Foodborne Cereulide Causes Beta-Cell Dysfunction and Apoptosis

<div><p>Aims/Hypothesis</p><p>To study the effects of cereulide, a food toxin often found at low concentrations in take-away meals, on beta-cell survival and function.</p><p>Methods</p><p>Cell death was quantified by Hoechst/Propidium Iodide in mouse (MIN6) and rat (INS-1E) beta-cell lines, whole mouse islets and control cell lines (HepG2 and COS-1). Beta-cell function was studied by glucose-stimulated insulin secretion (GSIS). Mechanisms of toxicity were evaluated in MIN6 cells by mRNA profiling, electron microscopy and mitochondrial function tests.</p><p>Results</p><p>24 h exposure to 5 ng/ml cereulide rendered almost all MIN6, INS-1E and pancreatic islets apoptotic, whereas cell death did not increase in the control cell lines. In MIN6 cells and murine islets, GSIS capacity was lost following 24 h exposure to 0.5 ng/ml cereulide (P<0.05). Cereulide exposure induced markers of mitochondrial stress including <i>Puma</i> (p53 up-regulated modulator of apoptosis, P<0.05) and general pro-apoptotic signals as <i>Chop</i> (CCAAT/-enhancer-binding protein homologous protein). Mitochondria appeared swollen upon transmission electron microscopy, basal respiration rate was reduced by 52% (P<0.05) and reactive oxygen species increased by more than twofold (P<0.05) following 24 h exposure to 0.25 and 0.50 ng/ml cereulide, respectively.</p><p>Conclusions/Interpretation</p><p>Cereulide causes apoptotic beta-cell death at low concentrations and impairs beta-cell function at even lower concentrations, with mitochondrial dysfunction underlying these defects. Thus, exposure to cereulide even at concentrations too low to cause systemic effects appears deleterious to the beta-cell.</p></div

Cereulide disrupts normal mitochondrial structure in MIN6 cells.

<p>Electron microscopic evaluation of MIN6 cells in control condition (A) or after 24 h exposure to 0.5 ng/ml cereulide (B) (magnification 2500x). Arrows indicate normal mitochondria in control condition (A) or enlarged mitochondria with less cristae after cereulide exposure (B). Morphometric analysis confirmed fewer mitochondria (D) and swelling of the remaining mitochondria (E) in the exposed cells, as is calculated by dividing the mitochondrial area by the number of mitochondria. In the cereulide exposed condition, we noted cells with chromatin condensation (indicated by CC in panel 4C), compared to normal nuclei (indicated by NC in panel A). After cereulide exposure there were more autophagic vacuoles (F, indicated by white arrow in panel C) Data are presented as mean ± SEM. * P<0.05 vs control; *** P<0.001 vs control (student t test).</p

Cereulide negatively affects mitochondrial function in MIN6 cells after 24 h exposure.

<p>Oxygen consumption rate in basal conditions was reduced (A, n = 7). Cereulide caused a rise in reactive oxygen species, as shown by the increase in dichlorofluorescindiacetate fluorescence (B, n = 8). Cereulide exposure caused cytochrome c release into the cytoplasm (C, n = 4). Data are presented as mean ± SEM. * P<0.05 vs control, **P<0.01 vs control.</p

Cereulide causes apoptotic cell death in 3 beta-cell models, but not in other mammalian cell lines after 24 h exposure.

<p>Cell death rate was determined with Hoechst-PI staining for MIN6 cells (A, n = 6), mouse islets (B, n = 3), INS-1E (C, n = 3), COS-1 (D, n = 3) and HepG2 (E, n = 3), as well as caspase 3/7 activation (F, n = 3) in MIN6 cells after 24 h exposure to cereulide. Data are presented as mean ± SEM. * P<0.05, ** P<0.01 *** P<0.001 vs control.</p

Cereulide impairs glucose-stimulated insulin secretion.

<p>MIN6 cells (A, n = 11) and whole mouse islets (B, n = 3) were exposed to cereulide for 24 h, and incubated with KREBS containing low (white bars) or high (black bars) concentrations of glucose. Insulin secretion is expressed as a percentage of total insulin content. Data are presented as mean ± SEM. * P<0.05, *** P<0.001 vs control (2 way ANOVA, followed by Bonferroni test for MIN6; student t test for islets).</p

Silicone Wristband Passive Samplers Yield Highly Individualized Pesticide Residue Exposure Profiles

Monitoring human exposure to pesticides and pesticide residues (PRs) remains crucial for informing public health policies, despite strict regulation of plant protection product and biocide use. We used 72 low-cost silicone wristbands as noninvasive passive samplers to assess cumulative 5-day exposure of 30 individuals to polar PRs. Ethyl acetate extraction and LC-MS/MS analysis were used for the identification of PRs. Thirty-one PRs were detected of which 15 PRs (48%) were detected only in worn wristbands, not in environmental controls. The PRs included 16 fungicides (52%), 8 insecticides (26%), 2 herbicides (6%), 3 pesticide derivatives (10%), 1 insect repellent (3%), and 1 pesticide synergist (3%). Five detected pesticides were not approved for plant protection use in the EU. Smoking and dietary habits that favor vegetable consumption were associated with higher numbers and higher cumulative concentrations of PRs in wristbands. Wristbands featured unique PR combinations. Our results suggest both environment and diet contributed to PR exposure in our study group. Silicone wristbands could serve as sensitive passive samplers to screen population-wide cumulative dietary and environmental exposure to authorized, unauthorized and banned pesticides

mRNA levels of intrinsic mitochondrial apoptosis and ER stress mediators are upregulated in MIN6 cells after 24 h of exposure to cereulide.

<p>Death protein 5 (<i>Dp5</i>, n = 4, A) and p53 upregulated modulator of apoptosis (<i>Puma</i>, n = 8, B) are dose dependently upregulated, as are activating transcription factor 4 (<i>Atf4</i>, n = 8, C) and CCAAT/-enhancer-binding protein homologous protein (<i>Chop</i>, n = 8, D), while binding immunoglobulin protein (<i>Bip</i>, n = 8, E) and spliced X-box binding protein are not (<i>Xbp1s</i>, n = 8, F). Data are presented as mean ± SEM (n 4–8). * P<0.05 vs control, *** P<0.001 vs control.</p

6‑Alkoxy-5-aryl-3-pyridinecarboxamides, a New Series of Bioavailable Cannabinoid Receptor Type 1 (CB1) Antagonists Including Peripherally Selective Compounds

We identified 6-alkoxy-5-aryl-3-pyridinecarboxamides as potent CB1 receptor antagonists with high selectivity over CB2 receptors. The series was optimized to reduce lipophilicity compared to rimonabant to achieve peripherally active molecules with minimal central effects. Several compounds that showed high plasma exposures in rats were evaluated in vivo to probe the contribution of central vs peripheral CB1 agonism to metabolic improvement. Both rimonabant and <b>14g</b>, a potent brain penetrant CB1 receptor antagonist, significantly reduced the rate of body weight gain. However, <b>14h</b>, a molecule with markedly reduced brain exposure, had no significant effect on body weight. PK studies confirmed similarly high exposure of both <b>14h</b> and <b>14g</b> in the periphery but 10-fold lower exposure in the brain for <b>14h</b>. On the basis of these data, which are consistent with reported effects in tissue-specific CB1 receptor KO mice, we conclude that the metabolic benefits of CB1 receptor antagonists are primarily centrally mediated as originally believed

Additional file 1: of A phase III randomized trial of gantenerumab in prodromal Alzheimer’s disease

List of institutional review boards and independent ethics committees. (DOCX 20 kb