Abnormal Sperm Chromatin Research Articles

Protamine 2 deficiency results in Septin 12 abnormalities.

There is a well-established link between abnormal sperm chromatin states and poor motility, however, how these two processes are interdependent is unknown. Here, we identified a possible mechanistic insight by showing that Protamine 2, a nuclear DNA packaging protein in sperm, directly interacts with cytoskeletal protein Septin 12, which is associated with sperm motility. Septin 12 has several isoforms, and we show, that in the Prm2 -/- sperm, the short one (Mw 36kDa) is mis-localized, while two long isoforms (Mw 40 and 41kDa) are unexpectedly lost in Prm2 -/- sperm chromatin-bound protein fractions. Septin 12 co-immunoprecipitated with Protamine 2 in the testicular cell lysate of WT mice and with Lamin B1/2/3 in co-transfected HEK cells despite we did not observe changes in Lamin B2/B3 proteins or SUN4 expression in Prm2 -/- testes. Furthermore, the Prm2 -/- sperm have on average a smaller sperm nucleus and aberrant acrosome biogenesis. In humans, patients with low sperm motility (asthenozoospermia) have imbalanced histone-protamine 1/2 ratio, modified levels of cytoskeletal proteins and we detected retained Septin 12 isoforms (Mw 40 and 41kDa) in the sperm membrane, chromatin-bound and tubulin/mitochondria protein fractions. In conclusion, our findings present potential interaction between Septin 12 and Protamine 2 or Lamin B2/3 and describe a new connection between their expression and localization, contributing likely to low sperm motility and morphological abnormalities.

Methylation Status of cAMP-responsive Element Modulator (CREM) Gene in Infertile Men and Its Association with Sperm Parameters.

The methylation pattern of non-imprinting genes was little studied, although it is widely known that the abnormal methylation levels of imprinting genes are associated with different forms of male infertility. The purpose of this research was to assess the CREM gene's methylation status and seminal characteristics in infertile individuals who were potential intracytoplasmic sperm injection (ICSI) candidates. A total of 45 semen samples (15 normospermia, 15 asthenospermia, and 15 oligoasthenoteratospermia) were examined. Using aniline blue (AB) staining, we carried out conventional semen analysis, chromatin quality, and sperm maturity testing. DNA was taken from semen samples, and all isolated DNA was assessed using Nanodrop and gel electrophoresis. A quantitative methylation-specific polymerase chain reaction (Q-MSP) approach was used to quantify the methylation at the DMRs of the CREM gene. According to our findings, sperm count (P=0.012), concentration (P= 0.019), motility (P=0.006), progression (P=0.006), and normal morphology (P=0.004) were all inversely correlated with abnormal sperm chromatin condensation. Additionally, we noted that the methylation level of the CREM gene was considerably more significant in the oligoasthenoteratospermia group compared to the asthenospermia and normospermia groups (P<0.05). Additionally, sperm count (P=0.043), progression (P=0.026), and normal morphology (P=0.024) were all inversely linked with CREM methylation. Overall, the abnormal CREM methylation patterns have a negative impact on sperm parameters. Additionally, the CREM gene's DNA methylation status may serve as an epigenetic indicator of male infertility.

The Relationship between Body Mass Index, Metal Elements, and Antioxidant Capacity of Semen on The Human Sperm Chromatin.

Sperm chromatin abnormalities are defects in nuclear maturation and DNA integrity. These defects originated from defective spermatogenesis due to a lack of DNA repair during chromatin remodeling. Changes in semen elements can cause damage to chromatin. There is little information about the relationship between changes in trace metal elements and total antioxidant capacity (TAC) with sperm chromatin damage. The present study was conducted to determine the relationship between Selenium (Se), Iron (Fe), Zinc (Zn), Copper (Cu) and the TAC of semen with the status of human sperm chromatin. In this experimental study, semen samples (n=30) were collected from healthy men referred to Kermanshah Motazadi Hospital and stored in liquid nitrogen; after thawing and centrifugation, sperm were separated. The atomic absorption method was used to measure the concentration of metal elements. The TAC was evaluated using the ferric-reducing antioxidant capacity of the plasma method. Furthermore, the integrity of sperm chromatin was measured using the sperm DNA fragmentation (SDF) method. The status of sperm chromatin had a non-significant correlation with body mass index (BMI, P=0.25, r=0.21) and a non-significant negative correlation with sperm count (P=0.71, r=-0.71) and motility (P=0.75, r=0.61). In addition, there was no significant relationship between sperm chromatin and the TAC of semen (P=0.92, r=0.01). Additionally, there was no significant correlation between Se, Zn, or Cu concentration (P>0.05) and Fe concentration, which had a partially positive relationship with the chromatin state of sperm (P=0.24, r=0.20). The trace metal elements in the seminal fluid did not play a significant role in the status of sperm chromatin.

