The use of ex vivo ovary culture for assessment of alterations in steroidogenesis following neonatal exposure to poly(ethylene glycol)-block-polylactide methyl ether or titanium dioxide nanoparticles in Wistar rats.

OBJECTIVES: Rapid development and widespread application of different types of nanoparticles (NPs) may result in increased exposure of humans and animals to NPs. Recently, reproductive toxicity due to NP exposure has become a major component of risk assessment. Current data have suggested that NPs may pose adverse effects on male and female reproductive health by altering normal testis and ovarian structure, and sex hormone levels. To detect possible alterations in steroidogenesis in adult and infantile rats following neonatal exposure to polymeric poly(ethylene glycol)-block-polylactide methyl ether (PEG-b-PLA) or titanium dioxide (TiO2) NPs, whole ovary cultures were used. METHODS: Newborn female Wistar rats were intraperitoneally (i.p.) injected daily with two different doses of PEG-b-PLA NPs (20 and 40 mg/kg body weight, b.w.) or TiO2 NPs (1% LD50 TiO2=59.2 µg/kg b.w. and 10% LD50 TiO2=592 µg/kg b.w.) from postnatal day 4 (PND 4) to PND 7. The ovaries were collected on PND73 and PND15 of PEG-b-PLA- and TiO2 NP-treated rats, respectively, and their corresponding control animals. Minced ovaries were cultured in vitro in the absence (basal conditions) or presence of gonadotropins (follicle-stimulating hormone, FSH and luteinizing hormone, LH) and insulin-like growth factor-1 (IGF-1) (stimulated conditions) for 6 days. At indicated time intervals, culture media were collected for steroid hormone (progesterone, estradiol) analysis by specific radioimmunoassay (RIA) and enzyme-linked immunosorbent assay (ELISA) kits. RESULTS: Basal progesterone and estradiol secretion by ovaries from adult rats (PND73) were significantly decreased (p<0.01) in both PEG-b-PLA-treated groups after 3 days and 1 day of ex vivo ovary culture, respectively, compared with control group. With the presence of FSH/LH and IGF-1 in the culture medium, progesterone and estradiol production significantly increased (p<0.001) compared to basal levels. Stimulated progesterone production was significantly decreased (p<0.05) in PEG-b-PLA40-treated group after 3 days of culture compared with controls. After ex vivo culture of rat ovaries collected on PND15, basal progesterone and estradiol levels measured in the culture media did not differ between control and both TiO2 NP-treated groups. The ovaries from rats neonatally exposed to both doses of TiO2 NPs failed to respond to FSH/IGF stimulation in progesterone secretion at all time intervals. CONCLUSIONS: The obtained results indicate that neonatal exposure to NPs in female rats may alter ovarian steroidogenic output (steroid hormone secretion) and thereby might subsequently induce perturbation of mammalian reproductive functions. Possible mechanisms (induction of oxidative stress, inflammation) of adverse effects of NPs on ovarian function should be further elucidated.

Hydrophobic sodium fluoride-based nanocrystals doped with lanthanide ions: assessment of in vitro toxicity to human blood lymphocytes and phagocytes.

In vitro immunotoxicity of hydrophobic sodium fluoride-based nanocrystals (NCs) doped with lanthanide ions was examined in this study. Although there is already a significant amount of optical and structural data on NaYF4 NCs, data on safety assessment are missing. Therefore, peripheral whole blood from human volunteers was used to evaluate the effect of 25 and 30 nm hydrophobic NaYF4 NCs dissolved in cyclohexane (CH) on lymphocytes, and of 10 nm NaYF4 NCs on phagocytes. In the concentration range 0.12-75 µg cm(-2) (0.17-106 µg ml(-1) ), both 25 and 30nm NaYF4 NCs did not induce cytotoxicity when measured as incorporation of [(3) H]-thymidine into DNA. Assessment of lymphocyte function showed significant suppression of the proliferative activity of T-lymphocytes and T-dependent B-cell response in peripheral blood cultures (n = 7) stimulated in vitro with mitogens phytohemagglutinin (PHA) and pokeweed (PWM) (PHA > PWM). No clear dose-response effect was observed. Phagocytic activity and respiratory burst of leukocytes (n = 5-8) were generally less affected. A dose-dependent suppression of phagocytic activity of granulocytes in cultures treated with 25 nm NCs was observed (vs. medium control). A decrease in phagocytic activity of monocytes was found in cells exposed to higher doses of 10 and 30 nm NCs. The respiratory burst of phagocytes was significantly decreased by exposure to the middle dose of 30 nm NCs only. In conclusion, our results demonstrate immunotoxic effects of hydrophobic NaYF4 NCs doped with lanthanide ions to lymphocytes and to lesser extent to phagocytes. Further research needs to be done, particularly faze transfer of hydrophobic NCs to hydrophilic ones, to eliminate the solvent effect.