Some features about transposition of the maize elementDissociation inNicotiana plumbaginifolia

We have recently shown that a plasmid-borneDissociation (Ds) element can excise from extrachromosomal plasmid DNA and integrate into a plant genome in the presence of theActivator (Ac) transposase.Ds andAc-carrying plasmids were used to co-transformNicotiana plumbaginifolia protoplasts. Transgenic plants were regenerated and analyzed. Here we describe further characterization of the system and discuss its efficiency in terms of DNA transformation and transposon tagging.     

Protocadherin-12 cleavage is a regulated process mediated by ADAM10 protein: evidence of shedding up-regulation in pre-eclampsia

Protocadherins are a group of transmembrane proteins with homophilic binding activity, members of the cadherin superfamily. Apart from their role in adhesion, the cellular functions of protocadherins are essentially unknown. Protocadherin (PCDH)12 was previously identified in invasive trophoblasts and endothelial and mesangial cells in the mouse. Invalidation studies revealed that the protein was required for optimal placental development. In this article, we show that its human homolog is abundantly expressed in various trophoblast subtypes of the human placenta and at lower levels in endothelial cells. We demonstrate that PCDH12 is shed at high rates in vitro. The shedding mechanism depends on ADAM10 and results in reduced cellular adhesion in a cell migration assay. PCDH12 is subsequently cleaved by the γ-secretase complex, and its cytoplasmic domain is rapidly degraded by the proteasome. PCDH12 shedding is regulated by interlinked intracellular pathways, including those involving protein kinase C, PI3K, and cAMP, that either increase or inhibit cleavage. In endothelial cells, VEGF, prostaglandin E(2), or histamine regulates PCDH12 shedding. The extracellular domain of PCDH12 was also detected in human serum and urine, thus providing evidence of PCDH12 shedding in vivo. Importantly, we observed an increase in circulating PCDH12 in pregnant women who later developed a pre-eclampsia, a frequent pregnancy syndrome and a major cause of maternal and fetal morbidity and mortality. In conclusion, we speculate that, like in mice, PCDH12 may play an important role in human placental development and that proteolytic cleavage in response to external factors, such as cytokines and pathological settings, regulates its activity.     

Nitric Oxide Contributes to Hypoxia-Reoxygenation-Induced P-Glycoprotein Expression in Rat Brain Endothelial Cells

Ischemia–reperfusion leads to increased levels at the blood–brain barrier of the multidrug efflux transporter, P-glycoprotein that provides protection to the brain by limiting access of unwanted substances. This is coincident with the production of nitric oxide. This present study using immortalized rat brain endothelial cells (GPNTs) examines whether following hypoxia-reoxygenation, nitric oxide contributes to the alterations in P-glycoprotein levels. After 6 h of hypoxia, both nitric oxide and reactive oxygen species, detected intracellularly using fluorescent monitoring dyes, were produced in the subsequent reoxygenation phase coincident with increased P-glycoprotein. The evidence that nitric oxide can directly affect P-glycoprotein expression was sought by applying S-nitroso-N-acetyl-dl-penicillamine that as shown increased the nitric oxide generation. Sodium nitroprusside, though more effective at increasing P-glycoprotein expression, appeared to produce different reactive species. Real time RT-PCR analysis revealed the predominant form of nitric oxide synthase in these cells to be endothelial, inhibition of which partially prevented the increase in P-glycoprotein during reoxygenation. These data indicate that the production of nitric oxide by endothelial nitric oxide synthase during reoxygenation can influence P-glycoprotein expression in cells of the blood-rat brain barrier, highlighting another route by which nitric oxide may protect the brain.     

An Automated Blur Detection Method for Histological Whole Slide Imaging

Whole slide scanners are novel devices that enable high-resolution imaging of an entire histological slide. Furthermore, the imaging is achieved in only a few minutes, which enables image rendering of large-scale studies involving multiple immunohistochemistry biomarkers. Although whole slide imaging has improved considerably, locally poor focusing causes blurred regions of the image. These artifacts may strongly affect the quality of subsequent analyses, making a slide review process mandatory. This tedious and time-consuming task requires the scanner operator to carefully assess the virtual slide and to manually select new focus points. We propose a statistical learning method that provides early image quality feedback and automatically identifies regions of the image that require additional focus points.     

Life expectancy estimates as a key factor in over-treatment: The case of prostate cancer

Objective: To estimate the magnitude of over-diagnosis and of potential and actual over-treatment regarding prostate cancer, taking comorbidities into account. Materials and methods: We used a sample collected by the French cancer registries of 1840 cases (T1: 583; T2: 1257) diagnosed in 2001. The proportion of over-diagnosed and over-treated patients was estimated by comparing life expectancy (LE), including or not comorbidities, with natural LE with cancer, using several assumptions from the literature. We distinguished potential and actual over-treatment according to the treatment that patients actually received. Results: Among patients with T1 tumors the proportion of potential over-treatment using LE adjusted for comorbidity varied from 29.5% to 53.5%, using LE adjusted on comorbidities, and varied from 9.3% to 22.2% regarding actual over-treatment. Between 7.7% and 24.4% of patient's receiving a radical prostatectomy, and between 30.8% and 62.5% of those receiving radiotherapy, were over-treated. Among patients with T2 tumors, the proportions of potential and actual over-treatment were 0.9% and 2.0%. Two per cent of patients receiving a radical prostatectomy and 4.9% of those receiving radiotherapy were over-treated. Comorbidities dramatically increased these proportions to nearly 100% of patients, with more than two comorbidities being potentially over-treated and around 33% actually over-treated. Conclusions: According to the French incidence, 3200–4800 French patients may be over-treated, among whom a large proportion of patients had comorbidities. The real issue is to offer the most appropriate treatment to people with low-grade tumors and comorbidities.     

