Exosomes are defined as extracellular membrane vesicles, 30–150 nm in diameter, derived from all types of cells. They originate via endocytosis and then they are released through exocytosis to the extracellular space, being found in various biological fluids as well as in cell culture medium. In the last few years, exosomes have gained considerable scientific interest due to their potential use as biomarkers, especially in the field of cancer research. This report describes a method to isolate, quantify and identify serum- and cell culture-derived exosomes from dog samples, using small volumes (100 μL and 1 mL, respectively). Quantification and sizing of exosomes contained in serum and cell culture samples were assessed by utilizing nanoparticle tracking analysis, transmission electron microscopy and immunoelectron microscopy. Detected particles showed the normal size (30–150 nm) and morphology described for exosomes, as well as presence of the transmembrane protein CD63 known as exosomal marker. Based on a validated rapid isolation procedure of nanoparticles from small volumes of different types of dog samples, a characterization and exploration of intact exosomes, as well as facilitation for their analysis in downstream applications was introduced.

Pharmacological levels of zinc oxide can promote growth and health of weaning piglets, but the underlying molecular mechanisms are yet not fully understood. The aim of this study was to determine changes in the global hepatic protein expression in response to dietary zinc oxide in weaned piglets. Nine half-sib piglets were allocated to three dietary zinc treatment groups (50, 150, 2500 mg/kg dry matter). After 14 d, pigs were euthanized and liver samples taken. The increase in hepatic zinc concentration following dietary supplementation of zinc was accompanied by up-regulation of metallothionein mRNA and protein expression. Global hepatic protein profiles were obtained by two-dimensional difference gel electrophoresis following matrix-assisted laser desorption ionization/time-of-flight mass spectrometry. A total of 15 proteins were differentially (P<0.05) expressed between groups receiving control (150 mg/kg) or pharmacological levels of zinc (2500 mg/kg) with 7 down- (e.g. arginase1, thiosulfate sulfurtransferase, HSP70) and 8 up-regulated (e.g. apolipoprotein AI, transferrin, C1-tetrahydrofolate synthase) proteins. Additionally, three proteins were differentially expressed with low zinc supply (50 mg/kg Zn) in comparison to the control diet. The identified proteins were mainly associated with functions related to cellular stress, transport, metabolism, and signal transduction. The differential regulation was evaluated at the mRNA level and a subset of three proteins of different functional groups was selected for confirmation by western blotting. The results of this proteomic study suggest that zinc affects important liver functions such as blood protein secretion, protein metabolism, detoxification and redox homeostasis, thus supporting the hypothesis of intermediary effects of pharmacological levels of zinc oxide fed to pigs.

This review, which has been prepared within the frame of the European Union (EU)-funded project MARLON, surveys the organisation and characteristics of specific livestock and feed production chains (conventional, organic, GM-free) within the EU, with an emphasis on controls, regulations, traceability, and common production practices. Furthermore, an overview of the origin of animal feed used in the EU as well as an examination of the use of genetically modified organisms (GMOs) in feed is provided. From the data, it shows that livestock is traceable at the herd or individual level, depending on the species. Husbandry practices can vary widely according to geography and animal species, whilst controls and checks are in place for notifiable diseases and general health symptoms (such as mortality, disease, productive performance). For feeds, it would be possible only to make coarse estimates, at best, for the amount of GM feed ingredients that an animal is exposed to. Labeling requirements are apparently correctly followed. Provided that confounding factors are taken into account, practices such as organic agriculture that explicitly involve the use of non-GM feeds could be used for comparison to those involving the use of GM feed.

The mammalian oviduct provides the optimal environment for gamete maturation including sperm capacitation, fertilization, and development of the early embryo. Various cell culture models for primary bovine oviductal epithelial cells (BOEC) were established to reveal such physiological events. The aim of this study was to evaluate 17 candidate mRNA expression patterns in oviductal epithelial cells (1) in transition from in vivo cells to in vitro cells; (2) during three consecutive cell culture passages; (3) affected by the impact of LOW or HIGH glucose content media; and (4) influenced by different phases of the estrous cycle in vivo and in vitro. In addition, the release of a metabolite and proteins from BOEC at two distinct cell culture passage numbers was estimated to monitor the functionality. BOEC from 8 animals were isolated and cultured for three consecutive passages. Total RNA was extracted from in vivo and in vitro samples and subjected to reverse transcription quantitative polymerase chain reaction to reveal mRNA expression of selected candidate genes. The release of prostaglandin E2 (PGE2), oviduct-specific glycoprotein 1 (OVGP1) and interleukin 8 (IL8) by BOEC was measured by EIA or ELISA after 24 h. Almost all candidate genes (prostaglandin synthases, enzymes of cellular metabolism and mucins) mRNA expression pattern differed compared in vivo with in vitro state. In addition, transcription of most candidate genes was influenced by the number of cell culture passages. Different glucose medium content did not affect mRNA expression of most candidate genes. The phase of the estrous cycle altered some candidate mRNA expression in BOEC in vitro at later passages. The release of PGE2 and OVGP1 between passages did not differ. However, BOEC in passage 3 released significantly higher amount of IL8 compared with cells in passage 0. This study supports the hypothesis that candidate mRNA expression in BOEC was influenced by transition from the in vivo situation to the new in vitro environment and during consecutive passages. The consequence of cell culture passaging on BOEC ability to release bioactive compounds should be considered.

Formic acid (FA) has been used for decades to control Varroa destructor, one of the most important parasites of the western honey bee, Apis mellifera. The rather unselective molecular mode of action of FA and its possible effects on honeybees have long been a concern of beekeepers, as it has undesirable side effects that affect the health of bee colonies. This study focuses on short-term transcriptomic changes as analysed by RNAseq in both larval and adult honey bees and in mites after FA treatment under applied conditions. Our study aims to identify those genes in honey bees and varroa mites differentially expressed upon a typical FA hive exposure scenario. Five detoxification-related genes were identified with significantly enhanced and one gene with significantly decreased expression under FA exposure. Regulated genes in our test setting included members of various cytochrome P450 subfamilies, a flavin-dependent monooxygenase and a cytosolic 10-formyltetrahydrofolate dehydrogenase (FDH), known to be involved in formate metabolism in mammals. We were able to detect differences in the regulation of detoxification-associated genes between mites and honey bees as well as between the two different developmental stages of the honey bee. Additionally, we detected repressed regulation of Varroa genes involved in cellular respiration, suggesting mitochondrial dysfunction and supporting the current view on the mode of action of FA—inhibition of oxidative phosphorylation. This study shows distinct cellular effects induced by FA on the global transcriptome of both host and parasite in comparison. Our expression data might help to identify possible differences in the affected metabolic pathways and thus make a first contribution to elucidate the mode of detoxification of FA.