Age-Related Dynamics of Typical Bioessential Elements in Mouse Sperm at Single-Cell Resolution

Bioessential elements are essential for maintaining reproductive function, yet their homeostasis may be perturbed during aging. The dynamic regulation of these elements during aging remains insufficiently characterized. Here, a multielement single-cell inductively coupled plasma mass spectrometry (SC-ICP-MS) method was established for the quantification of seven bioessential elements, including iron (Fe), copper (Cu), calcium (Ca), zinc (Zn), magnesium (Mg), phosphorus (P) and manganese (Mn). This method was optimized using an integrated strategy to balance sensitivity, transport efficiency, and signal-to-noise ratio, and subsequently applied to quantify elemental contents in sperm collected from mice at different ages. Distinct age-dependent patterns were observed at the single-sperm level. Fe, Cu, and Ca exhibited early life maxima, whereas Zn, Mg, and P reached peak levels in midlife; Mn remained consistently low across all age groups. Interestingly, elemental heterogeneity followed a conserved inverted U-shaped trajectory, reaching a maximum at week 40. Correlation network analysis revealed a significant association between Fe and Mn at the content level, whereas elevated P content and increased heterogeneity were associated with reduced sperm concentration and impaired morphology. Functionally, Zn content was positively correlated with sperm motility, whereas elevated P content and increased heterogeneity were associated with reduced sperm concentration and impaired morphology. Ca, P, Mn, and Fe were more susceptible to dysregulation linked to compromised sperm function, whereas Mg, Zn, and Cu were associated with beneficial outcomes. These findings provide single-cell insights into age-dependent elemental homeostasis and its functional implications for male reproductive aging.