TLR-1, TLR-2, and TLR-6 MYD88-dependent signaling pathway: A potential factor in the interaction of high-DNA fragmentation human sperm with fallopian tube epithelial cells.

The DNA integrity of spermatozoa that attach to fallopian tube (FT) cells is higher than spermatozoa that do not attach. FT epithelial cells can distinguish normal and abnormal sperm chromatin. This study investigated the effects of sperm with a high-DNA fragmentation index (DFI) from men with unexplained repeated implantation failure (RIF) on the Toll-like receptor (TLR) signaling pathway in human FT cells in vitro. Ten men with a RIF history and high-DFI and 10 healthy donors with low-DFI comprised the high-DFI (>30%) and control (<30%) groups, respectively. After fresh semen preparation, sperm were co-cultured with a human FT epithelial cell line (OE-E6/E7) for 24 hours. RNA was extracted from the cell line and the human innate and adaptive immune responses were tested using an RT2 profiler polymerase chain reaction (PCR) array. The PCR array data showed significantly higher TLR-1, TLR-2, TLR-3, TLR-6, interleukin 1α (IL-1α), IL-1β, IL-6, IL-12, interferon α (IFN-α), IFN-β, tumor necrosis factor α (TNF-α), CXCL8, GM-CSF, G-CSF, CD14, ELK1, IRAK1, IRAK2, IRAK4, IRF1, IRF3, LY96, MAP2K3, MAP2K4, MAP3K7, MAP4K4, MAPK8, MAPK8IP3, MYD88, NFKB1, NFKB2, REL, TIRAP, and TRAF6 expression in the high-DFI group than in the control group. These factors are all involved in the TLR-MyD88 signaling pathway. The MyD88-dependent pathway through TLR-1, TLR-2, and TLR-6 activation may be one of the main inflammatory pathways activated by high-DFI sperm from men with RIF. Following activation of this pathway, epithelial cells produce inflammatory cytokines, resulting in neutrophil infiltration, activation, phagocytosis, neutrophil extracellular trap formation, and apoptosis.

Sperm Chromatin Condensation Defect Accelerates the Kinetics of Early Embryonic Development but Does Not Modify ICSI Outcome.

The origin and quality of gametes are likely to influence the kinetics of embryonic development. The purpose of the study was to assess the impact of sperm nuclear quality, and in particular sperm chromatin condensation, on the kinetics of early embryo development after intracytoplasmic sperm injection (ICSI). Our study included 157 couples who benefitted from ICSI for male factor infertility. Chromatin condensation and DNA fragmentation were assessed in spermatozoa prior to ICSI. Above the 20% threshold of sperm condensation defect, patients were included in the abnormal sperm chromatin condensation (ASCC) group; below the 20% threshold, patients were included in the normal sperm chromatin condensation (NSCC) group. After ICSI, the oocytes were placed in the time-lapse incubator. The kinetics of the cohort's embryonic development have been modeled. The fading times of pronuclei and the time to two blastomeres (t2, first cleavage) and four blastomeres (t4, third cleavage) differed significantly between the NSCC and ASCC groups, with earlier events occurring in the ASCC group. On the other hand, the state of sperm chromatin condensation did not seem to have an impact on live birth rates or the occurrence of miscarriages. The kinetics of early embryonic development was accelerated in males with a sperm chromatin condensation defect without compromising the chances of pregnancy or promoting miscarriage. However, our study highlights the paternal contribution to early embryonic events and potentially to the future health of the conceptus.

Using Deep Learning Algorithm: The Study of Sperm Head Vacuoles and Its Correlation with Protamine mRNA Ratio.

