Sperm Oxidative Stress Research Articles

Malathion-induced spermatozoal oxidative damage and alterations in sperm quality of endangered trout Salmo coruhensis.

The use of pesticides has been increased along with increasing the farming activities and has caused environmental impacts deleteriously. In particular, non-target organisms including fish can be affected by toxic effects of pesticides. Therefore, the impacts of malathion (MTN) on oxidative stress and sperm quality were investigated in vitro. The MTN concentrations used on this study were 0 (control), 75, 100, and 125μg/L. Lipid peroxidation (MDA), non-enzymatic (GSH), and enzymatic (SOD, GSH-Px, and CAT) activities in spermatozoa were examined for determination of oxidative stress status. Our findings showed that motility rate and period of sperm cells significantly decreased with exposure to MTN. Biochemical assays revealed that CAT activity and levels of MDA, GSH increased in spermatozoa based on concentration while activity of GSH-Px and SOD decreased. Consequently, spermatozoa were highly sensitive to MTN exposure. MTN has disruptive effects on sperm quality and caused to oxidative stress in spermatozoa.

Effect of the organophosphate insecticide chlorpyrifos exposure on oxidative stress and quality of Salmo coruhensis spermatozoa

The use of pesticides has been increasing along with increasing farming activities and has caused deleterious environmental impacts. Non-target organisms in particular, including fish, are affected by pesticides. In this work, the impacts of Chlorpyrifos (CPF) on sperm oxidative stress markers and sperm motility were investigated in vitro. CPF concentrations were 0 μg/L (control), 5 μg/L, 10 μg/L and 15 μg/L. Lipid peroxidation [malondialdehyde (MDA)], nonenzymatic antioxidants [glutathione (GSH)] and enzymatic [superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and catalase (CAT)] activities in sperm cells were examined for the determination of oxidative stress status. Our findings showed that motility and survival of sperm cells significantly decreased with exposure to Chlorpyrifos. Biochemical assays revealed that CAT activity and levels of MDA and, GSH increased in spermatozoa based on CPF concentration while activity of GSH-Px and SOD decreased. Consequently, spermatozoa were highly sensitive to CPF exposure. It can be deduced that CPF has the potential to disrupt sperm quality and to cause to oxidative stress in sperm cells of S. coruhensis.

Validation of the CellRox Deep Red® fluorescent probe to oxidative stress assessment in equine spermatozoa

Considering the importance of ROS influence on sperm functionality and some limitations in sperm oxidative stress assessment methods, a field to studies of new techniques are still open. In this sense, the aim of this study is to validate the ROS detection technique through the CellRox Deep Red Reagent® probe in stallion sperm. Four stallions were used and the analyses were conducted on four replicates of semen samples from each of stallion (n = 16). The results of the polynomial regression presented a quadratic effect, high determination coefficient value (R2 = 0.88) and high significant P value (P < 0.0001). The CellRox Deep Red® fluorescent probe is able to detect reactive oxygen species in equine sperm, indicating accurately the occurrence of oxidative stress in stallion semen.

Peroxiredoxins prevent oxidative stress during human sperm capacitation.

Do peroxiredoxins (PRDXs) control reactive oxygen species (ROS) levels during human sperm capacitation? PRDXs are necessary to control the levels of ROS generated during capacitation allowing spermatozoa to achieve fertilizing ability. Sperm capacitation is an oxidative event that requires low and controlled amounts of ROS to trigger phosphorylation events. PRDXs are antioxidant enzymes that not only act as scavengers but also control ROS action in somatic cells. Spermatozoa from infertile men have lower levels of PRDXs (particularly of PRDX6), which are thiol-oxidized and therefore inactive. Semen samples were obtained from a cohort of 20 healthy nonsmoker volunteers aged 22-30 years old over a period of 1 year. Sperm from healthy donors was capacitated with fetal cord serum ultrafiltrate (FCSu) in the absence or presence of thiostrepton (TSP), inhibitor of 2-Cys PRDXs or 1-Hexadecyl-3-(trifluoroethyl)-sn-glycero-2-phosphomethanol lithium (MJ33), inhibitor of calcium independent-phospholipase A2 (Ca2+-iPLA2) activity of PRDX6, added at different times of incubation. Capacitation was also induced by the dibutyryl cAMP+3-isobuty1-1-methylxanthine system. Sperm viability and motility were determined by the hypo-osmotic swelling test and computer-assisted semen analysis system, respectively. Capacitation was determined by the ability of spermatozoa to undergo the acrosome reaction triggered by lysophosphatidylcholine. Percentages of acrosome reaction were obtained using the FITC-conjugated Pisum sativum agglutinin assay. Phosphorylation of tyrosine residues and of protein kinase A (PKA) substrates were determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis immunoblotting with specific antibodies. Actin polymerization was determined by phalloidin labeling. TSP and MJ33 prevented sperm capacitation and its associated actin polymerization in spermatozoa incubated with 10% FCSu (capacitation inducer) compared to non-capacitated controls (P < 0.05) without altering sperm viability. PKA substrates and tyrosine phosphorylations were prevented in FCSu-treated spermatozoa in a differential fashion depending on the type and the time of addition of the inhibitor used compared to non-capacitated controls (P < 0.05). TSP and MJ33 promoted an increase of lipid peroxidation in spermatozoa (P < 0.01) and these levels were higher in those spermatozoa incubated with the inhibitors and FCSu compared to those capacitated spermatozoa incubated without the inhibitors (P < 0.0001). Inhibition of 2-Cys PRDXs by TSP generated an oxidative stress in spermatozoa, affecting their viability compared to controls (P < 0.05). This oxidative stress was prevented by nuclephile D-penicillamine (PEN). MJ33 also promoted an increase of lipid peroxidation and impaired sperm viability compared to non-treated controls (P < 0.05) but its effect was not circumvented by PEN, suggesting that not only peroxidase but also Ca2+-iPLA2 activity of PRDX6 are necessary to guarantee viability in human spermatozoa. Not applicable. We focused on the global effect of PRDXs inhibitors on human sperm capacitation and in two of its associated phosphorylation events. Thus, other phosphorylation events and mechanisms necessary for capacitation may also be affected. PRDXs are the major antioxidant system in ejaculated spermatozoa and are necessary to allow spermatozoon to achieve fertilizing ability (capacitation and acrosome reaction). This research was supported by Canadian Institutes of Health Research (MOP 133661) and the Fonds de Recherché en Santé Quebec (FRSQS #22151) to C.O. The authors have nothing to disclose.

