Genomic instability related to zinc deficiency and excess in an in vitro model: is the upper estimate of the physiological requirements recommended for children safe?

Micronutrients are important for the prevention of degenerative diseases due to their role in maintaining genomic stability. Therefore, there is international concern about the need to redefine the optimal mineral and vitamin requirements to prevent DNA damage. We analyzed the cytostatic, cytotoxic, and genotoxic effect of in vitro zinc supplementation to determine the effects of zinc deficiency and excess and whether the upper estimate of the physiological requirement recommended for children is safe. To achieve zinc deficiency, DMEM/Ham's F12 medium (HF12) was chelated (HF12Q). Lymphocytes were isolated from healthy female donors (age range, 5-10 yr) and cultured for 7 d as follows: negative control (HF12, 60 µg/dl ZnSO4); deficient (HF12Q, 12 µg/dl ZnSO4); lower level (HF12Q + 80 µg/dl ZnSO4); average level (HF12Q + 180 µg/dl ZnSO4); upper limit (HF12Q + 280 µg/dl ZnSO4); and excess (HF12Q + 380 µg/dl ZnSO4). The comet (quantitative analysis) and cytokinesis-block micronucleus cytome assays were used. Differences were evaluated with Kruskal-Wallis and ANOVA (p < 0.05). Olive tail moment, tail length, micronuclei frequency, and apoptotic and necrotic percentages were significantly higher in the deficient, upper limit, and excess cultures compared with the negative control, lower, and average limit ones. In vitro zinc supplementation at the lower and average limit (80 and 180 µg/dl ZnSO4) of the physiological requirement recommended for children proved to be the most beneficial in avoiding genomic instability, whereas the deficient, upper limit, and excess (12, 280, and 380 µg/dl) cultures increased DNA and chromosomal damage and apoptotic and necrotic frequencies.

Possible radioprotective effect of folic acid supplementation on low dose ionizing radiation-induced genomic instability in vitro.

Ionizing radiation (IR) induces DNA damage through production of single and double-strand breaks and reactive oxygen species (ROS). Folic acid (FA) prevents radiation-induced DNA damage by modification of DNA synthesis and/or repair and as a radical scavenger. We hypothesized that in vitro supplementation with FA will decrease the sensitivity of cells to genetic damage induced by low dose of ionizing radiation. Annexin V, comet and micronucleus assays were performed in cultured CHO cells. After 7 days of pre-treatment with 0, 100, 200 or 300 nM FA, cultures were exposed to radiation (100 mSv). Two un-irradiated controls were executed (0 and 100 nM FA). Data were statistically analyzed with X2-test and linear regression analysis (P 0.05). We observed a significantly decreased frequency of apoptotic cells with the increasing FA concentration (P <0.05). The same trend was observed when analyzing DNA damage and chromosomal instability (P <0.05 for 300 nM). Only micronuclei frequencies showed significant differences for linear regression analysis (R2=94.04; P <0.01). Our results have demonstrated the radioprotective effect of folic acid supplementation on low dose ionizing radiation-induced genomic instability in vitro; folate status should be taken into account when studying the effect of low dose radiation in environmental or occupational exposure.

Assessment of the adverse effects of the acaricide amitraz: in vitro evaluation of genotoxicity.

Amitraz is a formamidine widely used in Veterinary Medicine for the treatment of ectoparasites. It is a highly liposoluble compound that is quickly absorbed through the skin and mucous membranes, thus making exposure potentially dangerous for humans and animals. The aim of this study was to compare the genotoxic potential of the active constituent of the insecticide amitraz and a commercial product containing amitraz in vitro in hamster cells. The induction of primary DNA damage was evaluated by alkaline single-cell gel electrophoresis (comet assay) and the apoptotic ability was examined by the Annexin V/propidium iodide staining assay. The commercial formulation significantly increased the index of DNA damage at concentrations of 2.50-3.75 µg/mL compared to the control. The active constituent only induced significant DNA damage with the highest concentration (3.75 µg/mL). Although both tested products increased the frequency of cell death, neither of them induced significant differences. Genotoxic potential is a primary risk factor for long-term effects such as carcinogenic and reproductive toxicology. Results presented here highlight the need for further investigation of the potential health risk of this veterinary medicine.