Objective It is necessary to evaluate fertility effective agents to predict assisted reproduction outcomes. This study was designed to examine sperm vacuole characteristics, and its association with sperm chromatin status and protamine-1 (PRM1) to protamine-2 (PRM2) ratio, to predict assisted pregnancy outcomes. Materials and Methods In this experimental study, ninety eight semen samples from infertile men were classified based on Vanderzwalmen’s criteria as follows: grade I: no vacuoles; grade II: <2 small vacuoles; grade III: <1 large vacuole and grade IV: large vacuole with other abnormalities. The location, frequency and size of vacuoles were assessed using high magnification, a deep learning algorithm, and scanning electron microscopy (SEM). The chromatin integrity, condensation, viability and acrosome integrity, and protamination status were evaluated for vacuolated samples by toluidine blue (TB) staining, aniline blue, triple staining, and CMA3 staining, respectively. Also, Protamine-1 and protamine-2 genes expression was analysed by reverse transcription-quantitative polymerase chain reaction (PCR). The assisted reproduction outcomes were also followed for each cycle. Results The results show a significant correlation between the vacuole size (III and IV) and abnormal sperm chromatin condensation (P=0.03 and P=0.02, respectively), and also, protamine-deficient (P=0.04 and P=0.03, respectively). The percentage of reacting acrosomes was significantly higher in the grades III and IV spermatozoa in comparison with normal group. The vacuolated spermatozoa with grade IV showed a high protamine mRNA ratio (PRM-2 was underexpressed, P=0.01). In the IVF cycles, we observed a negative association between sperm head vacuole and fertilization rate (P=0.01). This negative association was also significantly observed in pregnancy and live birth rate in the groups with grade III and IV (P=0.04 and P=0.03, respectively). ConclusionThe results of our study highlight the importance sperm parameters such as sperm head vacuole characteristics, particularly those parameters with the potency of reflecting protamine-deficiency and in vitro fertilization/ intracytoplasmic sperm injection (IVF/ICSI) outcomes predicting.

Male Factors: the Role of Sperm in Preimplantation Embryo Quality.

Few studies have been conducted on the paternal effects on embryogenesis as compared with studies on maternal effects. The fertility potential of sperm decreases with genomic material abnormalities. Damaged DNA of sperm has been correlated with poor fertilization, reduced implantation and pregnancy rates, and increased production of aneuploid embryos. Evidence suggests that the role of sperm in embryogenesis goes beyond genomic material transfer, and centrosomes, sperm-derived cytoplasmic factors, paternal mRNA, and small RNAs are essential for early embryonic development. Epigenetic factors like histone modification and DNA methylation participate in the regulation of gene expression in sperm. The etiology of sperm chromatin abnormalities is important in male fertility and may affect reproductive outcomes. Success in implantation depends on the quality of the fertilized sperm and oocyte as well as the type of assisted reproductive techniques. Therefore, male factors affecting development of embryo can play a role in the failure of assisted reproductive techniques. Further studies are needed to evaluate clinical aspects and the risks of transmitting genetic or epigenetic disorders to provide safe therapies for infertility.

Relationship between Sperm Parameters with Sperm Function Tests in Infertile Men with at Least One Failed Cycle after Intracytoplasmic Sperm Injection Cycle.

BackgroundImbalance between production of reactive oxygen species (ROS) and total antioxidant capacity in testis, epididymis, and seminal fluid can eventually lead to infertility. Abnormal sperm chromatin packaging, and DNA fragmentation is considered as the main underlying causes of infertility. Therefore, we aimed to assess relationship between sperm parameters with DNA damage, protamine deficiency, persistent histones, and lipid peroxidation in infertile men with at least one failed cycle after intracytoplasmic sperm injection (ICSI). Materials and MethodsIn this experimental study, semen samples were collected from infertile men with at least one failed intracytoplasmic sperm injection (ICSI) cycle (n=20). Sperm parameters, DNA damage, protamine de- ficiency, persistent histones, and lipid peroxidation were assessed using computer-assisted sperm analysis (CASA) system, sperm chromatin structure assay (SCSA) and Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assays, chromomycin A3, aniline blue, and BODIPY C11 staining, respectively.ResultsA negative significant correlation was observed between sperm concentration with percentage of sperm persistent histones (r=-0.56, P=0.02), while positive significant correlations were found between percentage of sperm persistent histones with percentage of abnormal morphology (r=0.54, P=0.02), CMA3-positive spermatozoa (r=o.6, P=0.008) and intensity of lipid peroxidation (r=0.6, P=0.01). In addition, significant correlation was observed be- tween sperm DNA damage with intensity and percentage of lipid peroxidation (r=0.62, P=0.009). Correlation between CMA3-positive spermatozoa and intensity of lipid peroxidation (r=0.5, P=0.03) were also significant.ConclusionObserved significant correlations between sperm functional tests in infertile men with at least one failed ICSI cycle, indicated that the reduction of oxidative stress by antioxidant supplementation may be considered as one therapy approach that can improve sperm function and increase the chance of successful clinical outcomes in next assisted reproductive cycle.

