Seminal cadmium affects human sperm motility through stable binding to the cell membrane

Abstract

Environmental pollutants are claimed to be major factors involved in the progressive decline of the fertility rate worldwide. Exposure to the heavy metal Cadmium (Cd) has been associated with reproductive toxicity due to its ionic mimicry. However, the possible direct accumulation of Cd in human sperm cells has been poorly investigated. In this study, we aimed to clarify the possible direct effect of Cd exposure on sperm function through the analysis of its cell accumulation. Semen samples from 30 male subjects residing in high environmental impact areas and adhering to the "Exposoma e Plurifocalità nella Prevenzione Oncologica" campaign for testis cancer prevention were compared with semen samples from 15 males residing in low exposure areas. Semen levels and cell Cd content were quantified by inductively coupled plasma (ICP) spectroscopy. Cell Cd distribution was assessed by scanning electron microscopy coupled with energy dispersive spectroscopy (SEM-EDS). The impact of Cd on sperm function was evaluated by the in vitro exposure to the heavy metal, whilst possible scavenging approaches/agents were assessed. In addition to higher values of semen Cd, exposed subjects showed a reduction in total motile sperm fraction compared to not-exposed controls (59.6% ± 13.6% vs. 66.3% ± 7.3%, p = 0.037). Semen Cd levels were also significantly correlated with SEM-EDS signals of Cd detected on the head and neck of sperm (respectively p = 0.738, p < 0.001 and ρ = 0.465, p < 0.001). A total of 2 h of in vitro exposure to 0.5 μM Cd was associated with a significant reduction of sperm progressive motility. Scavenging approaches with either hypo-osmotic swelling or 10 μM reduced glutathione were ineffective in blunting cell Cd and restoring motility. The reduction of exposure levels appears to be the main approach to reducing the reproductive issues associated with Cd.

Keywords: heavy metals; inductively-coupled plasma spectroscopy; ionic mimicry; scanning electron microscopy/energy dispersive spectroscopy; scavenging.

FIGURE 1

Representative images of Cadmium (Cd) mapping at scanning electron microscopy coupled with energy dispersive spectroscopy (SEM-EDS) analysis in sperm cells from normozoospermic healthy donors at basal not exposed conditions (CTRL) and upon in vitro exposure to Cd 0.5 μM for 1 hour at 37°C. The red boxes in the spectra underline the specific signals deriving from Cd detection in elemental analysis by SEM-EDS. The images in the inserts show the levels of Cd detection on the cell through a color intensity scale.

FIGURE 2

(A) Representative images of Cadmium (Cd) mapping at scanning electron microscopy coupled with energy dispersive spectroscopy (SEM-EDS) analysis, conducted on isolated sperm cells from 30 male subjects residing in high environmental impact areas. (B) Analysis of Cd signal reported as the percentage of carbon (C) signals in 747 representative cell sites, further distinguished into head, tail, and neck cell domains (249 mearurement each). (C) Correlation analysis of the Cd/C% signals, with semen Cd levels, for each cell domain. p values refer to Spearman’s correlation test.

FIGURE 3

(A) Induction coupled plasma spectroscopy analysis of cell Cadmium (Cd) content in sperm cells from normozoospermic healthy donors (n = 4) at basal (CTRL), after in vitro exposure to Cd 0.5 μM in for 1 hour at 37°C, and after further scavenging treatment with hypo-osmotic swelling (HOS) procedure or after 1-h treatment with 10 μM reduced glutathione (r-Glut). (B) Effect of Cd exposure (1 h at 37°C at 0.1 or 0.5 μM) on sperm progressive motility, monitored up to 2 h after exposure. In control condition (CTRL), Cd was omitted. Data are reported as percentage fold increase in the common basal condition. The effect of 10 μM r-Glut was also evaluated. Each experiment was performed on five different normozoospermic healthy donors. Significance: p values among the indicated condition are reported; n.s. = p > 0.05 between the indicated conditions.