Association between a marker of sperm DNA damage and sperm indices in infertile males in Benin City, Nigeria: A cross-sectional study

Abstract Background Studies have shown oxidative DNA damage is associated with male infertility. Objective This study determines the levels of 8-hydroxy-2'-deoxyguanosine (8-OHdG) and some markers of oxidative stress in seminal fluid of males investigated for infertility and men of proven fertility in Benin City, Nigeria. Materials and Methods Semen samples produced by self or assisted masturbation were analyzed by microscopic technique according to the World Health Organization guidelines. Thereafter, samples were centrifuged and seminal fluid plasma separated and stored at -20°C prior to assay for 8-OHdG and oxidative stress biomarkers. Based on the sperm concentration/count, the overall samples were grouped into the following categories: normospermia (n = 20), oligozoospermia (n = 30), and azoospermia (n = 20). The control group comprised of 30 age-matched males of proven fertility. The seminal fluid 8-OHdG, total antioxidant status, superoxide dismutase and malondialdehyde (MDA) were assayed through ELISA and spectrophotometric methods, respectively. Results Seminal plasma level of 8-OHdG and MDA were significantly higher (p = 0.01) in infertile subjects than controls. The mean levels of 8-OHdG and MDA in infertile subjects were higher in azoospermia than oligospermia than normospermia and so, was least in the normospermia. Conversely, the mean levels of total antioxidant status and superoxide dismutase were significantly lower (p = 0.01) in infertile than fertile the control male subjects with levels higher in normospermia than oligospermia and least in azoospermia. Moreover, the seminal 8-OHdG correlated negatively with sperm count (r = -0.359, p = 0.01), percent motility (r = -0.388, p = 0.04), and percent morphology (r = -0.327, p = 0.02). Conclusion The assessment of sperm DNA damage in addition to routine seminal fluid analysis may play an important role in specific diagnosis and management of male infertility.


Introduction
Very few studies in Nigeria have investigated the role of sperm nuclear DNA integrity among infertile males. It has been suggested that sperm DNA integrity may be a potential predictor of male fertility and/or may serve as an adjunct to routine semen analysis. It has also been reported that some men who were unable to achieve pregnancy had sperm concentrations within normal range (1). Some have suggested that sperm of infertile men contain more DNA damage than fertile men and as such may have negative effects on fertility potential of these individuals (2,3).
Male infertility is present in approximately half of all infertile couples. Currently, the routine analysis of male infertility includes aphysical examination and semen evaluation for sperm concentration, morphology, seminal volume, motility, and pH (4). Although, about 15% of patients with male factor infertility have been reported to have normal semen analysis (5), a conclusive and definitive diagnosis of male factor infertility can only be made through a routine semen analysis (4).
Most laboratories in our setting currently rely on microscopic examination of semen in the diagnosis and management of the male infertility.
This baseline microscopic semen analysis may not sufficient to detect the cause (s) since the etiology of male infertility is multifactorial.
The etiologies of male infertility could be anatomic defects, hormonal abnormalities, immune disorders, gene mutation, radiation, chemotherapy, ejaculatory failures, and environmental toxicants (6). In some cases, the identification of the exact cause (s) of male infertility may not be obvious and the established processes that led to the poor semen quality remain unclear (7). A standard semen analysis using a light microscope has been widely used in most laboratories for the initial assessment of male fertility potential; however, detecting defective sperm function by standard semen analysis is problematic because the spermatozoon is a highly specialized cell that exhibits manifold array of biological properties to achieve fertilization.
In addition, results of standard semen analyses are selfhood or internal and subject to intra-and inter-observer variability (8

SOD ELISA kit (Elabscience; catalog number E-EL_H 1113) Principle
The assay method is based on the Competitive

MDA assay kit (NWLSS, Canada) Principle
This assay is based on the reaction, n of MDA with thiobarbituric acid (TBA), forming a MDA-TBA2 adduct that absorbs strongly at 532 nm.

Ethical considerations
All included participants were informed about the nature of the study and informed consent was obtained prior to the specimens were collected.

Statistical analysis
Data generated from the study were compared between the groups using the analysis of variance A p-value ≤ 0.05 was considered as statistically significant.

Results
No statistically significant differences were seen in age and semen volume between groups. The sperm concentration, motility and morphology percentages were significantly higher among normospermia than oligospermia (p = 0.01) ( Table I). Table II (Table III).

International Journal of Reproductive BioMedicine
Yusuf et al.

Discussion
The routine laboratory evaluation of males who failed to achieve conception naturally for one or more years includes at least two semen analyses for sperm count, percent motility, and percent morphology in order to establish the reproductive potential of the subjects. Assessment of markers of sperm DNA damage, oxidative stress, lipid peroxidation, and SOD activity are not routinely done even though their roles in male infertility are recognized (17)(18)(19).
The results from this study indicate that there is an evidence of oxidative DNA damage, higher

Conclusion
Seminal