New Markers for Predicting Fertility of the Male Gametes in the Post Genomic Era.

A number of tests have been proposed to assess male fertility potential, ranging from routine testing by light microscopic method for evaluating semen samples, to screening test for DNA integrity aimed to look at sperm chromatin abnormalities. Spermatozoa are an extremely differentiated cell population, having critical functions for embryo development and heredity, apart from giving haploid paternal DNA to the oocyte. Some aspects are essential to set this specific purpose. The ability of spermatozoa to perform its reproductive function takes place in the spermatogenesis, a highly specialized process depending on multiple factors with effect on male fertility. In the past 30 years, large-scale analyses of transcriptomic and proteomic analyses in mammals have generated a large amount of information on numerous biomolecules involved in spermatogenesis and male germ cell reproductive function. Sperm proteome represents the protein content that spermatozoa needs to survive and work correctly and modifications of sperm proteome play a role in determining functional changes leading to a decrease of reproductive competence into affected spermatozoa. The post-genomic approach consists of different methodologies for concurrently testicular transcriptome studies, protein compositional analysis and metabolomics findings of the spermatozoa in humans.

The effect of human sperm chromatin maturity on ICSI outcomes.

Because sperm chromatin may play a key role in reproductive success, we verify the associations between sperm chromatin abnormalities, embryo development and the ability to achieve pregnancy. The evaluation of sperm chromatin maturity using aniline blue (AB), chromomycin A3 (CMA3) and toluidine blue (TB) staining were carried out in group of males from infertile couples that underwent ICSI. Low levels of sperm chromatin abnormalities (<16%) were found in most subjects (>50%). A higher percentage of TB-positive sperm cells were discovered in the men from couples who achieved≤50% fertilized oocytes compared to men who achieved>50%. No significant differences were discovered by the applied tests between the men from couples who achieved≤50% and those who achieved>50% high-quality embryos on the 3rd or 5th day after fertilization, nor between the men from couples who achieved pregnancy and those who failed. The sperm chromatin maturity did not correlate with the ICSI results. However, the ROC analysis revealed a significant predictive value of TB-positive spermatozoa only for fertilization. Therefore, the TB assay can be considered as a useful test for the prediction of fertilization. Our findings suggest that the level of sperm chromatin abnormalities of the examined men was not clinically significant. No found associations between sperm chromatin maturity and embryo development and the ability to achieve pregnancy. We could not exclude the effects of the repairing processes in the fertilized oocyte. The use of complementary tests that verify the status of the sperm chromatin seems justified.

Is there any correlation between sperm parameters and chromatin quality with embryo morphokinetics in patients with male infertility?

The main goal was to evaluate the correlation between sperm parameters and chromatin quality with embryo kinetics via time-lapse monitoring system (TLM). A total of 40 couples involved in the ICSI program as a result of male infertility. For assessment of sperm chromatin and DNA quality, we used aniline blue, toluidine blue, chromomycin A3, acridine orange and terminal transferase-mediated deoxyuridine triphosphate biotin end labelling assays. All mature oocytes were injected, and the generated zygotes (2PNs) were cultured in TLM. In day 3 after injection, single embryo transfer (SET) was carried out according to the morphology and morphokinetics. The patients were followed up until delivery. There were positive significant correlations between sperm count with CC2 (r=.330, p=.049), T4 (r=.329, p=.038), T6 (r=.342, p=.035) and T7 (r=.374, p=.025). Also, there were positive significant correlations between nonprogressive motility and T2 (r=0.323, p=.042), T3 (r=.411, p=.013) and T4 (r=.418, p=.007). Regarding the sperm chromatin quality assays, there were negative significant correlations between CMA3 and CC2 (r=-.272, p=.049) and between acridine orange and T5 (r=-.221, p=.040). It seems that the abnormal sperm parameters and chromatin alteration affect the normal embryo kinetics in ICSI program.

