Potential importance of transition metals in the induction of DNA damage by sperm preparation media

STUDY QUESTION What are the mechanisms by which the preparation of spermatozoa on discontinuous density gradients leads to an increase in oxidative DNA damage? SUMMARY ANSWER The colloidal silicon solutions that are commonly used to prepare human spermatozoa for assisted reproduction technology (ART) purposes contain metals in concentrations that promote free radical-mediated DNA damage. WHAT IS KNOWN ALREADY Sporadic reports have already appeared indicating that the use of colloidal silicon-based discontinuous density gradients for sperm preparation is occasionally associated with the induction of oxidative DNA damage. The cause of this damage is however unknown. STUDY DESIGN, SIZE, DURATION This study comprised a series of experiments designed to: (i) confirm the induction of oxidative DNA damage in spermatozoa prepared on commercially available colloidal silicon gradients, (ii) compare the levels of damage observed with alterative sperm preparation techniques including an electrophoretic approach and (iii) determine the cause of the oxidative DNA damage and develop strategies for its prevention. The semen samples employed for this analysis involved a cohort of >50 unselected donors and at least three independent samples were used for each component of the analysis. PARTICIPANTS/MATERIALS, SETTING, METHODS The setting was a University biomedical science laboratory. The major techniques employed were: (i) flow cytometry to study reactive oxygen species generation, lipid peroxidation and DNA damage, (ii) computer-aided sperm analysis to measure sperm movement and (iii) inductively coupled mass spectrometry to determine the elemental composition of sperm preparation media. MAIN RESULTS AND THE ROLE OF CHANCE Oxidative DNA damage is induced in spermatozoa prepared on PureSperm® discontinuous colloidal silicon gradients (P < 0.001 versus repeated centrifugation) because this medium contains metals, particularly Fe, Al and Cu, which are known to promote free radical generation in the immediate vicinity of DNA. This damage can be significantly accentuated by reducing agents, such as ascorbate (P < 0.001) and inhibited by selective chelation (P < 0.001). This problem is not confined to PureSperm®; analysis of additional commercial sperm preparation media revealed that metal contamination is a relatively constant feature of such products. LIMITATIONS, REASONS FOR CAUTION While the presence of metals, particularly transition metals, may exacerbate the levels of ox