (alpha-GalCer). However, unlike inv. NKT cells from non-Tg mice, the majority of NKT cells in V alpha 24 Tg mice display a double-negative phenotype, as well as a significant increase in TCR V beta 7 and a corresponding decrease in TCR V beta 8.2 use. Despite the forced expression of the human CD1d-restricted TCR in C alpha(-/-) mice, staining with mCD1d-alpha-GalCer tetramers reveals that the absolute numbers of peripheral CD1d-dependent T lymphocytes increase at most by 2-fold. This increase is accounted for mainly by an increased fraction of NK1.1(-) T cells that bind CD1d-alpha-GalCer tetramers. These findings indicate that human inv. V alpha 24 TCR supports the development of CD1d-dependent lymphocytes in mice, and argue for a tight homeostatic control on the total number of inv. NKT cells. Thus, human inv. V alpha 24 TCR-expressing mice are a valuable model to study different aspects of the inv. NKT cell subset.

The changing preference of T and B cells for partners as T-dependent antibody responses develop.

Recirculating virgin CD4+ T cells spend their life migrating between the T zones of secondary lymphoid tissues where they screen the surface of interdigitating dendritic cells. T-cell priming starts when processed peptides or superantigen associated with class II MHC molecules are recognised. Those primed T cells that remain within the lymphoid tissue move to the outer T zone, where they interact with B cells that have taken up and processed antigen. Cognate interaction between these cells initiates immunoglobulin (Ig) class switch-recombination and proliferation of both B and T cells; much of this growth occurs outside the T zones B cells migrate to follicles, where they form germinal centres, and to extrafollicular sites of B-cell growth, where they differentiate into mainly short-lived plasma cells. T cells do not move to the extrafollicular foci, but to the follicles; there they proliferate and are subsequently involved in the selection of B cells that have mutated their Ig variable-region genes. During primary antibody responses T-cell proliferation in follicles produces many times the peak number of T cells found in that site: a substantial proportion of the CD4+ memory T-cell pool may originate from growth in follicles.

The average crossing number of equilateral random polygons.

In this paper, we study the average crossing number of equilateral random walks and polygons. We show that the mean average crossing number ACN of all equilateral random walks of length n is of the form . A similar result holds for equilateral random polygons. These results are confirmed by our numerical studies. Furthermore, our numerical studies indicate that when random polygons of length n are divided into individual knot types, the for each knot type can be described by a function of the form where a, b and c are constants depending on and n0 is the minimal number of segments required to form . The profiles diverge from each other, with more complex knots showing higher than less complex knots. Moreover, the profiles intersect with the ACN profile of all closed walks. These points of intersection define the equilibrium length of , i.e., the chain length at which a statistical ensemble of configurations with given knot type -upon cutting, equilibration and reclosure to a new knot type -does not show a tendency to increase or decrease . This concept of equilibrium length seems to be universal, and applies also to other length-dependent observables for random knots, such as the mean radius of gyration Rg.

Metabolic effects of diets differing in glycaemic index depend on age and endogenous glucose-dependent insulinotrophic polypeptide in mice.

AIMS/HYPOTHESIS: High- vs low-glycaemic index (GI) diets unfavourably affect body fat mass and metabolic markers in rodents. Different effects of these diets could be age-dependent, as well as mediated, in part, by carbohydrate-induced stimulation of glucose-dependent insulinotrophic polypeptide (GIP) signalling. METHODS: Young-adult (16 weeks) and aged (44 weeks) male wild-type (C57BL/6J) and GIP-receptor knockout (Gipr ( -/- )) mice were exposed to otherwise identical high-carbohydrate diets differing only in GI (20-26 weeks of intervention, n = 8-10 per group). Diet-induced changes in body fat distribution, liver fat, locomotor activity, markers of insulin sensitivity and substrate oxidation were investigated, as well as changes in the gene expression of anorexigenic and orexigenic hypothalamic factors related to food intake. RESULTS: Body weight significantly increased in young-adult high- vs low-GI fed mice (two-way ANOVA, p < 0.001), regardless of the Gipr genotype. The high-GI diet in young-adult mice also led to significantly increased fat mass and changes in metabolic markers that indicate reduced insulin sensitivity. Even though body fat mass also slightly increased in high- vs low-GI fed aged wild-type mice (p < 0.05), there were no significant changes in body weight and estimated insulin sensitivity in these animals. However, aged Gipr ( -/- ) vs wild-type mice on high-GI diet showed significantly lower cumulative net energy intake, increased locomotor activity and improved markers of insulin sensitivity. CONCLUSIONS/INTERPRETATION: The metabolic benefits of a low-GI diet appear to be more pronounced in younger animals, regardless of the Gipr genotype. Inactivation of GIP signalling in aged animals on a high-GI diet, however, could be beneficial.