The Influence of Advanced Paternal Age on Sperm Chromatin Integrity and Early Embryo Morphological Development during ICSI

The study determined the impact of advanced male ageing (≥50 years) on sperm chromatin integrity and early embryo morphological development in intra-cytoplasmic sperm injection (ICSI) technique cycles. Six hundred subfertile men were age-grouped; X1 (50 to 59 years), X2 (60 to 69), and X3 (≥70), were compared with 600 fertile males of known fertility (Y, age 25 - 35 years). Oocytes from 254 women, aged ≤ 30 years, were included. Sperm were analyzed using acridine orange fluorescence test (AOT) and categorized: “low”, “inter-mediate” and “high” damage. After ICSI, embryos were evaluated and categorized as “good”, “fair” or “poor” quality. Embryonic morphological development was assessed at three stages: fertilization, early and late paternal effect. The AOT results were: X1: low = 29, intermediate = 53 and high = 118; X2: low = 11, intermediate = 42 and high = 147; X3: low = 8, intermediate = 24 and high = 168; Y: Low = 486, intermediate = 71 and high = 43. The fertilization rate was: X1, 329/350 (93.7%); X2, 298/350 (85.1%); X3, 225/350 (64.1%) and, Y, 350/350 (100%). Associations between increasing age and sperm chromatin damage (χ2 (723.249, 6) p < 0.0001), increasing age and inability to fertilize (χ2 (210.990, 3) p < 0.0001) were observed. Associated with increasing age was the significant proportion of morphologically poor quality embryos over the five days after fertilization. Male age ≥ 50 years, is highly associated with abnormal sperm chromatin organization, an inability to adequately fertilize with ICSI methodology, an increase in the number of poor quality embryos and, a corresponding decrease in the number of good quality embryos five days after fertilization.

The effect of cigarette smoking on human seminal parameters, sperm chromatin structure and condensation.

Considerable debate still exists regarding the effects of cigarette smoking on male fertility. This work aimed to explore effects of cigarette smoking on semen parameters and DNA fragmentation on 95 infertile patients who were divided into infertile male nonsmokers (45) and infertile male smokers (50). Smokers were subdivided according to a number of cigarettes smoked per day into mild (≤10), moderate (11-20) and heavy smokers (≥21). Semen analysis, sperm chromatin condensation integrity with aniline blue staining and sperm viability were compared between the study groups. A significant decrease has been shown in sperm count (p=.006), progressive motility (p=<.001), percentage of normal forms (p=<.001) and viability (p=.002) between infertile nonsmoker and infertile smokers. The percentage of abnormal sperm chromatin condensation was significantly higher in smokers compared to nonsmokers (p=<.001). A linear correlation was detected between the extent of cigarette smoking and the degree of worsening in progressive motility (p=.001), total motility (p<.001), viability (p<.001) and normal morphology (p<.001). These results indicate that cigarette smoking has detrimental effects on semen parameters. It negatively affected all conventional semen parameters in addition to sperm chromatin condensation and sperm viability. These abnormalities were also proportional to the number of cigarettes smoked per day and to the duration of smoking.

Effects of Chlamydia trachomatis infection on sperm chromatin condensation and DNA integrity.

The present study was performed to investigate the relation of Chlamydia trachomatis infection to sperm chromatin/DNA integrity in a population of infertile men (male partner of infertile couples) from Iran. Blood, semen and first-void urine samples were obtained from 250 infertile men. Data were analysed with regard to the results of (i) serological analysis for specific antibodies to C.trachomatis in serum; (ii) the presence of C.trachomatis and DNA in first-void urine; and (iii) in the semen sample of the male partner, in addition to sperm analysis, four different tests (aniline blue, chromomycin A3, acridine orange and TUNEL) were used to detect sperm chromatin and DNA abnormalities. The main conclusions of the results were: (i) no evidence of C.trachomatis infection in semen samples was found; (ii) sperm DNA fragmentation and chromatin studies were not correlated with C.trachomatis diagnosis; (iii) the percentage of DNA fragmentation is positively correlated with the percentage of immotile sperm but negatively with semen volume, normal morphology; and (iv) in sperm chromatin evaluations, only the percentage of chromatin protamination was related to male age.

