Abstract The ecotoxicity of three different sizes of titanium dioxide (TiO2 ) particles (primary particles sizes: 10, 30, and 300 nm) to the freshwater green alga Pseudokirchneriella subcapitata was ...investigated in this study. Algal growth inhibition was found for all three particle types, but the physiological mode of action is not yet clear. It was possible to establish a concentration/dose–response relationship for the three particle sizes. Reproducibility, however, was affected by concentration-dependent aggregation of the nanoparticles, subsequent sedimentation, and possible attachment to vessel surfaces. It is also believed that heteroaggregation, driven by algal exopolymeric exudates, is occurring and could influence the concentration–response relationship. The ecotoxicity of cadmium to algae was investigated both in the presence and absence of 2 mg/L TiO2 . The presence of TiO2 in algal tests reduced the observed toxicity due to decreased bioavailability of cadmium resulting from sorption/complexation of Cd2+ ions to the TiO2 surface. However, for the 30 nm TiO2 nanoparticles, the observed growth inhibition was greater than what could be explained by the concentration of dissolved Cd(II) species, indicating a possible carrier effect, or combined toxic effect of TiO2 nanoparticles and cadmium. These results emphasize the importance of systematic studies of nanoecotoxicological effects of different sizes of nanoparticles and underline the fact that, in addition to particle toxicity, potential interactions with existing environmental contaminants are also of crucial importance in assessing the potential environmental risks of nanoparticles.
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Alternative processing of the precursor protein pro-GIP results in endogenously produced GIP(1–30)NH2, that by DPP-4 cleavage in vivo results in the metabolite GIP(3–30)NH2. We showed ...previously that GIP(3–30)NH2 is a high affinity antagonist of the human GIPR in vitro. Here we determine whether it is suitable for studies of GIP physiology in rats since effects of GIP agonists and antagonists are strictly species-dependent. Transiently transfected COS-7 cells were assessed for cAMP accumulation upon ligand stimulation or assayed in competition binding using human 125I-GIP(1–42) as radioligand. In isolated perfused rat pancreata, insulin, glucagon, and somatostatin-releasing properties were evaluated. Competition binding demonstrated that on the rat GIP receptor (GIPR), rat GIP(3–30)NH2 bound with high affinity (Ki of 17nM), in contrast to human GIP(3–30)NH2 (Ki of 250nM). In cAMP studies, rat GIP(3–30)NH2 inhibited GIP(1–42)-induced rat GIPR activation and schild-plot analysis showed competitive antagonism with a pA2 of 13nM and a slope of 0.9±0.09. Alone, rat GIP(3–30)NH2 displayed weak, low-potent partial agonistic properties (EC50>1μM) with an efficacy of 9.4% at 0.32μM compared to GIP(1–42). In perfused rat pancreata, rat GIP(3–30)NH2 efficiently antagonized rat GIP(1–42)-induced insulin, somatostatin, and glucagon secretion. In summary, rat GIP(3–30)NH2 is a high affinity competitive GIPR antagonist and effectively antagonizes GIP-mediated G protein-signaling as well as pancreatic hormone release, while human GIP(3–30)NH2, despite a difference of only one amino acid between the two (arginine in position 18 in rat GIP(3–30)NH2; histidine in human), is unsuitable in the rat system. This underlines the importance of species differences in the GIP system, and the limitations of testing human peptides in rodent systems.
Background and Purpose
Specific, high potency receptor antagonists are valuable tools when evaluating animal and human physiology. Within the glucose‐dependent, insulinotropic polypeptide (GIP) ...system, considerable attention has been given to the presumed GIP receptor antagonist, (Pro3)GIP, and its effect in murine studies. We conducted a pharmacological analysis of this ligand including interspecies differences between the rodent and human GIP system.
Experimental Approach
Transiently transfected COS‐7 cells were assessed for cAMP accumulation upon ligand stimulation and assayed in competition binding using 125I‐human GIP. Using isolated perfused pancreata both from wild type and GIP receptor‐deficient rodents, insulin‐releasing, glucagon‐releasing and somatostatin‐releasing properties in response to species‐specific GIP and (Pro3)GIP analogues were evaluated.
