High-level dietary cadmium exposure is associated with global DNA hypermethylation in the gastropod hepatopancreas

5-methylcytosine (5mC) is a key epigenetic mark which influences gene expression and phenotype. In vertebrates, this epigenetic mark is sensitive to Cd exposure, but there is no information linking such an event with changes in global 5mC levels in terrestrial gastropods despite their importance as excellent ecotoxicological bioindicators of metal contamination. Therefore, we first evaluated total 5mC content in DNA of the hepatopancreas of adult Cantareus aspersus with the aim to determine whether this epigenetic mark is responsive to Cd exposure. The experiment was conducted under laboratory conditions and involved a continuous exposure, multiple dose- and time-point (14, 28, and 56 days) study design. Hepatopancreas cadmium levels were measured using Flame Atomic Absorption Spectrometry and the percentage of 5-mC in samples using an ELISA-based colorimetric assay. Snail death rates were also assessed. Our results, for the first time, reveal the presence of 5mC in C. aspersus and provide evidence for Cd-induced changes in global 5mC levels in DNA of gastropods and mollusks. Although less sensitive than tissue accumulation, DNA methylation levels responded in a dose- and time-dependent manner to dietary cadmium, with exposure dose having a much stronger effect than exposure duration. An obvious trend of increasing 5mC levels was observed starting at 28 days of exposure to the second highest dose and this trend persisted at the two highest treatments for close to one month, when the experiment was terminated after 56 days. Moreover, a strong association was identified between Cd concentrations in the hepatopancreas and DNA methylation levels in this organ. These data indicate an overall trend towards DNA hypermethylation with elevated Cd exposure. No consistent lethal effect was observed, irrespective of time point and Cd-dosage. Overall, our findings suggest that the total 5mC content in DNA of the hepatopancreas of land snails is responsive to sublethal Cd exposure and give new insights into invertebrate environmental epigenetics

Short-term effects of very low dose cadmium feeding on copper, manganese and iron homeostasis:A gastropod perspective

The available information on the interplay between low-dose cadmium intake and copper, manganese, and iron homeostasis in invertebrates is limited. We have currently studied the accumulation of these trace metals in the hepatopancreas of adult snails, Cantareus aspersus, following 14 and 28 days of exposure to low doses of dietary cadmium, up to 1 mg/kg dw (dry weight). The cadmium dose, but not the duration of exposure, had a significant effect on hepatopancreas copper deposition, the values being significantly elevated compared to controls. A significant peak in manganese levels at 14 days was found in snails administered the lowest cadmium dose. These increases occurred even in the absence of cadmium increase in the hepatopancreas. Our data suggest that low dose cadmium feeding can produce a transient disturbance in hepatopancreas copper and manganese homeostasis. Such responses may serve as early biomarkers of physiological changes occurring during the initial stages of cadmium intoxication

Effect of dietary cadmium on genome-wide 5mC levels in DNA of the hepatopancreas of <i>C</i>. <i>aspersus</i> adults.

<p>(<b>A</b>) The mean contents of 5mC in hepatopancreas DNA in each treatment group at each time point. (<b>B</b>) Estimated marginal means for dietary dose (as the main effect). (<b>C</b>) Estimated marginal means for time (as the main effect). Four snails were sampled for each treatment group at each time point (14, 28, and 56 days). Data are shown on a log₁₀ scale as mean (point) with one standard error (box) and one standard deviation (error bar). In Fig 2A the outliers (white circles) and the extremes (whiskers) are also given (if they exist). Marked boxes(*) indicate significant differences as compared to the reference group (Newman-Keuls test, ***—<i>p</i> < 0.001, **—<i>p</i> < 0.01, *—<i>p</i> < 0.05).</p

Survival times and mortalities for each time period.