Ageing-related cardiomyocyte functional decline is sex and angiotensin II dependent.

Clinically, heart failure is an age-dependent pathological phenomenon and displays sex-specific characteristics. The renin-angiotensin system mediates cardiac pathology in heart failure. This study investigated the sexually dimorphic functional effects of ageing combined with angiotensin II (AngII) on cardiac muscle cell function, twitch and Ca(2+)-handling characteristics of isolated cardiomyocytes from young (~13 weeks) and aged (~87 weeks) adult wild type (WT) and AngII-transgenic (TG) mice. We hypothesised that AngII-induced contractile impairment would be exacerbated in aged female cardiomyocytes and linked to Ca(2+)-handling disturbances. AngII-induced cardiomyocyte hypertrophy was evident in young adult mice of both sexes and accentuated by age (aged adult ~21-23 % increases in cell length relative to WT). In female AngII-TG mice, ageing was associated with suppressed cardiomyocyte contractility (% shortening, maximum rate of shortening, maximum rate of relaxation). This was associated with delayed cytosolic Ca(2+) removal during twitch relaxation (Tau ~20 % increase relative to young adult female WT), and myofilament responsiveness to Ca(2+) was maintained. In contrast, aged AngII-TG male cardiomyocytes exhibited peak shortening equivalent to young TG; yet, myofilament Ca(2+) responsiveness was profoundly reduced with ageing. Increased pro-arrhythmogenic spontaneous activity was evident with age and cardiac AngII overexpression in male mice (42-55 % of myocytes) but relatively suppressed in female aged transgenic mice. Female myocytes with elevated AngII appear more susceptible to an age-related contractile deficit, whereas male AngII-TG myocytes preserve contractile function with age but exhibit desensitisation of myofilaments to Ca(2+) and a heightened vulnerability to arrhythmic activity. These findings support the contention that sex-specific therapies are required for the treatment of age-progressive heart failure.

Tissue-specific and SRSF1-dependent splicing of fibronectin, a matrix protein that controls host cell invasion.

Cell invasion targets specific tissues in physiological placental implantation and pathological metastasis, which raises questions about how this process is controlled. We compare dermis and endometrium capacities to support trophoblast invasion, using matching sets of human primary fibroblasts in a coculture assay with human placental explants. Substituting endometrium, the natural trophoblast target, with dermis dramatically reduces trophoblast interstitial invasion. Our data reveal that endometrium expresses a higher rate of the fibronectin (FN) extra type III domain A+ (EDA+) splicing isoform, which displays stronger matrix incorporation capacity. We demonstrate that the high FN content of the endometrium matrix, and not specifically the EDA domain, supports trophoblast invasion by showing that forced incorporation of plasma FN (EDA-) promotes efficient trophoblast invasion. We further show that the serine/arginine-rich protein serine/arginine-rich splicing factor 1 (SRSF1) is more highly expressed in endometrium and, using RNA interference, that it is involved in the higher EDA exon inclusion rate in endometrium. Our data therefore show a mechanism by which tissues can be distinguished, for their capacity to support invasion, by their different rates of EDA inclusion, linked to their SRSF1 protein levels. In the broader context of cancer pathology, the results suggest that SRSF1 might play a central role not only in the tumor cells, but also in the surrounding stroma.

Prevalence of social phobia in a clinical sample of drug dependent patients.

INTRODUCTION: Social phobia is among the most frequent psychiatric disorders and can be classified into two subtypes, nongeneralized and generalized. Whereas it significantly worsens the morbidity of comorbid substance abuse disorders, and it often is associated with reduced treatment responses, there is still lacking data on its prevalence in clinical populations of drug abusing patients. METHODS: The study sample consisted of 75 inpatients and 75 outpatients meeting DSM-IV criteria for drug dependence. Symptoms of social phobia were assessed with the French-language version of the Liebowitz Social Anxiety Scale (LSAS). RESULTS: Prevalence rate were 20% for the generalized subtype and 42.6% for the nongeneralized subtype. Gender difference emerged in the severity of fear, women reporting significantly greater fear relating to performance situations than men. CONCLUSIONS: An important proportion of patients with substance dependence present a comorbid generalized or nongeneralized social phobia. Early recognition of social phobia and adequate interventions is warranted for these patients in order to improve their treatment response with regard to quality of life and relapse prevention.

Education of Murine NK Cells Requires Both cis and trans Recognition of MHC Class I Molecules.

