A gene encoding a germin-like protein,identified by a cDNA–AFLP approach,is specifically expressed during germination of<Emphasis Type="Italic"> Phaseolus vulgaris</Emphasis>

In order to identify markers of germination in Phaseolus vulgaris L., a cDNA-amplified fragment length polymorphism (AFLP) approach was conducted on mRNAs from embryo axes and from cotyledons. Among changes observed throughout the germination process, a cDNA fragment not detected 9 h after imbibition (HAI) but present specifically in axes 24 HAI was further studied. The complete cDNA was recovered by rapid amplification of cDNA ends, then cloned and sequenced. It includes an open reading frame predicting a 206-amino-acid polypeptide of 21.8 kDa. Analysis of the nucleotide sequence and deduced amino acid sequence revealed a high homology with germin-like proteins (GLPs), and particularly with an auxin-binding protein from peach, ABP19, that belongs to the GLP family. Thus, we propose that this cDNA encodes the first GLP described in P. vulgaris, designated PvGLP1. Northern blot analysis carried out on mRNAs from seed axes showed a dramatic increase in PvGLP1 expression a few hours before radicle emergence (17 HAI). Among mature vegetative tissues, PvGLP1 expression was very weak in pods and not detected in leaves, stems or roots. Immunoblot analysis using antibodies raised against AtGER3 from Arabidopsis thaliana showed that the protein could be detected only in axes from the dry seed stage onwards, at a steady-state level. Then, PvGLP1 expression seems to be associated with the early stages of embryo axis growth. The high homology indicated with ABP19 led us to study the effect of different concentrations of indole-3-acetic acid (IAA) on PvGLP1 expression during germination. Whereas no effect was noticed at low concentrations (1, 5, 10 microM), a marked decrease in PvGLP1 mRNA level was observed in axes of seeds imbibed with 100 microM IAA. Thus, PvGLP1 gene expression is not stimulated by auxin and, moreover, it might be inhibited by high concentrations of IAA.     

Comparison of the expression and activity of the lipogenic pathway in human and rat adipose tissue

Lipogenesis is considered less active in human than in rat adipose tissue. This could be explained by different nutritional conditions, namely high-carbohydrate (HCHO) diet in rats and high-fat (HF) diet in humans. Adipose tissue was sampled (postabsorptive state) in rats and humans receiving HCHO or HF diets, ad libitum fed humans, and obese subjects. We measured 1) mRNA concentrations of fatty acid synthase (FAS), acetyl-CoA carboxylase 1 (ACC1), sterol regulatory element binding protein 1c (SREBP-1c), and carbohydrate response element binding protein (ChREBP), 2) SREBP-1c protein, and 3) FAS activity. FAS, ACC1, ChREBP, and SREBP1-c mRNA concentrations were unaffected by diet in humans or in rats. FAS and ACC1 mRNA levels were lower in humans than in rats (P < 0.05). FAS activity was unaffected by diet and was lower in humans (P < 0.05). SREBP-1c mRNA concentrations were similar in rats and humans, but the precursor and mature forms of SREBP-1c protein were less abundant in humans (P < 0.05). ChREBP mRNA concentrations were lower in humans than in rats. In conclusion, the lipogenic capacity of adipose tissue is lower in humans than in rats. This is not related to differences in diet and is probably explained by lower abundance of SREBP-1c protein. A decreased expression of ChREBP could also play a role.     

Impact of feed supplementation with antimicrobial agents on growth performance of broiler chickens, Clostridium perfringens and enterococcus counts, and antibiotic resistance phenotypes and distribution of antimicrobial resistance determinants in Escherichia coli isolates

