The effects of bacterial infection on human sperm nuclear protamine P1/P2 ratio and DNA integrity.

The purpose of this study was to detect the effects of bacterial infection on human sperm nuclear protamines and DNA fragmentation. In this study, 120 semen samples were collected from unselected male partners of couples consulting for infertility in infertility and obstetrics clinic. All the samples were screened bacteriologically according to World Health Organization guidelines, and also sperm parameters and DNA fragmentation were evaluated. The concentrations of protamines P1 and P2 were quantified using acid urea acrylamide gel electrophoresis. Of a total number of 120 sample, 36 (30%) of them were infected with bacteria. Nine species of bacteria belonging to five genera, Staphylococcus, Escherichia, Streptococcus, Enterococcus and Klebsiella, were identified. The comparison between infected (36) and noninfected (84) samples appeared the negative impact of bacterial infection on sperm parameters and P1/P2 ratios. The percentages of P1/P2 ratio abnormality were significantly higher in infected patients. Sperm concentration, motility, progression and chromatin condensation were significantly lower in infected patients (p < .010). Depending on these results, we concluded that the bacterial infections have significant negative effects on sperm chromatin condensation and protamine P1/P2 ratio. Moreover, the negative relationship between the bacterial infections and sperm parameters, such as concentration, motility and progressive motility, has been shown.

Impact of cigarette smoking on histone (H2B) to protamine ratio in human spermatozoa and its relation to sperm parameters.

Smoking is strongly associated with abnormalities in histone-to-protamine transition and with alteration of protamine expression in human spermatozoa. A proper protamine to histone ratio is, however, essential for sperm chromatin maturity and DNA integrity. Alterations in these sperm nuclear proteins were observed in infertile men. The present prospective study is aimed at evaluating the possible relationship among smoking, semen quality and the histone-to-protamine transition ratio in mature spermatozoa. Histone H2B and protamine 1 (P1) and 2 (P2) were quantified using acid-urea polyacrylamide gel electrophoresis in the spermatozoa of 35 smokers and 19 non-smokers. Levels of lipid peroxidation marker malondialdehyde (MDA) were measured in seminal plasma by thiobarbituric acid assay. Cotinine concentrations were determined in seminal plasma using an enzyme-linked immunosorbent assay. Histone H2B levels in smokers (292.27 ± 58.24 ng/10(6)) were significantly higher (p = 0.001) than that of non-smokers (109.1 ± 43.70 ng/10(6)), besides, a significant difference (p > 0.0001) was found for the P1 and P2 ratio between smokers (1.71 ± 0.071) and non-smokers (1.05 ± 0.033). The H2B/(H2B+P1 + P2) ratio (0.29 ± 0.71) of smokers were significantly higher (p = <0.0001) than that of non-smokers (0.12 ± 0.01). The concentrations of MDA (µm) (7.13 ± 1.15) and cotinine (ng/mL) (60.44 ± 31.32) in seminal plasma of smokers were significantly higher (p = 0.001) than those in the samples of the non-smoker group (4.42 ± 1.16 and 2.01 ± 2.84 respectively). In addition, smokers showed significantly (p ≤ 0.002) lower sperm count, motility (p = 0.018), vitality (p = 0.009) and membrane integrity (p = 0.0001) than non-smokers. These results reveal that patients who smoke possess a higher proportion of spermatozoa with an alteration of the histone to protamine ratio than patients who do not smoke, and suggest that cigarette smoking may inversely affect male fertility.

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

BACKGROUND: 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. METHODS: 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. RESULTS: 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%). CONCLUSIONS: 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.