Complementary adhesion molecules promote neutrophil-Kupffer cell interaction and the elimination of bacteria taken up by the liver.

Most bacteria that enter the bloodstream are taken up by the liver. Previously, we reported that such organisms are initially bound extracellularly and subsequently killed by immigrating neutrophils, not Kupffer cells as widely presumed in the literature. Rather, the principal functions of Kupffer cells demonstrated herein are to clear bacteria from the peripheral blood and to promote accumulation of bactericidal neutrophils at the principal site of microbial deposition in the liver, i.e., the Kupffer cell surface. In a mouse model of listeriosis, uptake of bacteria by the liver at 10 min postinfection i.v. was reduced from approximately 60% of the inoculum in normal mice to approximately 15% in mice rendered Kupffer cell deficient. Immunocytochemical analysis of liver sections derived from normal animals at 2 h postinfection revealed the massive immigration of neutrophils and their colocalization with Kupffer cells. Photomicrographs of the purified nonparenchymal liver cell population derived from these infected mice demonstrated listeriae inside neutrophils and neutrophils within Kupffer cells. Complementary adhesion molecules promoted the interaction between these two cell populations. Pretreatment of mice with mAbs specific for CD11b/CD18 (type 3 complement receptor) or its counter-receptor, CD54, inhibited the accumulation of neutrophils in the liver and the elimination of listeriae. Complement was not a factor; complement depletion affected neither the clearance of listeriae by Kupffer cells nor the antimicrobial activity expressed by infiltrating neutrophils.     

Metal ion-mobilizing additives for comprehensive detection of femtomole amounts of phosphopeptides by reversed phase LC-MS

It is hypothesized that metal ion-mediated adsorption of phosphorylated peptides on stationary phases of LC-columns is the major cause for their frequently observed poor detection efficiency in LC-MS. To study this phenomenon in more detail, sample solutions spiked with metal ion-mobilizing additives were analyzed by reversed phase μLC-ICP-MS or nanoLC-ESI-MS. Using μLC-ICP-MS, metal ions were analyzed directly as atomic ions. Using electrospray ionization, either metal ion chelates or phosphopeptide standard mixtures injected in subpicomole amounts were analyzed. Deferoxamine, imidazole, ascorbate, citrate, EDTA, and the tetrapeptide pSpSpSpS were tested as sample additives for the interlinked purposes of metal ion-mobilization and improvement of phosphopeptide recovery. Iron probably represents the major metal ion contamination of reversed phase columns. Based on the certified iron level in LC-grade solvents, a daily metal ion load of >10 pmol was estimated for typical nanoLC flow rates. In addition, phosphopeptide fractions from IMAC columns were identified as source for metal ion contamination of the LC column, as demonstrated for Ga³⁺-IMAC. The three metal ion-chelating additives, EDTA, citrate and pSpSpSpS, were found to perform best for improving the LC recovery of multiply phosphorylated peptides injected at subpicomole amounts. The benefits of metal ion-mobilizing LC (mimLC) characterized by metal ion complexing sample additives is demonstrated for three different instrumental setups comprising (a) a nanoUPLC-system with direct injection on the analytical column, (b) a nanoLC system with inclusion of a trapping column, and (c) the use of a HPLC-Chip system with integrated trapping and analytical column.     

Cases Concerning Sovereignty over Islands before the International Court of Justice and the Dokdo/Takeshima Issue

The article analyzes the recent jurisprudence of the International Court of Justice in relation to disputes concerning sovereignty over islands and considers the potential implications of this jurisprudence for the resolution of the Dokdo/Takeshima issue. It does so by examining the principles and rules of international law applied by the Court to the determination of title to territory, especially those concerning the question of original title and its interplay with state conduct in general (effectivités). The article also pays special attention to the different legal techniques applied by the Court in resolving each particular dispute and the Court's practical approach to dealing with questions of historical facts and other evidentiary matters in relation to small and uninhabited islands.     

Saponin production is not qualitatively changed upon callus regeneration in the medicinal shrub Maesa perlarius

Maesa perlarius is a medicinal plant that produces maesabalides, which possess selective and strong anti-leishmania activity. In this study, M. perlarius plants were regenerated from leaf-derived calli. Shoots were induced in Murashige and Skoog medium in the presence of thidiazuron in combination with α-naphthalene acetic acid. In contrast to seed-derived plants, callus-derived regenerants were tetraploid showing typical characteristics of higher ploidy phenotypes. We assessed the impact of indirect plant regeneration and associated increase in ploidy on the production of saponin by means of LC–MS analysis. Tetraploid M. perlarius produce a saponin profile, which was not significantly different from seed grown plants. Based on this study, we concluded that saponin production in M. perlarius is not qualitatively changed by a genome-doubling event.     

