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Sperm head defects and disturbances in spermatozoal chromatin and DNA integrities in idiopathic infertile subjects: association with cigarette smoking

Date: 12-6-2008

Elshal MF, El-Sayed IH, Elsaied MA, El-Masry SA, Kumosani TA. Molecular Biology Department, Genetic Engineering and Biotechnology Research Institute, Menoufiya University, Egypt. .Clin Biochem. 2009 May;42(7-8):589-94. Epub 2008 Dec 6.

CONCLUSION: DFI%, HDS% and round-head sperms are increased in idiopathic infertile men; this increase is associated with cigarette smoking. These defects may be attributed to increased oxidative stress and insufficient scavenging antioxidant enzymes in the seminal fluid of infertile patients.

OBJECTIVES: To evaluate sperm chromatin and DNA integrities in idiopathic infertile men and determine the possible association(s) of cigarette smoking on oxidative stress markers, antioxidant capacity and semen quality. SUBJECTS AND METHODS: Semen samples from men referring to the andrology laboratory were categorized into 3 groups: fertile non-smokers (n=16), infertile non-smokers (n=36), and infertile smokers (n=34). Semen analysis was performed according to WHO criteria. The percentage of sperm DNA fragmentation index (%DFI) and the percentage of sperm with abnormally high DNA stainability (HDS%; immature spermatozoa) were determined by SCSA using the metachromatic properties of acridine orange. Lipid peroxidation, superoxide dismutase (SOD), catalase (CAT) and reduced glutathione (GSH) levels in seminal plasma and spermatozoa were measured by spectrophotometer assays. RESULTS: The classical semen parameters were negatively correlated with lipid peroxidation in spermatozoa; motility and morphology were negatively correlated with %DFI (p<0.05). HDS% was also negatively correlated with above markers except for morphology (r=-0.352, p=0.081). DFI% and HDS% were significantly higher in the infertile smokers group than in infertile non-smokers (p=0.032; p=0.001 respectively). Cigarette smoking was significantly associated with DFI%, HDS%, TBARS and the fraction of "round-headed" sperm (r=0.796, p=0.0001; r=0.371, p=0.033; r=0.606, r=0.591, p=0.001 respectively), and decreased SOD levels (r=-0.545). CONCLUSION: DFI%, HDS% and round-head sperms are increased in idiopathic infertile men; this increase is associated with cigarette smoking. These defects may be attributed to increased oxidative stress and insufficient scavenging antioxidant enzymes in the seminal fluid of infertile patients

Oxidative Stress Responsible for Significant Amount of DNA Damage in Human Spermatozoa (from J. Aitken laboratory, Australia).
DNA damage in human spermatozoa has been linked to impaired fertilization, disturbed preimplantation embryonic growth, poor implantation rates, high miscarriage rates, elevated rates of morbidity in offspring, and reduced fertility in vivo and in vitro.  The cause of this DNA damage is unclear; however, physical factors including heat and electromagnetic radiation, in addition to a range of xenobiotics, abnormalities in lipid metabolism, and age have been suggested as possible factors in this process. The molecular basis of DNA fragmentation seen in this patient population also remains unresolved.
"Clinically, the relevance of the present results is that they suggest a significant proportion of the DNA damage observed in human spermatozoa is oxidative in nature," wrote De Julius et al. "This finding provides a solid rationale for use of antioxidants in the amelioration of such damage." 

Full abstract: Oxidative Stress Responsible for Significant Amount of DNA Damage in Human Spermatozoa

DNA damage in human spermatozoa has been linked to impaired fertilization, disturbed preimplantation embryonic growth, poor implantation rates, high miscarriage rates, elevated rates of morbidity in offspring, and reduced fertility in vivo and in vitro.  The cause of this DNA damage is unclear; however, physical factors including heat and electromagnetic radiation, in addition to a range of xenobiotics, abnormalities in lipid metabolism, and age have been suggested as possible factors in this process. The molecular basis of DNA fragmentation seen in this patient population also remains unresolved. In the present study, Geoffry N. De Iuliis and colleagues performed a study to test their hypothesis that there is a close association between the efficiency of chromatin remodeling, oxidative base damage to sperm DNA, and DNA fragmentation in human spermatozoa ("DNA Damage in Human Spermatozoa is Highly Correlated with the Efficiency of Chromatin Remodeling and the Formation of 8-Hydroxy-2'- Deoxyguanosine, a Marker of Oxidative Stress.  These results clearly highlight the importance of oxidative stress in the induction of sperm DNA damage and carry significant implications for the clinical management of this condition," wrote De Iuliis et al. Flow cytometry/fluorescence microscopy was used to examine DNA fragmentation, chromatin protamination, mitochondrial membrane potential, and the development of the oxidative base adduct, 8-hydroxy-2'-deoxyguanosine (8OHdG). Impairment of DNA protamination throughout late spermatogenesis was closely associated (P<0.001) with DNA damage in human spermatozoa. The disturbance of chromatin remodeling was also associated with a significant rise in levels of 8OHdG (P<0.001). Additionally, the latter itself closely corresponded with DNA fragmentation (P<0.001). The importance of oxidative stress in the formation of 8OHdG was displayed experimentally with H2O2/Fe2+ and by the relationship exhibited between this base adduct and superoxide generation (P<0.001). "That 8OHdG formation was inversely associated with mitochondrial membrane potential (P<0.001) suggested a possible role for these organelles in the creation of oxidative stress," stated the researchers.

"Clinically, the relevance of the present results is that they suggest a significant proportion of the DNA damage observed in human spermatozoa is oxidative in nature," wrote De Iuliis et al. "This finding provides a solid rationale for use of antioxidants in the amelioration of such damage."

Macrophage activity in semen is significantly correlated with sperm quality in infertile men
K. Tremellen and O. Tunc  Research Centre for Reproductive Health, Discipline of Obstetrics and Gynaecology, School of Paediatrics and Reproductive Health, University of Adelaide, Adelaide, SA, Australia , and  Repromed, Dulwich, SA, Australia

PARTIAL ABSTRACT
The presence of leucocytes within semen has the potential to impair sperm function. Neutrophils and macrophages make up 95% of seminal leucocytes, with both having the ability to damage sperm via the generation of reactive oxygen species, proteases and the induction of apoptosis.  Neopterin, a molecule released from activated macrophages, may be a useful marker for macrophage activity in the male reproductive tract. To examine this possibility a total of 63 asymptomatic subjects with male factor infertility and 11 fertile controls provided semen samples for measurement of various inflammatory markers. We were able to confirm for the first time that seminal plasma does indeed contain neopterin and that the levels of this macrophage activity marker are threefold higher in infertile than fertile men. Furthermore, seminal plasma neopterin concentration was significantly correlated with sperm oxidative stress, DNA fragmentation (TUNEL) and apoptosis (Annexin V), making it a useful marker of sperm quality.