Differential nanoreprotoxicity of silver nanoparticles in male somatic cells and spermatogonial stem cells

Abstract

Background Silver nanoparticles (AgNPs) possess unique physical, chemical, and biological properties. AgNPs have been increasingly used as anticancer, antiangiogenic, and antibacterial agents for the treatment of bacterial infections in open wounds as well as in ointments, bandages, and wound dressings. The present study aimed to investigate the effects of two different sizes of AgNPs (10 nm and 20 nm) in male somatic Leydig (TM3) and Sertoli (TM4) cells and spermatogonial stem cells (SSCs). Methods Here, we demonstrate a green and simple method for the synthesis of AgNPs using Bacillus cereus culture supernatants. The synthesized AgNPs were characterized using ultraviolet and visible absorption spectroscopy, X-ray diffraction, Fourier transform infrared spectroscopy, and transmission electron microscopy (TEM). The toxicity of the synthesized AgNPs was evaluated by the effects on cell viability, metabolic activity, oxidative stress, apoptosis, and expression of genes encoding steroidogenic and tight junction proteins. Results AgNPs inhibited the viability and proliferation of TM3 and TM4 cells in a dose- and size-dependent manner by damaging cell membranes and inducing the generation of reactive oxygen species, which in turn affected SSC growth on TM3 and TM4 as feeder cells. Small AgNPs (10 nm) were more cytotoxic than medium-sized nanoparticles (20 nm). TEM revealed the presence of AgNPs in the cell cytoplasm and nucleus, and detected mitochondrial damage and enhanced formation of autosomes and autolysosomes in the AgNP-treated cells. Flow cytometry analysis using Annexin V/propidium iodide staining showed massive cell death by apoptosis or necrosis. Real-time polymerase chain reaction and western blot analyses indicated that in TM3 and TM4 cells, AgNPs activated the p53, p38, and pErk1/2 signaling pathways and significantly downregulated the expression of genes related to testosterone synthesis (TM3) and tight junctions (TM4). Furthermore, the exposure of TM3 and TM4 cells to AgNPs inhibited proliferation and self-renewal of SSCs. Conclusion Our results suggest that AgNPs exhibit size-dependent nanoreprotoxicity in male somatic cells and SSCs, strongly suggesting that applications of AgNPs in commercial products must be carefully evaluated. Further studies of AgNPs-induced nanoreprotoxicity in animal models are required.

Acute reproductive toxicology after intratesticular injection of silver nanoparticles (AgNPs) in Wistar rats

“…Moreover, at concentrations !10 lg/ml, Ag-NPs induced a significant decline in SSC proliferation by disrupting components of the Glial cell-derived neurotrophic factor (GDNF) signaling pathway, which is essential for SSC self-renewal in vivo and in vitro (Braydich-Stolle et al 2010). AgNPs was proven to be toxic for the cells involved in spermatogenesis, such as Leydig and Sertoli cells and SSCs and this toxicity was linked to a rapid ROS production and accumulation (Zhang et al 2015). High ROS levels in seminal plasma have been correlated with poor sperm morphology and is associated with the inhibition of sperm function and viability (Aziz et al 2004).…”

“…Studies have reported that AgNPs cause: DNA damage in germ cells, decrease sperm count and promote morphometric changes in seminiferous tubules (Dziendzikowska et al 2012;Gromadzka-Ostrowska et al 2012); decrease spermatogonia proliferation (Braydich-Stolle et al 2005, and increase germ cell apoptosis (Garcia et al 2014). In an elegant study, Zhang et al (2015) demonstrated that AgNPs induced massive cell death in TM3 (Leydig cell line) and TM4 (Sertoli cell line) cells, and inhibited proliferation and self-renewal of spermatogonial stem cells (SSCs). In this study AgNPs induced expression of autophagy-related genes and activated signaling molecules involved in apoptosis.…”

Silver nanoparticles: Synthesis, medical applications and biosafety

“…AgNPs can cross the biological barriers to deposit in reproductive organs including testis, epididymis, ovary and uterus. Thus, AgNPs may damage germ cells and related cells, such as primary and secondary follicles, germline stem cells, Sertoli cells and Leydig cells 42 , 347 . Besides, AgNPs can also cause changes in sexual behavior by affecting the secretion of hormones within the reproductive organs and glands.…”

“…Further studies confirmed that the reproductive toxicities of AgNPs are achieved by increasing inflammation, disrupting DNA structure, down-regulating gene expression, decreasing mitochondrial function, inducing ROS production and apoptosis. These toxicities of AgNPs to the reproductive system are size-, time- and dose-dependent 347 , 348 . Zhang et al 347 investigated the effects of AgNPs with different sizes (10 nm and 20 nm) on male somatic Leydig cells and Sertoli cells, and found that cell viability was inhibited by AgNPs in size- and concentration-dependent manners.…”

Sodium Selenite Protects Against Silver Nanoparticle-Induced Testicular Toxicity and Inflammation

“…Henceforth, the toxicity of AgNPs has been studied widely in many in vitro [4][5][6] and in vivo models [1,7,8]. Recent studies have demonstrated the adverse effects of AgNPs on the male reproductive tract, particularly spermatogenesis and the quality of sperm and male somatic cells and spermatogonial stem cells [9,10]. Early exposure to AgNPs has the potential to disrupt fetal and postnatal health in the embryo and abnormal development of the placenta [11].…”