Effects of Titanium Dioxide Nanoparticles on Porcine Prepubertal Sertoli Cells: An “In Vitro” Study

Abstract

The increasing use of nanomaterials in a variety of industrial, commercial, medical products, and their environmental spreading has raised concerns regarding their potential toxicity on human health. Titanium dioxide nanoparticles (TiO2 NPs) represent one of the most commonly used nanoparticles. Emerging evidence suggested that exposure to TiO2 NPs induced reproductive toxicity in male animals. In this in vitro study, porcine prepubertal Sertoli cells (SCs) have undergone acute (24 h) and chronic (from 1 up to 3 weeks) exposures at both subtoxic (5 µg/ml) and toxic (100 µg/ml) doses of TiO2 NPs. After performing synthesis and characterization of nanoparticles, we focused on SCs morphological/ultrastructural analysis, apoptosis, and functionality (AMH, inhibin B), ROS production and oxidative DNA damage, gene expression of antioxidant enzymes, proinflammatory/immunomodulatory cytokines, and MAPK kinase signaling pathway. We found that 5 µg/ml TiO2 NPs did not induce substantial morphological changes overtime, but ultrastructural alterations appeared at the third week. Conversely, SCs exposed to 100 µg/ml TiO2 NPs throughout the whole experiment showed morphological and ultrastructural modifications. TiO2 NPs exposure, at each concentration, induced the activation of caspase-3 at the first and second week. AMH and inhibin B gene expression significantly decreased up to the third week at both concentrations of nanoparticles. The toxic dose of TiO2 NPs induced a marked increase of intracellular ROS and DNA damage at all exposure times. At both concentrations, the increased gene expression of antioxidant enzymes such as SOD and HO-1 was observed whereas, at the toxic dose, a clear proinflammatory stress was evaluated along with the steady increase in the gene expression of IL-1α and IL-6. At both concentrations, an increased phosphorylation ratio of p-ERK1/2 was observed up to the second week followed by the increased phosphorylation ratio of p-NF-kB in the chronic exposure. Although in vitro, this pilot study highlights the adverse effects even of subtoxic dose of TiO2 NPs on porcine prepubertal SCs functionality and viability and, more importantly, set the basis for further in vivo studies, especially in chronic exposure at subtoxic dose of TiO2 NPs, a condition closer to the human exposure to this nanoagent.

Impact of chitosan administration on titanium dioxide nanoparticles induced testicular dysfunction

“…At a higher dose (300 mg/kg), TiO 2 NPs enhanced oxidative stress via increasing testicular levels of MDA and reducing testicular levels of SOD, CAT, and GSH 16 . Similar outcomes were reported at the cellular level using rat, human, or prepubertal porcine primary cultured Sertoli cells 25 , 26 . Nano-titanium particles stimulated production of reactive oxygen species in mouse testes after a 60-day exposure period with a subsequent alteration in the activities of spermatogenesis-dependent enzymes 27 .…”

RETRACTED: Long-Lasting Exendin-4-Loaded PLGA Nanoparticles Ameliorate Cerebral Ischemia/Reperfusion Damage in Diabetic Rats

“…The increasing use of nanomaterials in a variety of industrial, commercial, and medical products and their environmental spreading have raised concerns regarding their potential toxicity to human health. For example, titanium dioxide nanoparticles may induce ultrastructural alterations appearing at the third week by inducing activation of caspase-3 and increasing intracellular ROS and DNA damage [ 24 ]. On the other hand, PLGA nanoparticles were beneficial for ensuring structural stability, enhancing absorption, followed by sustained drug release [ 25 ].…”

Sperm-carried IGF2 downregulated the expression of mitogens produced by Sertoli cells: A paracrine mechanism for regulating spermatogenesis?

“…Porcine SCs were isolated from pre-pubertal neonatal (7-15 days of age) Large White pigs using established methods ( 49 , 50 ). These cells were incubated with increasing concentrations (0.33 ng/mL, 3.33 ng/mL, 10 ng/mL) of recombinant human IGF2 (rhIGF2) for 48 hours.…”

“…Cells were cultured for 3 days. Therefore, the purity and functional competence of the SC monolayers were evaluated according to previously established methods ( 50 ). In particular, purity was assessed by immunofluorescence analysis of specific markers for both SC (AMH positivity and vimentin positivity >95 ± 2%) and non-SC cells (3β-HSD for Leydig cells and ASMA for peritubular cells <5 ± 2%), (data not shown).…”

“…Aliquots of the culture media of treated and untreated SCs were stored at −20°C for the assessment of AMH (AMH Gen IIELISA, Beckman Coulter; intra-assay CV = 3.89%; inter-assay CV = 5.77%) and inhibin B (Inhibin B Gen II ELISA, Beckman Coulter, Webster, TX, USA; intra-assay CV =2.81%; inter-assay CV= 4.33%) secretion as previously described ( 50 ).…”