Key Results
Human (Pro3)GIP is a full agonist at human GIP receptors with similar efficacy (Emax) for cAMP production as human GIP, while both rat and mouse(Pro3)GIP were partial agonists on their corresponding receptors. Rodent GIPs are more potent and efficacious at their receptors than human GIP. In perfused pancreata in the presence of 7 mM glucose, both rodent (Pro3)GIP analogues induced modest insulin, glucagon and somatostatin secretion, corresponding to the partial agonist activities observed in cAMP production.
Conclusions and Implications
When evaluating new compounds, it is important to consider interspecies differences both at the receptor and ligand level. Thus, in rodent models, human GIP is a comparatively weak partial agonist. Human (Pro3)GIP was not an antagonist at human GIP receptors, so there is still a need for a potent antagonist in order to elucidate the physiology of human GIP.
Background and Purpose
Glucose‐dependent insulinotropic polypeptide (GIP) affects lipid, bone and glucose homeostasis. High‐affinity ligands for the GIP receptor are needed to elucidate the ...physiological functions and pharmacological potential of GIP in vivo. GIP(1–30)NH2 is a naturally occurring truncation of GIP(1–42). Here, we have characterized eight N‐terminal truncations of human GIP(1–30)NH2.
Experimental Approach
COS‐7 cells were transiently transfected with human GIP receptors and assessed for cAMP accumulation upon ligand stimulation or competition binding with 125I‐labelled GIP(1–42), GIP(1–30)NH2, GIP(2–30)NH2 or GIP(3–30)NH2.
Key Results
GIP(1–30)NH2 displaced 125I‐GIP(1–42) as effectively as GIP(1–42) (Ki 0.75 nM), whereas the eight truncations displayed lower affinities (Ki 2.3–347 nM) with highest affinities for GIP(3–30)NH2 and GIP(5–30)NH2 (5–30)NH2. Only GIP(1–30)NH2 (Emax 100% of GIP(1–42)) and GIP(2–30)NH2 (Emax 20%) were agonists. GIP(2‐ to 9–30)NH2 displayed antagonism (IC50 12–450 nM) and Schild plot analyses identified GIP(3–30)NH2 and GIP(5–30)NH2 as competitive antagonists (Ki 15 nM). GIP(3–30) NH2 was a 26‐fold more potent antagonist than GIP(3–42). Binding studies with agonist (125I‐GIP(1–30)NH2), partial agonist (125I‐GIP(2–30)NH2) and competitive antagonist (125I‐GIP(3–30)NH2) revealed distinct receptor conformations for these three ligand classes.
Conclusions and Implications
The N‐terminus is crucial for GIP agonist activity. Removal of the C‐terminus of the endogenous GIP(3–42) creates another naturally occurring, more potent, antagonist GIP(3–30)NH2, which like GIP(5–30)NH2, was a high‐affinity competitive antagonist. These peptides may be suitable tools for basic GIP research and future pharmacological interventions.
Nanomaterials (NMs) display many unique and useful physico-chemical properties. However, reliable approaches are needed for risk assessment of NMs. The present study was performed in the FP7-MARINA ...project, with the objective to identify and evaluate in vitro test methods for toxicity assessment in order to facilitate the development of an intelligent testing strategy (ITS). Six representative oxide NMs provided by the EC-JRC Nanomaterials Repository were tested in nine laboratories. The in vitro toxicity of NMs was evaluated in 12 cellular models representing 6 different target organs/systems (immune system, respiratory system, gastrointestinal system, reproductive organs, kidney and embryonic tissues). The toxicity assessment was conducted using 10 different assays for cytotoxicity, embryotoxicity, epithelial integrity, cytokine secretion and oxidative stress. Thorough physico-chemical characterization was performed for all tested NMs. Commercially relevant NMs with different physico-chemical properties were selected: two TiO2 NMs with different surface chemistry - hydrophilic (NM-103) and hydrophobic (NM-104), two forms of ZnO - uncoated (NM-110) and coated with triethoxycapryl silane (NM-111) and two SiO2 NMs produced by two different manufacturing techniques - precipitated (NM-200) and pyrogenic (NM-203). Cell specific toxicity effects of all NMs were observed; macrophages were the most sensitive cell type after short-term exposures (24-72h) (ZnO>SiO2>TiO2). Longer term exposure (7 to 21 days) significantly affected the cell barrier integrity in the presence of ZnO, but not TiO2 and SiO2, while the embryonic stem cell test (EST) classified the TiO2 NMs as potentially 'weak-embryotoxic' and ZnO and SiO2 NMs as 'non-embryotoxic'. A hazard ranking could be established for the representative NMs tested (ZnO NM-110 > ZnO NM-111 > SiO2 NM-203 > SiO2 NM-200 > TiO2 NM-104 > TiO2 NM-103). This ranking was different in the case of embryonic tissues, for which TiO2 displayed higher toxicity compared with ZnO and SiO2. Importantly, the in vitro methodology applied could identify cell- and NM-specific responses, with a low variability observed between different test assays. Overall, this testing approach, based on a battery of cellular systems and test assays, complemented by an exhaustive physico-chemical characterization of NMs, could be deployed for the development of an ITS suitable for risk assessment of NMs. This study also provides a rich source of data for modeling of NM effects.
