Investigation of the potential toxicity of zinc oxide nanoparticles on mouse spermatogonial stem cells

Azam Javadi,1,* Maryam farzaneh,2 Saadat mokhtari,3 Seyede faezeh moraveji,4 Fereshteh esfandiari,5 Hamid gourabi,6

1. Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
2. Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
3. Department of Physics, Shahid Beheshti University, Tehran, Iran
4. Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
5. Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
6. Department of Genetics, Reproductive Biomedicine Research Center, Royan Institute for Reproductive Biomedicine, ACECR, Tehran, Iran

Abstract

Introduction

The widespread applications of nanoparticles have made it important to investigate their side effects on the environment and living organisms. among the metal oxide nanoparticles, zinc oxide nanoparticles (zno nps) are widely used in various industries and consumer products such as food packaging and additives, cosmetic products, sunscreens and biomedical applications. spermatogonial stem cells (sscs) that are sperms precursors are very important due to the role of them in transmitting genetic information to the next generation. zno nps according to their nano scale can easily pass through the blood–testis barriers and affect these important cells. herein, we evaluated the toxicity effect of zno nps on mouse sscs.

Methods

We synthesized zno nps and have been characterized them by xrd and sem. sscs were isolated from testes of 5-7 days nmri mice and characterized by immunofluorescence (if) staining. based on a literature review, the cells were treated with different concentration of zno nps; 10, 20, 30 and 40 µg/ml and evaluated for the viability of them by pi staining at one and seven days after treatment. after determining the maximum and minimum harmful concentration on the survival of these cells, the damage to the dna of sscs by these concentrations were investigated by tunel test. moreover, the gene expression of cytokines: il-6, il-8 and tnfα were investigated by q-rt pcr method.

Results

xrd and sem analysis confirmed successful synthesis of zno nps. if for ssc-specific markers, plzf and gfrα, confirmed the presence of sscs. our results showed, the survival of treated cells with zno nps decreased significantly compared to the control group starting from concentrations of 30 µg/ml one day after treatment and starting from concentrations of 20 µg/ml, 7 days after treatment (p<0.05). also, the entry of nps into the cell was confirmed by using an electron microscope (tem). we observed dna damage at concentration of 30 µg/ml one day after treatment and also 7 days after treatment, starting from concentrations of 20 µg/ml. also the results showed that the zno nps at concentration of 10 μg/ml on the first day after treatment, increased the expression of cytokine il-6 and 7 days after treatment, zno nps increased the expression of il-8 (p<0.05). : xrd and sem analysis confirmed successful synthesis of zno nps. if for ssc-specific markers, plzf and gfrα, confirmed the presence of sscs. our results showed, the survival of treated cells with zno nps decreased significantly compared to the control group starting from concentrations of 30 µg/ml one day after treatment and starting from concentrations of 20 µg/ml, 7 days after treatment (p<0.05). also, the entry of nps into the cell was confirmed by using an electron microscope (tem). we observed dna damage at concentration of 30 µg/ml one day after treatment and also 7 days after treatment, starting from concentrations of 20 µg/ml. also the results showed that the zno nps at concentration of 10 μg/ml on the first day after treatment, increased the expression of cytokine il-6 and 7 days after treatment, zno nps increased the expression of il-8 (p<0.05).

Conclusion

Our data indicates cyto- and genotoxic effects as well as a pro-inflammatory potential of zno-nps in mouse sscs. therefore, the growing trend in daily application of zno nps can have an effect on the male fertility through the effect on the male sscs. considering this issue, it is advisable to reduce its consumption in industries and consumer products in order to reduce its harmful effects on fertility.

Keywords

Sscs, zinc oxide nanoparticles, dna damage, toxicity, inflammatory