Although NK cells use invariant receptors to identify diseased cells, they nevertheless adapt to their environment, including the presence of certain MHC class I (MHC-I) molecules. This NK cell education, which is mediated by inhibitory receptors specific for MHC-I molecules, changes the responsiveness of activating NK cell receptors (licensing) and modifies the repertoire of MHC-I receptors used by NK cells. The fact that certain MHC-I receptors have the unusual capacity to recognize MHC-I molecules expressed by other cells (trans) and by the NK cell itself (cis) has raised the question regarding possible contributions of the two types of interactions to NK cell education. Although the analysis of an MHC-I receptor variant suggested a role for cis interaction for NK cell licensing, adoptive NK cell transfer experiments supported a key role for trans recognition. To reconcile some of these findings, we have analyzed the impact of cell type-specific deletion of an MHC-I molecule and of a novel MHC-I receptor variant on the education of murine NK cells when these mature under steady-state conditions in vivo. We find that MHC-I expression by NK cells (cis) and by T cells (trans), and MHC-I recognition in cis and in trans, are both needed for NK cell licensing. Unexpectedly, modifications of the MHC-I receptor repertoire are chiefly dependent on cis binding, which provides additional support for an essential role for this unconventional type of interaction for NK cell education. These data suggest that two separate functions of MHC-I receptors are needed to adapt NK cells to self-MHC-I.

What was your fracture risk evaluated by FRAX(®) the day before your osteoporotic fracture?

Osteoporotic fracture (OF) is one of the major causes of morbidity and mortality in industrialized countries. Switzerland is among the countries with the greatest risk. Our aim was (1) to calculate the FRAX(®) in a selected Swiss population the day before the occurrence of an OF and (2) to compare the results with the proposed Swiss FRAX(®) thresholds. The Swiss Association Against Osteoporosis proposed guidelines for the treatment of osteoporosis based on age-dependent thresholds. To identify a population at a very high risk of osteoporotic fracture, we included all consecutive patients in the active OF pathway cohort from the Lausanne University Hospital, Switzerland. FRAX(®) was calculated with the available data the day before the actual OF. People with a FRAX(®) body mass index (BMI) or a FRAX(®) (bone mineral density) BMD lower than the Swiss thresholds were not considered at high risk. Two-hundred thirty-seven patients were included with a mean age of 77.2 years, and 80 % were female. Major types of fracture included hip (58 %) and proximal humerus (25 %) fractures. Mean FRAX(®) BMI values were 28.0, 10.0, 13.0, 26.0, and 37.0 % for age groups 50-59, 60-69, 70-79, and 80-89 years old, respectively. Fifty percent of the population was not considered at high risk by the FRAX(®) BMI. FRAX(®) BMD was available for 95 patients, and 45 % had a T score < -2.5 standard deviation. Only 30 % of patients with a normal or osteopenic BMD were classified at high risk by FRAX(®) BMD. The current proposed Swiss thresholds were not able to classify at high risk in 50 to 70 % of the studied population the day before a major OF.

Interleukin-8 secretion by fibroblasts induced by low density lipoproteins is p38 MAPK-dependent and leads to cell spreading and wound closure.

We have previously reported (Dobreva, I., Waeber, G., Mooser, V., James, R. W., and Widmann, C. (2003) J. Lipid Res. 44, 2382-2390) that low density lipoproteins (LDLs) induce activation of the p38 MAPK pathway, resulting in fibroblast spreading and lamellipodia formation. Here, we show that LDL-stimulated fibroblast spreading and wound sealing are due to secretion of a soluble factor. Using an antibody-based human protein array, interleukin-8 (IL-8) was identified as the main cytokine whose concentration was increased in supernatants from LDL-stimulated cells. Incubation of supernatants from LDL-treated cells with an anti-IL-8 blocking antibody completely abolished their ability to induce cell spreading and mediate wound closure. In addition, fibroblasts treated with recombinant IL-8 spread to the same extent as cells incubated with LDL or supernatants from LDL-treated cells. The ability of LDL and IL-8 to induce fibroblast spreading was mediated by the IL-8 receptor type II (CXCR-2). Furthermore, LDL-induced IL-8 production and subsequent wound closure required the activation of the p38 MAPK pathway, because both processes were abrogated by a specific p38 inhibitor. Therefore, the capacity of LDLs to induce fibroblast spreading and accelerate wound closure relies on their ability to stimulate IL-8 secretion in a p38 MAPK-dependent manner. Regulation of fibroblast shape and migration by lipoproteins may be relevant to atherosclerosis that is characterized by increased LDL cholesterol levels, IL-8 production, and extensive remodeling of the vessel wall.