<p>Survival times and mortalities for each time period.</p

Effects of Quercetin on Blood Pressure: A Systematic Review and Meta‐Analysis of Randomized Controlled Trials

BACKGROUND: Quercetin, the most abundant dietary flavonol, has antioxidant effects in cardiovascular disease, but the evidence regarding its effects on blood pressure (BP) has not been conclusive. We assessed the impact of quercetin on BP through a systematic review and meta‐analysis of available randomized controlled trials. METHODS AND RESULTS: We searched PUBMED, Cochrane Library, Scopus, and EMBASE up to January 31, 2015 to identify placebo‐controlled randomized controlled trials investigating the effect of quercetin on BP. Meta‐analysis was performed using either a fixed‐effects or random‐effect model according to I(2) statistic. Effect size was expressed as weighted mean difference (WMD) and 95% CI. Overall, the impact of quercetin on BP was reported in 7 trials comprising 9 treatment arms (587 patients). The results of the meta‐analysis showed significant reductions both in systolic BP (WMD: −3.04 mm Hg, 95% CI: −5.75, −0.33, P=0.028) and diastolic BP (WMD: −2.63 mm Hg, 95% CI: −3.26, −2.01, P<0.001) following supplementation with quercetin. When the studies were categorized according to the quercetin dose, there was a significant systolic BP and diastolic BP‐reducing effect in randomized controlled trials with doses ≥500 mg/day (WMD: −4.45 mm Hg, 95% CI: −7.70, −1.21, P=0.007 and −2.98 mm Hg, 95% CI: −3.64, −2.31, P<0.001, respectively), and lack of a significant effect for doses <500 mg/day (WMD: −1.59 mm Hg, 95% CI: −4.44, 1.25, P=0.273 and −0.24 mm Hg, 95% CI: −2.00, 1.52, P=0.788, respectively), but indirect comparison tests failed to significant differences between doses. CONCLUSIONS: The results of the meta‐analysis showed a statistically significant effect of quercetin supplementation in the reduction of BP, possibly limited to, or greater with dosages of >500 mg/day. Further studies are necessary to investigate the clinical relevance of these results and the possibility of quercetin application as an add‐on to antihypertensive therapy

Effects of Quercetin on Blood Pressure: A Systematic Review and Meta‐Analysis of Randomized Controlled Trials

Quercetin, the most abundant dietary flavonol, has antioxidant effects in cardiovascular disease, but the evidence regarding its effects on blood pressure (BP) has not been conclusive. We assessed the impact of quercetin on BP through a systematic review and meta-analysis of available randomized controlled trials. We searched PUBMED, Cochrane Library, Scopus, and EMBASE up to January 31, 2015 to identify placebo-controlled randomized controlled trials investigating the effect of quercetin on BP. Meta-analysis was performed using either a fixed-effects or random-effect model according to I(2) statistic. Effect size was expressed as weighted mean difference (WMD) and 95% CI. Overall, the impact of quercetin on BP was reported in 7 trials comprising 9 treatment arms (587 patients). The results of the meta-analysis showed significant reductions both in systolic BP (WMD: -3.04&nbsp;mm&nbsp;Hg, 95% CI: -5.75, -0.33, P=0.028) and diastolic BP (WMD: -2.63&nbsp;mm&nbsp;Hg, 95% CI: -3.26, -2.01, P&lt;0.001) following supplementation with quercetin. When the studies were categorized according to the quercetin dose, there was a significant systolic BP and diastolic BP-reducing effect in randomized controlled trials with doses ≥500&nbsp;mg/day (WMD: -4.45&nbsp;mm&nbsp;Hg, 95% CI: -7.70, -1.21, P=0.007 and -2.98&nbsp;mm&nbsp;Hg, 95% CI: -3.64, -2.31, P&lt;0.001, respectively), and lack of a significant effect for doses &lt;500&nbsp;mg/day (WMD: -1.59&nbsp;mm&nbsp;Hg, 95% CI: -4.44, 1.25, P=0.273 and -0.24&nbsp;mm&nbsp;Hg, 95% CI: -2.00, 1.52, P=0.788, respectively), but indirect comparison tests failed to significant differences between doses. The results of the meta-analysis showed a statistically significant effect of quercetin supplementation in the reduction of BP, possibly limited to, or greater with dosages of &gt;500&nbsp;mg/day. Further studies are necessary to investigate the clinical relevance of these results and the possibility of quercetin application as an add-on to antihypertensive therapy