Developments in nanotechnology are leading to a rapid proliferation of new materials that are likely to become a source of engineered nanoparticles (ENPs) to the environment, where their possible ...ecotoxicological impacts remain unknown. The surface properties of ENPs are of essential importance for their aggregation behavior, and thus for their mobility in aquatic and terrestrial systems and for their interactions with algae, plants and, fungi. Interactions of ENPs with natural organic matter have to be considered as well, as those will alter the ENPs aggregation behavior in surface waters or in soils. Cells of plants, algae, and fungi possess cell walls that constitute a primary site for interaction and a barrier for the entrance of ENPs. Mechanisms allowing ENPs to pass through cell walls and membranes are as yet poorly understood. Inside cells, ENPs might directly provoke alterations of membranes and other cell structures and molecules, as well as protective mechanisms. Indirect effects of ENPs depend on their chemical and physical properties and may include physical restraints (clogging effects), solubilization of toxic ENP compounds, or production of reactive oxygen species. Many questions regarding the bioavailability of ENPs, their uptake by algae, plants, and fungi and the toxicity mechanisms remain to be elucidated.
Exaggerated postprandial secretion of glucagon-like peptide-1 (GLP-1) and peptide YY (PYY) may explain appetite reduction and weight loss after Roux-en-Y gastric bypass (RYGB), but causality has not ...been established. We hypothesized that food intake decreases after surgery through combined actions from GLP-1 and PYY. GLP-1 actions can be blocked using the GLP-1 receptor antagonist Exendin 9-39 (Ex-9), whereas PYY actions can be inhibited by the administration of a dipeptidyl peptidase-4 (DPP-4) inhibitor preventing the formation of PYY
.
Appetite-regulating gut hormones and appetite ratings during a standard mixed-meal test and effects on subsequent ad libitum food intake were evaluated in two studies: in study 1, nine patients with type 2 diabetes were examined prospectively before and 3 months after RYGB with and without Ex-9. In study 2, 12 RYGB-operated patients were examined in a randomized, placebo-controlled, crossover design on four experimental days with: (1) placebo, (2) Ex-9, (3) the DPP-4 inhibitor, sitagliptin, to reduce formation of PYY
and (4) Ex-9/sitagliptin combined.
In study 1, food intake decreased by 35% following RYGB compared with before surgery. Before surgery, GLP-1 receptor blockage increased food intake but no effect was seen postoperatively, whereas PYY secretion was markedly increased. In study 2, combined GLP-1 receptor blockage and DPP-4 inhibitor mediated lowering of PYY
increased food intake by ~20% in RYGB patients, whereas neither GLP-1 receptor blockage nor DPP-4 inhibition alone affected food intake, perhaps because of concomitant marked increases in the unblocked hormone.
Blockade of actions from only one of the two L-cell hormones, GLP-1 and PYY
, resulted in concomitant increased secretion of the other, probably explaining the absent effect on food intake on these experimental days. Combined blockade of GLP-1 and PYY actions increased food intake after RYGB, supporting that these hormones have a role in decreased food intake postoperatively.
