Fluoride exposure during puberty induces testicular impairment via ER stress-triggered apoptosis in mice.

Fluoride, a ubiquitous environmental compound, carries significant health risks at excessive levels. This study investigated the reproductive toxicity of fluoride exposure during puberty in mice, focusing on its impact on testicular development, spermatogenesis, and underlying mechanisms. The results showed that fluoride exposure during puberty impaired testicular structure, induced germ cell apoptosis, and reduced sperm counts in mice. Additionally, the SOD activity and GSH content were significantly decreased, while MDA content was significantly elevated in the NaF group. Immunohistochemistry showed an increase in the number of cells positive for GRP78, a key ER stress marker. Moreover, qRT-PCR and Western blot analyses confirmed the upregulation of both Grp78 mRNA and protein expression, as well as increased mRNA expression of other ER stress-associated genes (Grp94, chop, Atf6, Atf4, and Xbp1) and enhanced protein expression of phosphorylated PERK, IRE1α, eIF2α, JNK, XBP-1, ATF-6α, ATF-4, and CHOP. In conclusion, our findings demonstrate that fluoride exposure during puberty impairs testicular structure, induces germ cell apoptosis, and reduces sperm counts in mice. ER stress may participate in testicular cell apoptosis, and contribute to the testicular damage and decreased sperm counts induced by fluoride.

NCOA4-mediated ferritinophagy participates in cadmium-triggered ferroptosis in spermatogonia.

Cadmium (Cd) is a common pollutant with reproductive toxicity. Our previous study revealed that Cd triggered spermatogonia ferroptosis. However, the underlying mechanisms remain unclear. Nuclear receptor coactivator 4 (NCOA4) mediates ferritinophagy and specific degradation of ferritin through lysosomes, resulting in the release of ferrous ions. Excessive autophagy can lead to ferroptosis. This study investigated the role of autophagy in Cd-triggered ferroptosis using GC-1 spermatogonial (spg) cells which exposed to CdCl2 (5 µM, 10 µM, or 20 µM) for 24 without/with CQ. The cells which transfected with Ncoa4-siRNA were used to explore the role of NCOA4-mediated ferritinophagy in Cd-triggered ferroptosis. The results revealed that Cd caused mitochondrial swelling, rupture of cristae, and vacuolar-like changes. The Cd-treated cells exhibited more autophagosomes. Simultaneously, Cd increased intracellular iron, reactive oxygen species, and malondialdehyde concentrations while decreasing glutathione content and Superoxide Dismutase-2 activity. Moreover, Cd upregulated mRNA levels of ferritinophagy-associated genes (Ncoa4, Lc3b and Fth1), as well as enhanced protein expression of NCOA4, LC3B, and FTH1. While Cd decreased the mRNA and protein expression of p62/SQSTM1. These results showed that Cd caused ferritinophagy and ferroptosis. The use of chloroquine to inhibit autophagy ameliorated Cd-induced iron overload and ferroptosis. Moreover, Ncoa4 knockdown in spermatogonia significantly reduced intracellular iron concentration and alleviated Cd-triggered ferroptosis. In conclusion, our findings demonstrate that Cd activates the ferritinophagy pathway mediated by NCOA4, resulting in iron accumulation through ferritin degradation. This causes oxidative stress, ultimately initiating ferroptosis in spermatogonia. Our results may provide new perspectives and potential strategies for preventing and treating Cd-induced reproductive toxicity.