SLY regulates genes involved in chromatin remodeling and interacts with TBL1XR1 during sperm differentiation

Sperm differentiation requires unique transcriptional regulation and chromatin remodeling after meiosis to ensure proper compaction and protection of the paternal genome. Abnormal sperm chromatin remodeling can induce sperm DNA damage, embryo lethality and male infertility, yet, little is known about the factors which regulate this process. Deficiency in Sly, a mouse Y chromosome-encoded gene expressed only in postmeiotic male germ cells, has been shown to result in the deregulation of hundreds of sex chromosome-encoded genes associated with multiple sperm differentiation defects and subsequent male infertility. The underlying mechanism remained, to date, unknown. Here, we show that SLY binds to the promoter of sex chromosome-encoded and autosomal genes highly expressed postmeiotically and involved in chromatin regulation. Specifically, we demonstrate that Sly knockdown directly induces the deregulation of sex chromosome-encoded H2A variants and of the H3K79 methyltransferase DOT1L. The modifications prompted by loss of Sly alter the postmeiotic chromatin structure and ultimately result in abnormal sperm chromatin remodeling with negative consequences on the sperm genome integrity. Altogether our results show that SLY is a regulator of sperm chromatin remodeling. Finally we identified that SMRT/N-CoR repressor complex is involved in gene regulation during sperm differentiation since members of this complex, in particular TBL1XR1, interact with SLY in postmeiotic male germ cells.

Administration of high dose of methamphetamine has detrimental effects on sperm parameters and DNA integrity in mice

Methamphetamine (MA) was shown to have harmful effects on male reproductive system. To investigate probable effects of daily administration of MA on sperm parameters and chromatin/DNA integrity in mouse. Thirty-five NMRI male mice were divided into five groups including low, medium, and high dosage groups which were injected intraperitoneally with 4, 8 and 15 mg/kg/day for 35 days, respectively. Normal saline was injected in sham group and no medications were used in control group. Then, the mice were killed and caudal epididymis of each animal was cut and placed in Ham's F10 medium for sperm retrieval. To evaluate sperm chromatin abnormalities, the aniline blue, toluidine blue and chromomycine A3 were used. For sperm DNA integrity and apoptosis, the acridine orange, sperm chromatin dispersion, and TUNEL assay were applied. For sperm morphology, Papanicolaou staining was done. Normal morphology and progressive motility of spermatozoa decreased in medium and high dosage groups in comparison with the control group (p=0.035). There was a significant increase in rate of aniline blue, toluidine blue, and chromomycine A3 positive spermatozoa in high dosage group. In a similar manner, there was an increase in rates of acridine orange, TUNEL and sperm chromatin dispersion positive sperm cells in high dosage group with respect to others. MA abuse in a dose-dependent manner could have detrimental effects on male reproductive indices including sperm parameters and sperm chromatin/DNA integrity in mice.

Assessment of human sperm DNA integrity using two cytochemical tests: Acridine orange test and toluidine blue assay.

Primary infertility affects approximately 15% of couples, with male factor infertility accounting for 50% of cases. Semen samples from 41 patients with asthenoteratospermia and 28 men with proven fertility were analysed according to World Health Organization guidelines. Abnormal sperm chromatin structure was assessed by toluidine blue assay (TBA), and DNA denaturation (DD) was detected by the acridine orange test (AOT). The mean (±SEM) rates of DD and abnormal chromatin structure were significantly higher in infertile subjects compared to fertile group respectively p=.003 and p<.001. A significant correlation was established between sperm DD and abnormal chromatin structure (R=.431, p<.001). Sperm DNA damage correlated significantly with abnormal morphology, sperm motility and necrozoospermia. Our study shows that men with increased levels of abnormal sperm chromatin structure have a high incidence of DNA denaturation and altered semen parameters. These findings suggest that male infertility has been linked to sperm DNA damage.

Fourier transform infrared spectroscopic analysis of sperm chromatin structure and DNA stability.

Sperm chromatin structure and condensation determine accessibility for damage, and hence success of fertilization and development. The aim of this study was to reveal characteristic spectral features coinciding with abnormal sperm chromatin packing (i.e., DNA-protein interactions) and decreased fertility, using Fourier transform infrared spectroscopy. Chromatin structure in spermatozoa obtained from different stallions was investigated. Furthermore, spermatozoa were exposed to oxidative stress, or treated with thiol-oxidizing and disulfide-reducing agents, to alter chromatin structure and packing. Spectroscopic studies were corroborated with flow cytometric analyses using the DNA-intercalating fluorescent dye acridine orange. Decreased fertility of individuals correlated with increased abnormal sperm morphology and decreased stability toward induced DNA damage. Treatment with the disulfide reducing agent dithiothreitol resulted in increased sperm chromatin decondensation and DNA accessibility, similar as found for less mature epididymal spermatozoa. In situ infrared spectroscopic analysis revealed that characteristic bands arising from the DNA backbone (ν1230, ν1086, ν1051cm(-1) ) changed in response to induced oxidative damage, water removal, and decondensation. This coincided with changes in the amide-I region (intensity at ν1620 vs. ν1640cm(-1) ) denoting concomitant changes in protein secondary structure. Reduction in protein disulfide bonds resulted in a decreased value of the asymmetric to symmetric phosphate band intensity (ν1230/ν1086cm(-1) ), suggesting that this band ratio is sensitive for the degree of chromatin condensation. Moreover, when analyzing spermatozoa from different individuals, it was found that the asymmetric/symmetric phosphate band ratio negatively correlated with the percentage of morphologically abnormal spermatozoa.