Poor adherence to complex multimodal therapies is a widely recognized problem in the daily care of dialysis patients, contributing to excess morbidity and mortality of this population. While a few ...studies have been devoted to understanding patient nonadherence, their results were somewhat controversial. The goals of this review are to quantify nonadherence to certain oral medications, to raise awareness of factors that may cause problems in a patient;s adherence to this treatment, and to describe strategies that may be used to improve adherence to prescribed pharmacotherapy.
A systematic literature review in the MEDLINE and PubMed database (1971-2008) was performed. Quantitative studies, which accurately indicated the total percentages of nonadherence to oral medication in adult patients receiving chronic hemodialysis, were identified.
A total of 19 studies fulfilled the search criteria. Rates of nonadherence to the oral medication ranged from 3 - 80%. More than half of the included studies reported nonadherence rates of > or = 50% (mean 67%). The use of phosphate binding therapy was the prevalent surveyed oral medication. Self reports, structured interviews, and predialysis serum phosphate levels were the most frequent assessment tools used to record adherence rates. Limitations of the reviewed studies included small patient cohorts, inconsistent definitions of adherence, and a lack of standardized methods for measuring nonadherence.
Nonadherence to oral medication in hemodialysis patients is still an underestimated, but life-threatening behaviour.
Microplastics (MPs) are omnipresent in our surroundings and in the environment, with drinking water being a potential pathway for human exposure. This study investigated the presence of MPs in Danish ...drinking water from 17 different households and workplaces in Denmark. Samples of tap water were collected using a closed sampling system to decrease airborne contamination, and QA/QC measurements were performed to assess background contamination. Particles > 100 µm were visually analysed by stereomicroscopy in combination with spectroscopy analysis (µ-FTIR) to evaluate morphology and chemical composition. An assessment of MP particles down to 10 µm was performed on water samples from three locations using hyperspectral image analysis. The results indicate a low level of MPs in Danish drinking water, with a total of seven MP particles across all samples, comprising PET, PP, PS, and ABS. Microfibers were the most common type of MP-like particles in both drinking water and blanks, but the concentration for all samples was below the limit of detection and could not be differentiated from background contamination. Most of the particles analysed by µ-FTIR were identified as cellulose fibres and a smaller subset as protein. Based on this work, we discuss the status of MP drinking water studies and address challenges and limitations regarding the analysis of MP in drinking water.
Aims
To evaluate the performances of commercially available glucagon‐like peptide‐1 (GLP‐1) assays and the implications for clinical studies.
Methods
Known concentrations (5–300 pmol/l) of synthetic ...GLP‐1 isoforms (GLP‐1 1‐36NH2, 7‐36NH2, 9‐36NH2, 1‐37, 7‐37 and 9‐37) were added to the matrix (assay buffer) supplied with 10 different kits and to human plasma, and recoveries were determined. Assays yielding meaningful results were analysed for precision and sensitivity by repeated analysis and ability to discriminate low concentrations. Endogenous GLP‐1 levels in clinical samples were assessed using three commercial kits.
Results
The USCN LIFE assay detected none of the GLP‐1 isoforms. The active GLP‐1 enzyme‐linked immunosorbent assays (ELISAs) from Millipore and DRG appeared identical and were specific for intact GLP‐1 in buffer and plasma. The Meso Scale Discovery (MSD) total GLP‐1 kit detected all six GLP‐1 isoforms, although recovery of non‐active forms was incomplete, especially in plasma. Millipore total GLP‐1 ELISA kit detected all isoforms in buffer, but mainly amidated forms in plasma. The Alpco, Phoenix and Bio‐Rad kits detected only amidated GLP‐1, but the Alpco kit had a limited measurement range (30 pmol/l), the Phoenix kit had incomplete recovery in plasma and the Bio‐Rad kit was insensitive (detection limit in plasma 40 pmol/l). The pattern of postprandial GLP‐1 responses in clinical samples was similar between the kits tested, but the absolute concentrations measured varied.
Conclusions
The specificity and sensitivity of commercially available kits for the analysis of GLP‐1 levels vary considerably. This should be taken into account when selecting which assay to use and when comparing data from different studies.
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