Protamine contents and P1/P2 ratio in human spermatozoa from smokers and non-smokers

Protamine content is necessary for proper sperm chromatin condensation and subsequent male fertility. The exact effect of smoking on male fertility remains controversial. The objective of this study was to evaluate the effect of smoking on protamine content of sperm in smoker and non-smoker patients.

Protamines 1 (P1) and 2 (P2) were quantified by gel electrophoresis in the sperm of 53 smokers and 63 non-smokers. Sperm DNA fragmentation was analyzed employing the terminal deoxynucleotidyltransferase-mediated dUTP nick-end labelling (TUNEL) assay and non-condensed chromatin was evaluated using chromomycin A(3) (CMA(3)). Levels of smoking and oxidative stress markers were determined in seminal plasma using an enzyme linked immunosorbant assay (ELISA) and chemical reactions.

Protamine 2 concentrations were significantly lower (P < 0.050) in smokers than in non-smokers. In contrast P1/P2 ratios were significantly higher (P < 0.010) in smokers (1.34 ± 0.46 ng/10(6) sperm) than in non-smokers (1.11 ± 0.20 ng/10(6) sperm). The oxidative stress and smoking markers, reactive oxygen species (ROS), malondialdehyde, 8-Hydroxyguanosine (8-OHdG) and cotinine were significantly higher (P < 0.010) in smokers than in non-smokers, and correlated significantly (P < 0.050) with P1/P2 ratios. P2 showed significant negative (P < 0.050) correlations with ROS, 8-OHdG and cotinine. CMA(3) and TUNEL were also significantly higher (P < 0.010) in smokers (36.4 ± 8.1 and 17.4 ± 5.3%) than in non-smokers (29.8 ± 7.1 and 11.3 ± 4.2%).

This is the first study to evaluate the effect of smoking on protamines. Abnormal elevation of the P1/P2 ratio appears to be associated with aberrant P2 expression in smokers. These results suggest that induced oxidative stress by cigarette smoking may have significant inverse effect on the protamination process by disrupting P2.