In vitro investigation of zinc oxide nanoparticle toxic effects in spermatogonial cells at the molecular level.

Because spermatogonia transmit genetic information across generations, their DNA must be protected from environmental damages, including exposure to zinc oxide nanoparticles (ZnO NPs), which are frequently used in modern technology. Here, we used an in vitro system enriched for spermatogonia and exposed them to 10 and 20 µg/ml ZnO NPs for one/seven days. We did not detect any significant cell death, chromosomal instability, or DNA fragmentation in the spermatogonia treated with the ZnO NPs following one-day treatment with 10 or 20 µg/ml ZnO NPs. However, ZnO NPs (both 10 and 20 µg/ml) induced chromosomal instability in the spermatogonia after seven days of treatment. Moreover, one-day exposure to these NPs induced reactive oxygen species (ROS) generation and upregulation of apoptotic pathway-related genes p53, Caspase3 and Il6, as an inflammatory factor. Taken together, our study provides preliminary evidence for possible damages induced by low concentrations of ZnO NPs in spermatogonia. We should pay increased attention when using these NPs because of the silent damages in spermatogonia that can be transmitted to the next generation and cause severe effects. However, more data and validation of these results are required to determine the extent of this concern.

In vitro cytotoxicity of zinc oxide nanoparticles in mouse ovarian germ cells.

Recently, metal oxide nanoparticles such as zinc oxide nanoparticles (ZnO-NPs) have received considerable attention and humans are exposed to them in everyday life. The increasing use of ZnO-NPs may lead to human health issues. However, little is known about their effects on female reproductive systems, particularly on female germ cells. Germ cells differentiation is a complex biological process that is sensitive to environmental insults and any negative effect on germ cells development may inhibit fertility. Therefore, this study aimed to determine the impact of ZnO-NPs on mouse ovarian germ cells in an in vitro system. The effects of ZnO-NPs on these cells were evaluated using light microscopy, cell proliferation assessment, reactive oxygen species (ROS) level determination, standard cytotoxicity assessment (cell viability assessed by PI staining) and gene expression analysis. Our results demonstrated that ZnO-NPs have cytotoxic effects in a concentration- and time-dependent manner in mouse ovarian germ cells. Exposure of cells to ZnO-NPs concentration-dependently enhanced ROS generation. Furthermore, molecular analysis of ZnO-NPs-treated cells showed a significant increase in expression of premeiotic germ cells markers but a decrease in meiotic and post-meiotic markers compared to un-treated cells. Taken together, our data provides a preliminary insight into possible adverse effects of ZnO-NPs on mouse ovarian germ cells differentiation even at low concentrations.

Short time exposure to low concentration of zinc oxide nanoparticles up-regulates self-renewal and spermatogenesis-related gene expression.

Extensive application of zinc oxide (ZnO) nanoparticles (NPs) in everyday life results in increased exposure to these NPs. Spermatogonial stem cells (SSCs) guarantee sperm production throughout the male reproductive life by providing a balance between self-renewal and differentiation. We used an in vitro platform to investigate the ZnO NPs effects on SSCs. We successfully synthesized ZnO NPs. In order to investigate these NPs, we isolated SSCs from mouse testes and cultured them in vitro. Our results confirmed the uptake of ZnO NPs by the cultured SSCs. We observed a dose- and time-dependent decrease in SSC viability. Both spherical and nanosheet ZnO NPs had the same cytotoxic effects on the SSCs, irrespective of their shapes. Moreover, we have shown that short time (one day) exposure of SSCs to a low concentration of ZnO NPs (10 µg/mL) promoted expressions of specific genes (Plzf, Gfr α1 and Bcl6b) for SSC self-renewal and differentiation genes (Vasa, Dazl, C-kit and Sycp3) expressed by spermatogonia during spermatogenesis. Our study provides the first insight into ZnO NPs function in SSCs and suggests a new function for ZnO NPs in the male reproductive system. We demonstrated that ZnO NPs might promote spermatogenesis via upregulation of gene expression related to SSC self-renewal and differentiation at low concentrations. Additional research should clarify the possible effect of ZnO NPs on the SSC genome and its effects on human SSCs.