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J Xenobiot
2024 Aug 22;143:1130-1142. doi: 10.3390/jox14030064.
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Vanadium Toxicity Is Altered by Global Warming Conditions in Sea Urchin Embryos: Metal Bioaccumulation, Cell Stress Response and Apoptosis.
Martino C
,
Geraci F
,
Scudiero R
,
Barone G
,
Naselli F
,
Chiarelli R
.
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In recent decades, the global vanadium (V) industry has been steadily growing, together with interest in the potential use of V compounds as therapeutics, leading to V release in the marine environment and making it an emerging pollutant. Since climate change can amplify the sensitivity of marine organisms already facing chemical contamination in coastal areas, here, for the first time, we investigated the combined impact of V and global warming conditions on the development of Paracentrotus lividus sea urchin embryos. Embryo-larval bioassays were carried out in embryos exposed for 24 and 48 h to sodium orthovanadate (Na3VO4) under conditions of near-future ocean warming projections (+3 °C, 21 °C) and of extreme warming at present-day marine heatwave conditions (+6 °C, 24 °C), compared to the control temperature (18 °C). We found that the concomitant exposure to V and higher temperature caused an increased percentage of malformations, impaired skeleton growth, the induction of heat shock protein (HSP)-mediated cell stress response and the activation of apoptosis. We also found a time- and temperature-dependent increase in V bioaccumulation, with a concomitant reduction in intracellular calcium ions (Ca2+). This work demonstrates that embryos' sensitivity to V pollution is increased under global warming conditions, highlighting the need for studies on multiple stressors.
Figure 1. Morphological analysis of P. lividus embryos. Upper panel: images of representative embryos and larvae at 24 and 48 h of development. Embryos were grown at three different temperatures (18 °C: (A1,A2,D1,D2)); 21 °C: (B1,B2,E1,E2); and 24 °C: (C1,C2,F1,F2)) in the presence (A2–F2) or absence (A1–F1) of V. Bar: 100 μm. Lower panel: Number of embryonic stages detected in all treatments. Developmental stages are indicated as EG (early gastrula); IG (intermediate gastrula); AG (advanced gastrula); Pr (prism); EPl (early pluteus); and APl (advanced pluteus). * Altered phenotypes. Data are presented as means of triplicate experiments (n = 3).
Figure 2. Histograms showing quantitative analysis of the amount of V (A,B) and Ca (C,D) incorporated after 24 and 48 h of development/treatment. Embryos were cultured in V 1 mM at 18, 21 and 24 °C. V and Ca contents were detected by Inductively Coupled Plasma Mass Spectrometry (ICP-MS), determining mean metal quantity in about 250,000 embryos. Data are presented as means ± standard deviations (SDs) of triplicate experiments (n = 3). Treatments with same letter do not differ (Tukey HSD).
Figure 3. Immunoreactive bands obtained by immunoblotting detection and quantitative analysis for HSP 60 and cleaved caspase-7. (A) Total lysates of control and treated embryos after 24 and 48 h of development/treatment. Actin was used as loading control. Histograms show densitometric analysis of bands identified for (B) HSP 60 and (C) cleaved caspase-7. Relative protein expression, reported as arbitrary units, was calculated as band density ratio to that of actin. Data are presented as means ± standard deviations (SD) of triplicate experiments (n = 3). Data were analyzed by two-way ANOVA. Treatments with same letter do not differ (Tukey HSD).
Figure 4. Apoptotic nuclei evalutation in whole-mount embryos detected by fluorescent TUNEL assay and densitometric analysis. Pictures of demonstrative embryos at 48 h of development/treatment. DNA fragmentation (A1–H1). Nuclei marked with propidium iodide (A2–H2). Merging of both signals (A3–H3). Control embryo reared at 18 °C (A1–A3); V-treated embryo at 18 °C (B1–B3); embryo grown at 21 °C (C1–C3); V-treated embryo at 21 °C (D1–D3); embryo grown at 24 °C (E1–E3); and V-treated embryo at 24 °C (F1–F3). Negative control embryo (G1–G3). Positive control embryo (H1–H3). Bar = 100 μm. Data in histograms report relative quantification of green fluorescence related to fragmented DNA as mean relative levels expressed in arbitrary units as fold change compared to 18 °C value, assumed as 1 in histogram, and are presented as means ± standard deviations (SDs) of triplicate experiments (n = 3) of entire morphological population. Data were analyzed by two-way ANOVA. Treatments with same letter do not differ (Tukey HSD).
