Mechanisms of nanosized titanium dioxide-induced testicular oxidative stress and apoptosis in male mice

Abstract

Background: Due to the increased application of titanium dioxide nanoparticles (TiO₂ NPs) in the food industry and daily life, their potential toxic effects in humans and animals have been investigated. However, very few studies have focused on testicular oxidative stress and/or apoptosis.

Methods: In order to understand the possible molecular mechanisms of testicular lesions following exposure to TiO₂ NPs, male mice were exposed to 2.5, 5, or 10 mg/kg body weight TiO₂ NPs for 90 consecutive days. Testicular oxidative stress and apoptosis were then evaluated, and the testicular mRNA expression of several genes and their proteins involved in oxidative stress and/or apoptosis was investigated.

Results: TiO₂ NPs entered Sertoli cells and caused severe testicular oxidative damage and/or apoptosis, accompanied by excessive production of reactive oxygen species and peroxidation of lipids, proteins and DNA as well as a significant reduction in antioxidant capacity. Furthermore, exposure to TiO₂ NPs resulted in the up-regulation of caspase-3, Nrbp2, and cytochrome c expression, and caused down-regulation of SOD, CAT, GPx, GST, GR, Cyp1b1, Car3, Bcl-2, Acaa2, and Axud1 expression in mouse testis.

Conclusions: TiO₂ NPs entered Sertoli cells via the blood-testis barrier and were deposited in mouse seminiferous cord and/or Sertoli cells, causing oxidative damage and apoptosis.

Figure 1

The (101) X-ray diffraction peak of anatase TiO₂NPs. The average particle size was about 5 nm calculated using Scherrer’s equation.

Figure 2

Transmission electron microscope images of anatase TiO₂NPs. (a) TiO₂ NP powder; (b) TiO₂ NPs suspended in HPMC solvent after incubation for 12 h. TEM images showed that the sizes of the TiO₂ NP powder or TiO₂ NPs suspended in HPMC solvent for 12 h ranged from 5 to 6 nm, respectively.

Figure 3

Histopathological observations of mouse testis following intragastric administration of TiO₂NPs for 90 consecutive days. The blue circle indicates few sperm, breakages or vacuolation, the yellow circle indicates apoptosis and/or necrosis of Sertoli cells, and the blue arrow indicates damaged seminiferous tubule. The red arrows indicate vacuolation, the yellow arrows indicate irregular arrangement of Sertoli cells, and the red circle indicates black agglomerates in the testis. The green arrow spot denotes a representative cell loaded with TiO₂ NPs. The right panel shows the corresponding Raman spectra identifying specific peaks at about 148 cm⁻¹.

Figure 4

Morphological changes in mouse sperm following intragastric administration of TiO ₂ NPs for 90 consecutive days.

Figure 5

Ultrastructure of Sertoli cells in mouse testis caused by intragastric administration of TiO₂NPs for 90 consecutive days. The yellow arrow indicates mitochondrial swelling, blushing, and/or cristae disappearance, the green arrow indicates TiO₂ NP aggregation in mitochondria, and the red arrow indicates nuclear membrane collapse and chromatin marginalization in Sertoli cells. The green arrow spot denotes a representative cell loaded with TiO₂ NPs. The right panel shows the corresponding Raman spectra identifying specific peaks at about 148 cm⁻¹.

Figure 6

Alterations in the mRNA expression of oxidative stress and/or apoptosis-related genes in mouse testis caused by intragastric administration of TiO₂NPs for 90 consecutive days. *p < 0.05, **p < 0.01, and ***p < 0.001. Values represent mean ± SD (n = 5).

Figure 7

Alterations in the expression of oxidative stress and/or apoptosis-related proteins in mouse testis caused by intragastric administration of TiO₂NPs for 90 consecutive days. *p < 0.05, **p < 0.01, and ***p < 0.001. Values represent mean ± SD (n = 5).