Separating Drought Effects from Roof Artifacts on Ecosystem Processes in a Grassland Drought Experiment

1

Given the predictions of increased drought probabilities under various climate change scenarios, there have been numerous experimental field studies simulating drought using transparent roofs in different ecosystems and regions. Such roofs may, however, have unknown side effects, called artifacts, on the measured variables potentially confounding the experimental results. A roofed control allows the quantification of potential artifacts, which is lacking in most experiments.

2

We conducted a drought experiment in experimental grasslands to study artifacts of transparent roofs and the resulting effects of artifacts on ecosystems relative to drought on three response variables (aboveground biomass, litter decomposition and plant metabolite profiles). We established three drought treatments, using (1) transparent roofs to exclude rainfall, (2) an unroofed control treatment receiving natural rainfall and (3) a roofed control, nested in the drought treatment but with rain water reapplied according to ambient conditions.

3

Roofs had a slight impact on air (+0.14°C during night) and soil temperatures (−0.45°C on warm days, +0.25°C on cold nights), while photosynthetically active radiation was decreased significantly (−16%). Aboveground plant community biomass was reduced in the drought treatment (−41%), but there was no significant difference between the roofed and unroofed control, i.e., there were no measurable roof artifact effects.

4

Compared to the unroofed control, litter decomposition was decreased significantly both in the drought treatment (−26%) and in the roofed control treatment (−18%), suggesting artifact effects of the transparent roofs. Moreover, aboveground metabolite profiles in the model plant species Medicago x varia were different from the unroofed control in both the drought and roofed control treatments, and roof artifact effects were of comparable magnitude as drought effects.

5

Our results stress the need for roofed control treatments when using transparent roofs for studying drought effects, because roofs can cause significant side effects.     

Metformin Protects Rat Hepatocytes against Bile Acid-Induced Apoptosis

Background

Metformin is used in the treatment of Diabetes Mellitus type II and improves liver function in patients with non-alcoholic fatty liver disease (NAFLD). Metformin activates AMP-activated protein kinase (AMPK), the cellular energy sensor that is sensitive to changes in the AMP/ATP-ratio. AMPK is an inhibitor of mammalian target of rapamycin (mTOR). Both AMPK and mTOR are able to modulate cell death.

Aim

To evaluate the effects of metformin on hepatocyte cell death.

Methods

Apoptotic cell death was induced in primary rat hepatocytes using either the bile acid glycochenodeoxycholic acid (GCDCA) or TNFα in combination with actinomycin D (actD). AMPK, mTOR and phosphoinositide-3 kinase (PI3K)/Akt were inhibited using pharmacological inhibitors. Apoptosis and necrosis were quantified by caspase activation, acridine orange staining and Sytox green staining respectively.

Results

Metformin dose-dependently reduces GCDCA-induced apoptosis, even when added 2 hours after GCDCA, without increasing necrotic cell death. Metformin does not protect against TNFα/ActD-induced apoptosis. The protective effect of metformin is dependent on an intact PI3-kinase/Akt pathway, but does not require AMPK/mTOR-signaling. Metformin does not inhibit NF-κB activation.

Conclusion

Metformin protects against bile acid-induced apoptosis and could be considered in the treatment of chronic liver diseases accompanied by inflammation.     

PCR-verified microarray analysis and functional in vitro studies indicate a role of α-tocopherol in vesicular transport

Global gene expression profiles of livers from mice, fed diets differing in α-tocopherol content, were compared using DNA microarray technology. Three hundred and eighty nine genes were found to significantly differ in their expression level by a factor of 2 or higher between the high and the low α-tocopherol group. Functional clustering using the EASE software identified 121 genes involved in transport processes. Twenty-one thereof were involved in (synaptic) vesicular trafficking. Up-regulation of syntaxin 1C (Stx1c), vesicle-associated membrane protein 1 (Vamp1), N-ethylmaleimide-sensitive factor (Nsf) and syntaxin binding protein 1 (Stxbp1, Munc18-1) was verified by real time PCR. At a functional level, α-tocopherol increased the secretory response in RBL and PC12 cells. Although here detected in liver, the α-tocopherol-responsive pathways are also relevant to neurotransmission. A role of α-tocopherol in the vesicular transport might not only affect its own absorption and transport but also explain the neural dysfunctions observed in severe α-tocopherol deficiency.     

The effects of manufacturing tolerances and assembly force on the volumetric wear at the taper junction in modular total hip arthroplasty

Abstract

Fretting and corrosion at the taper-head interface in total hip arthroplasty has been reported as a potential cause of early failure of the implant system. The finite element (FE) method can be used to study the mechanics at the taper junction that are difficult to assess experimentally. Taper mismatch is one of the factors that can influence the performance of the taper junction. In this study we have assessed the effect of taper mismatch, in combination with assembly force on the volumetric wear. The study showed that higher assembly forces and smaller mismatches result in the least volumetric wear.