The effects of feed supplementation with the approved antimicrobial agents bambermycin, penicillin, salinomycin, and bacitracin or a combination of salinomycin plus bacitracin were evaluated for the incidence and distribution of antibiotic resistance in 197 commensal Escherichia coli isolates from broiler chickens over 35 days. All isolates showed some degree of multiple antibiotic resistance. Resistance to tetracycline (68.5%), amoxicillin (61.4%), ceftiofur (51.3%), spectinomycin (47.2%), and sulfonamides (42%) was most frequent. The levels of resistance to streptomycin, chloramphenicol, and gentamicin were 33.5, 35.5, and 25.3%, respectively. The overall resistance levels decreased from day 7 to day 35 (P < 0.001). Comparing treatments, the levels of resistance to ceftiofur, spectinomycin, and gentamicin (except for resistance to bacitracin treatment) were significantly higher in isolates from chickens receiving feed supplemented with salinomycin than from the other feeds (P < 0.001). Using a DNA microarray analysis capable of detecting commonly found antimicrobial resistance genes, we characterized 104 tetracycline-resistant E. coli isolates from 7- to 28-day-old chickens fed different growth promoters. Results showed a decrease in the incidence of isolates harboring tet(B), bla(TEM), sulI, and aadA and class 1 integron from days 7 to 35 (P < 0.01). Of the 84 tetracycline-ceftiofur-resistant E. coli isolates, 76 (90.5%) were positive for bla(CMY-2). The proportions of isolates positive for sulI, aadA, and integron class 1 were significantly higher in salinomycin-treated chickens than in the control or other treatment groups (P < 0.05). These data demonstrate that multiantibiotic-resistant E. coli isolates can be found in broiler chickens regardless of the antimicrobial growth promoters used. However, the phenotype and the distribution of resistance determinants in E. coli can be modulated by feed supplementation with some of the antimicrobial agents used in broiler chicken production.     

A Qualitative Study to Appraise Patients and Family Members Perceptions,Knowledge, and Attitudes towards Venous Thromboembolism Risk

Objective

This study aimed to examine perception, knowledge and concerns developed by patients and their family as regards venous thromboembolism (VTE) risk.

Methods

We conducted a qualitative study. Participants were: (1) patients with unprovoked VTE with either factor V Leiden mutation or G20210A prothrombin gene mutation or not; and (2) their first-degree relatives. Interviews took place mostly at Brest University Hospital. Participants produced narratives of the patient’s illness, stressing their perception of the disorder, its mechanisms, etiology, circumstances and risk factors. Interviews were audiotaped and transcribed verbatim. On an ongoing basis, central themes were identified and data from narratives were categorized by these themes.

Results

A total of ten patients and 25 first-degree relatives were interviewed. Analyses of patient’s narratives suggested 4 main themes: (1) concerns about initial symptoms and suspicion of VTE. The longer the duration of the initial phase, the more likely anxiety took place and persisted after diagnosis; (2) underestimation of potential life-threatening episode once being managed in emergency; (3) possible biographical disruption with inability to cope with the event; and (4) secondary prevention attitudes motivated by remains of the episode and favoring general prevention attitudes. Analyses of the first-degree relatives narratives suggested 3 main themes: (1) common interpretation of the VTE episode shared within the family; (2) diverse and sometimes confusing interpretation of the genetic status; and, (3) interpretation of clinical signs linked to VTE transmission within the family.

Conclusions

Construction of the risk of VTE is based on patient’s initial experience and shared within the family. Collection of narratives illustrates the gap between these perceptions and current medical knowledge. These results support the need to collect the perceptions of the VTE episode and its consequences, as a prerequisite to any health education process.     

Metabolic Outcome of Female Mice Exposed to a Mixture of Low-Dose Pollutants in a Diet-Induced Obesity Model

Pollutants are suspected to contribute to the etiology of obesity and related metabolic disorders. Apart from occupational exposure which concerns a subset of chemicals, humans are mostly exposed to a large variety of chemicals, all life-long and at low doses. Food ingestion is a major route of exposure and it is suggested that pollutants have a worsened impact when combined with a high-fat diet. In the experimental studies described herein, we aimed to add further evidence on the metabolic impact of food pollutants using a recently set up model in which mice are life-long fed a high-fat/high-sucrose diet (HFSD) with/without common food pollutants shown to exhibit metabolic disrupting activities. Specifically, this mixture comprised bisphenol A, dioxin, polychlorobiphenyl PCB153, and phthalate and was added in HFSD at doses resulting in mice exposure at the Tolerable Daily Intake dose range for each pollutant. We herein focused on the 7-week-old females which exhibited early signs of obesity upon HFSD feeding. We observed no signs of toxicity and no additional weight gain following exposure to the mixture but alleviated HFSD-induced glucose intolerance in the absence of alteration of gluconeogenesis and steatosis. It suggested that the observed metabolic improvement was more likely due to effects on muscle and/or adipose tissues rather than on the liver. Consistently, female mice exhibited enhanced lean/fat mass ratio and skeletal muscle insulin sensitivity. Moreover, expression levels of inflammatory markers were reduced in adipose tissue at 7 but enhanced at 12 weeks of age in agreement with the inverse alterations of glucose tolerance observed at these ages upon pollutant exposure in the HFSD-fed females. Collectively, these data suggest apparent biphasic effects of pollutants upon HFSD feeding along with obesity development. These effects were not observed in males and may depend on interactions between diet and pollutants.     