Effect of Glutathione Adding to Freezing Media on Sperm Parameters and on Malondialdehyde Concentration and Abnormal Sperm Chromatin Percent of Oligozoospermic Patients

Objective: The study aimed to study the effect antioxidant Glutathione adding to sperm Freezing Media ( Sperm Freeze) on Sperm Parameters and Malondialdehyde concentration and abnormal sperm chromatin percent of Oligozoospermic patients Methodology: This study was performed in the laboratory of Intra cytoplasmic Injection and freezing of sperm in the fertility center of Al-Sader Medical City / Al-najaf Alashraf City during the period from May, 2012 to April 2013 , The study included 30 of Oligozoospermic patients, the statistical analysis of this study performed by using Student's t-test and paired t-test at the level of probability 0.05 Results: The study results shows that used of sperm Freezing Media ( Sperm Freeze) with adding of 1 mm from antioxidant Glutathione leads to a significant difference (p&lt;0.05) in sperm parameters of study included patients, there was a significant increase (p&lt;0.05) in the sperm progressive motility percent and sperm viability percent ,and there was a significant decrease (p&lt;0.05) In The Malondialdehyde concentration and abnormal sperm chromatin percent and no significant difference (p&lt;0.05) In the sperm concentration, normal sperm morphology percent and round cell concentration In comparison with their Means when used of sperm freezing media ( Sperm Freeze) alone. Conclusion: It can be concluded from this study that sperm rapid freezing and thawing processes had a negative effect on the semen and sperm parameters and Malondialdehyde concentration and abnormal sperm chromatin percent in all samples of study included patients and adding of 1 mm from antioxidant Glutathione shows resistance to this negative effect leads to a significant increase (p&lt;0.05) In the studying parameters . Recommendation: using of antioxidant Glutathione with sperm Freezing Media (Sperm Freeze) in sperm cryopreservation processes

Sperm epigenetic profile and risk of cancer

Introduction and objective. The integrity, stability and composition of sperm chromatin are of great importance in the fertilizing potential of male gametes and their capacity to support normal embryonic development. In this study, the author presents the current state of knowledge about the sperm epigenetic profile and risk of cancer. Abbreviated description of the state of knowledge. The obtaining of pregnancy and the state of health of the baby depends on the quality of the genetic material of both the female and the male. Health behaviours and environmental factors directly affect the quality of sperm, as well as the human egg cell and, consequently, on the reproductive capabilities, the course of pregnancy and the state of the newborn. There exist two thoroughly investigated epigenetic modifications: DNA methylation and histone modifications. The process of DNA methylation can be also a fundamental factor contributing to the development of cancer, where epigenotype undergoes significant modifications. When considering numerous DNA aberrations in the male gamete, the most commonly encountered is DNA fragmentation, particularly in infertile subjects. Surprisingly, an intracytoplasmatic sperm injection study of mice oocytes, using spermatozoa with a high DNA Fragmentation Index (DFI), revealed that a considerable percentage of adults born as a result of this method, showed a significant increase in the incidence of abnormal behavioural tests, malformations, cancer and signs of premature aging. Summary. The issue of assisted procreation raises the need to look for an appropriate treatment for males with sperm chromatin abnormalities. As a result, the fight against smoking addiction becomes the obvious necessity. Moreover, the reasonable solution nowadays seems to be supplementation with micronutrients and folic acid. It has been proved that the process of DNA fragmentation is a phenomenon that intensifies over time. Therefore, there should be a pursuance for, as close as possible, to the moment of ejaculation, application of semen to reproductive techniques. Finally, epigenetic changes are suspected of being one of the factors responsible for the deterioration of male sperm parameters observed in recent decades.