Toxicological Study Of Silver Nanoparticles In Aquatic Organisms

With the rapid development of nanotechnology, nanomaterials have been widely used in various fields. Silver nanoparticles (Ag NPs) have become one of the extensively applied nanomaterials in biomedical, consumer products and other fields for their distinctive physicochemical properties and advantageous antibacterial properties. The use of these products will release Ag NPs into sewage or industrial wastewater. However, little is known about their potential impacts on aquatic organisms. In this study, zebrafish embryos were selected as model organism. The aim of this study was to understand the environmental impacts of silver nanoparticles on early development of aquatic organisms.The teratogenic effect and particle size effect of Ag NPs were studied in developing zebrafish embryos. Selected zebrafish embryos were exposed to different concentrations of 4 nm- and 10 nm-Ag NPs respectively from 4 hpf to 96 hpf. The 96 h LC50 of S4 and S10 to zebrafish embryos were 6.69 mg/L and 11.75 mg/L, and 96 h EC50 of S4 and S10 were 4.12 mg/L and 5.91 mg/L respectively. The malformation percentages of zebrafish embryos increased with the exposure concentration of Ag NPs. Exposed zebrafish embryos exhibited malformations such as shorter body, yolk sac deformities, small head, small eye, cardiac anomalies and disappeared somites. And the percentages of small head, yolk sac deformities and disappeared somites were relatively high as compared to control. The toxicity of Ag NPs in exposed zebrafish embryos was size-dependent, and the toxicity of smaller sized-Ag NPs (4 nm) was higher than that of larger sized-Ag NPs (10 nm).The impacts of Ag NPs on the early development of zebrafish embryos were studied and compared to that of Ag+ according to short body and yolk sac deformities observed. Results indicated that there was no release of Ag+ in the Ag NPs suspension used in this study. The 96 h LC50 of Ag+ to zebrafish embryos was 0.054 mg/L which indicated the toxicity of Ag+ was much greater than that of Ag NPs. Both Ag NPs and Ag+ could delay the hatching and reduce hatching rate. The hatched zebrafish showed significant shorter body length, and the total protein content reduced accordingly. These results indicated both Ag NPs and Ag+ could inhibit development of zebrafish embryos. Cell apoptosis in the trunk and yolk sac of zebrafish embryos were observed respectively after treatment with Ag NPs and Ag+ through AO staining. The ROS levels in Ag NPs- or Ag+-exposed zebrafish embryos were significantly higher than that of control groups. The RT-PCR results further suggested that Ag NPs could induce apoptosis in zebrafish embryos via the caspase-dependent pathway. However, exposure to Ag+ not only affected the oxidation-related gene pathway, but also induced apoptosis via p53 pathway and inhibited the development of exposed zebrafish embryos.The neural malformations including head and eye deformities were further studied in exposed zebrafish embryos. The morphological observation showed Ag NPs-exposed zebrafish embryos exhibited unclear head boundaries, hindbrain hypoplasia and reduced eye pigment. At the molecular level, Ag NPs altered the neural development-related genes (gfap, huC and ngnl) and metal detoxification-related genes (MT and ABCC). And the up-regulated expression of ABCC2 in the head of exposed zebrafish was relatively more significant than in the trunk. These results indicated Ag NPs might effect the neural cell differentiation and thus showed neurotoxicity. The heart rate of exposed zebrafish embryos decreased significantly at 72 hpf. However, the expression of heart development- related genes (Gata5 and Nkx2.5) did not alter significantly. In addition, Ag NPs also changed the expression levels of AhR2 and CyplA which used to mediate PAH toxicity. The in vivo accumulation of Ag NPs in zebrafish embryos was also detected. The uptake of S4 into zebrafish embryos was higher than that of S10, and this was in line with the size-dependent toxicity of Ag NPs. The content of silver in the head of zebrafish was close to or slightly higher than that in the trunk, and this could further explain the expression of ABCC2 and the neurotoxicity presented by Ag NPs.This study suggested Ag NPs could produce strong teratogenic effects on aquatic organisms, and even influence the survival of them at relatively high concentration. The smaller sized Ag NPs were more toxic. The effects of Ag NPs on aquatic organisms were mainly inducing apoptosis and oxidative stress, thus hindering the early development of aquatic organisms, In addition, Ag NPs showed strong neurotoxicity and could affect the neural cell differentiation. However, there was no released Ag+ in the Ag NPs solution used in this study. This indicates the toxicity of Ag NPs could not be only attributed to the Ag+, and the toxicity of Ag+ was much higher than that of Ag NPs.The toxicological study showed that Ag NPs could potentially affect the normal hatching, development and breeding of aquatic organisms. The hatched aquatic organisms exhibited shorter body length which implies significantly delayed development. The exposed embryos also have neural defects such as head and eye hypoplasia. The negative and chronic environmental impacts of AgNPs to aquatic organisms should be put into consideration in its future production and usage.