Immunogenicity of Live Attenuated B. pertussis BPZE1 Producing the Universal Influenza Vaccine Candidate M2e

Background

Intranasal delivery of vaccines directed against respiratory pathogens is an attractive alternative to parenteral administration. However, using this delivery route for inactivated vaccines usually requires the use of potent mucosal adjuvants, and no such adjuvant has yet been approved for human use.

Methodology/Principal Findings

We have developed a live attenuated Bordetella pertussis vaccine, called BPZE1, and show here that it can be used to present the universal influenza virus epitope M2e to the mouse respiratory tract to prime for protective immunity against viral challenge. Three copies of M2e were genetically fused to the N-terminal domain of filamentous hemagglutinin (FHA) and produced in recombinant BPZE1 derivatives in the presence or absence of endogenous full-length FHA. Only in the absence of FHA intranasal administration of the recombinant BPZE1 derivative induced antibody responses to M2e and effectively primed BALB/c mice for protection against influenza virus-induced mortality and reduced the viral load after challenge. Strong M2e-specific antibody responses and protection were observed after a single nasal administration with the recombinant BPZE1 derivative, followed by a single administration of M2e linked to a virus-like particle without adjuvant, whereas priming alone with the vaccine strain did not protect.

Conclusions/Significance

Using recombinant FHA-3M2e-producing BPZE1 derivatives for priming and the universal influenza M2e peptide linked to virus-like particles for boosting may constitute a promising approach for needle-free and adjuvant-free nasal vaccination against influenza.     

A Liver-Specific Defect of Acyl-CoA Degradation Produces Hyperammonemia,Hypoglycemia and a Distinct Hepatic Acyl-CoA Pattern

Most conditions detected by expanded newborn screening result from deficiency of one of the enzymes that degrade acyl-coenzyme A (CoA) esters in mitochondria. The role of acyl-CoAs in the pathophysiology of these disorders is poorly understood, in part because CoA esters are intracellular and samples are not generally available from human patients. We created a mouse model of one such condition, deficiency of 3-hydroxy-3-methylglutaryl-CoA lyase (HL), in liver (HLLKO mice). HL catalyses a reaction of ketone body synthesis and of leucine degradation. Chronic HL deficiency and acute crises each produced distinct abnormal liver acyl-CoA patterns, which would not be predictable from levels of urine organic acids and plasma acylcarnitines. In HLLKO hepatocytes, ketogenesis was undetectable. Carboxylation of [2-¹⁴C] pyruvate diminished following incubation of HLLKO hepatocytes with the leucine metabolite 2-ketoisocaproate (KIC). HLLKO mice also had suppression of the normal hyperglycemic response to a systemic pyruvate load, a measure of gluconeogenesis. Hyperammonemia and hypoglycemia, cardinal features of many inborn errors of acyl-CoA metabolism, occurred spontaneously in some HLLKO mice and were inducible by administering KIC. KIC loading also increased levels of several leucine-related acyl-CoAs and reduced acetyl-CoA levels. Ultrastructurally, hepatocyte mitochondria of KIC-treated HLLKO mice show marked swelling. KIC-induced hyperammonemia improved following administration of carglumate (N-carbamyl-L-glutamic acid), which substitutes for the product of an acetyl-CoA-dependent reaction essential for urea cycle function, demonstrating an acyl-CoA-related mechanism for this complication.     

Comparative effect of inorganic N on plant growth and N<Subscript>2</Subscript> fixation of ten legume crops: towards a better understanding of the differential response among species

Aims

A better understanding of how plant growth, N nutrition and symbiotic nitrogen fixation (SNF) are influenced by soil inorganic N availability, for a wide range of legume species, is crucial to optimise legume productivity, N₂ fixation, while limiting environmental risks such as N leaching.

Methods

A comparative analysis was performed for ten legume crops, grown in a field experiment and supplied with four N fertiliser rates. Dry matter, N concentration and SNF were measured. In parallel, root elongation rates were studied in a greenhouse experiment.

Results

For most species, N fertilisation had little effect on plant growth and N accumulation. SNF was reduced by soil inorganic N available at sowing but with large differences in the magnitude of the response among species. The response varied according to plant N requirements for growth and plant ability to retrieve inorganic N. Accordingly, root lateral expansion rate measured in RhizoTubes was highly correlated with plant ability to retrieve inorganic N measured in the field experiment.

Conclusion

Combining SNF response to soil inorganic N, shoot N and plant ability to retrieve inorganic N, allowed a robust evaluation of differential response to soil inorganic N among a wide range of legume species.