The role of breeding system on ant ecological dominance: genetic analysis of Ectatomma tuberculatum

Social insects exhibit a great variability in their social organization,and this affects colony kin structure, relatedness among nestmates, and population genetic structure. In the mosaic of arborealants of neotropical habitats, mutually exclusive dominant antspecies occupy different territories, and their nest distributionis spatially aggregated in patches influencing patterns of populationgenetic structure. In this study, we performed an analysis ofthe population and colony genetic structure of the facultativepolygynous ant Ectatomma tuberculatum to investigate how theparticular breeding and social system of this species can explainits ecological dominance in the mosaic. Within-nest geneticanalysis revealed that relatedness between nest mate workerswas significantly greater than zero (r = 0.30) with an effectivenumber of queens per nest of Ne = 2.5–3, indicating thatpolygyny is functional in this species. Moreover, we found thatqueen number was highly variable, probably due to queen adoptionevents, leading to the prevalence of polygyny over monogyny.Finally, the strong population genetic structure and the significantisolation by distance suggested that both budding and polydomytake place in this species. The respective role of secondarypolygyny, budding, and polydomy are then discussed in the contextof the mosaic of arboreal ants, and we propose that this particularsocial organization ensures the ecological dominance of E. tuberculatumby optimizing the colonization of new available nesting sitesand by increasing territory size.     

Induction of autoantibodies to murine P-glycoprotein: consequences on drug sensitivity in MDR cancer cells and on the expression of mdr genes in organs

Overexpression of the 170 kDa plasma membrane P-glycoprotein (P-gp) represents the most common MDR mechanism in chemotherapy. In this work, specific autoantibodies to fragments from extracellular loops 1, 2, and 4 of the murine MDR1 P-gp were elicited in mice using synthetic palmitoylated peptides reconstituted in liposomes and alum. The highest IgG level was observed after the third immunization and the immune response against lipopeptides was still detected more than 200 days after immunizations. Immunocytochemichal studies revealed that these antibodies were specific for P-gp. When incubated with P-gp-expressing MDR cell lines, serum from immunized mice restored sensitivity to either doxorubicin or vinblastine, or had no effect in a cell type specific manner, suggesting that several mechanisms may occur in the establishment of the MDR phenotype. The expression of mdr1 and mdr3 genes was unchanged in organs from mice immunized with palmitoylpeptides grafted on liposomes. These results suggest that the induction of autoantibodies to P-gp is a safe strategy to overcome MDR in cancer chemotherapy.     

Chondrocalcin is internalized by chondrocytes and triggers cartilage destruction via an interleukin-1β-dependent pathway

Chondrocalcin is among the most highly synthesized polypeptides in cartilage. This protein is released from its parent molecule, type II pro-collagen, after secretion by chondrocytes. A participation of extracellular, isolated chondrocalcin in mineralization was proposed more than 25 years ago, but never demonstrated. Here, exogenous chondrocalcin was found to trigger MMP13 secretion and cartilage destruction ex vivo in human cartilage explants and did so by modulating the expression of interleukin-1β in primary chondrocyte cultures in vitro. Chondrocalcin was found internalized by chondrocytes. Uptake was found mediated by a single 18-mer peptide of chondrocalcin, which does not exhibit homology to any known cell-penetrating peptide. The isolated peptide, when artificially linked as a tetramer, inhibited gene expression regulation by chondrocalcin, suggesting a functional link between uptake and gene expression regulation. At the same time, the tetrameric peptide potentiated chondrocalcin uptake by chondrocytes, suggesting a cooperative mechanism of entry. The corresponding peptide from type I pro-collagen supported identical cell-penetration, suggesting that this property may be conserved among C-propeptides of fibrillar pro-collagens. Structural modeling localized this peptide to the tips of procollagen C-propeptide trimers. Our findings shed light on unexpected function and mechanism of action of these highly expressed proteins from vertebrates.