Neodymium Oxide-Induced Male Reproductive Dysfunction in Mice Is Associated with TFR1/FTH1 Axis Dysregulation and Ferroptosis-Related Alterations: A Proteomics-Based Study

Growing concerns regarding the environmental and occupational health risks associated with rare earth elements have highlighted the importance of elucidating the mechanisms underlying neodymium oxide (Nd2O3)-induced male reproductive toxicity. In this study, male C57BL/6J mice were exposed to Nd2O3 at concentrations of 250, 500, or 1000 mg/m3 through nose-only inhalation for 2 h/day over 90 consecutive days. The effects of Nd2O3 exposure on male reproductive function were investigated using histopathological examination, sperm analysis, transmission electron microscopy (TEM), quantitative proteomics, bioinformatics analysis, Western blotting, and biochemical assays. Nd2O3 exposure was associated with testicular histopathological alterations, decreased sperm count, and increased sperm abnormalities. Quantitative proteomic analysis identified 892 differentially expressed proteins enriched in biological processes and pathways associated with spermatogenesis, ribosome function, endoplasmic reticulum protein processing, programmed cell death, iron uptake and transport, lipid metabolism, and ferroptosis. Protein–protein interaction (PPI) analysis further identified a ferroptosis-associated module containing ACSL4, HMOX1, TF, HMGB1, and SIRT1. TEM qualitatively revealed apparent mitochondrial shrinkage and cristae alterations in testicular cells, compatible with ferroptosis-associated ultrastructural changes. Western blotting demonstrated dose-dependent upregulation of TFR1 and downregulation of FTH1 following Nd2O3 exposure. Moreover, Nd2O3 exposure decreased serum testosterone and estradiol levels, increased total iron, Fe2+, malondialdehyde, and 4-hydroxynonenal levels, and depleted reduced glutathione in both serum and testicular tissues, indicating disrupted iron homeostasis, impaired antioxidant capacity, and enhanced lipid peroxidation. Collectively, these proteomic, molecular, biochemical, and morphological findings indicate that Nd2O3-induced male reproductive dysfunction is associated with TFR1/FTH1 axis dysregulation, disrupted iron homeostasis, lipid peroxidation, and ferroptosis-related alterations.