Caspase 3 Activity and Lipoperoxidative Status in Raw Semen Predict the Outcome of Cryopreservation of Stallion Spermatozoa.

Stallion-to-stallion variability in the quality of cryopreserved ejaculates postthaw affects the commercial acceptability of frozen semen and thus is a major constraint for the equine industry. In recent years, the molecular mechanisms associated with sperm damage during cryopreservation have become better understood. Identification of the freezability of the ejaculates before the freezing process is initiated will have a major impact on the equine industry. We studied three markers of oxidative stress in sperm, including 8-iso-PGF2alpha, 8-OH guanosine, and 4-hydroxynonenal (4-HNE); the presence of active caspase 3; and their changes after sperm cryopreservation. Although 4-HNE levels increased after cryopreservation (from 7% to 33%, P < 0.001), 8OH-guanosine and 8-ISO-PGF2alpha levels decreased after cryopreservation (from 130 to 35 arbitrary fluorescence units, P < 0.01, and from 1280 to 1233, P < 0.01, respectively). Postthaw sperm quality was classified as poor, average, or good using the 25th and 75th percentiles of all assays of sperm quality studied (motility, velocity, membrane functionality, and thiol content) as thresholds. Using these values, a sperm postthaw quality index was proposed. Receiver operating characteristic curves and the Youden J statistic were used to investigate the value of the measured parameters in fresh sperm as predictors of potential freezability. Using these techniques, we identified markers of bad freezers (percentages of caspase 3-positive dead sperm [area under the curve (AUC) = 0.820, P < 0.05] and percentages of caspase 3- and 4-HNE-positive sperm [AUC = 0.872, P < 0.05]) and good freezers (percentages of caspase 3-negative live sperm [AUC = 0.815, P < 0.05], percentages of live sperm with high thiol content [AUC = 0.907, P < 0.01], and percentages of 8-ISO-PGF2alpha-positive sperm [AUC = 0.900, P < 0.01]. Moreover, we described for the first time the presence of 8-ISO-PGF2alpha in stallion spermatozoa and revealed the importance of considering different markers of oxidative stress.

Effect of brain-derived neurotrophic factor on sperm function, oxidative stress and membrane integrity in human.

Oxidative stress has negative impacts on the clinical outcomes of assisted reproduction techniques. The brain-derived neurotrophic factor (BDNF) promotes the viability of nerve cells and is known to decrease oxidative stress and apoptosis in different cells. The aim of this study was to evaluate the effect of BDNF treatment on human sperm functions that are known to be essential for fertilisation. Our findings showed that treatment of human spermatozoa with 0.133 nM BDNF significantly increased the percentages of both total (P=0.001) and progressive (P<0.01) motile sperm cells compared to those observed in the nontreated (control) group. We also showed that the mean fluorescence intensity of DCFH-DA, as an indicator of intracellular reactive oxygen species, was significantly lower (P<0.05) in spermatozoa treated with BDNF compared to the control group. Treatment of spermatozoa with BDNF significantly decreased the percentages of both dead (P=0.001) and apoptotic-like sperm cells (P<0.05) compared to the control group. On the other hand, BDNF treatment significantly increased the percentage of viable sperm cells compared to the control (P=0.001). In conclusion, BDNF has protective effects against oxidative stress in spermatozoa and could improve sperm functions that are essential for sperm-egg fusion and subsequent fertilisation.

Lower dietary n-6 : n-3 ratio and high-dose vitamin E supplementation improve sperm morphology and oxidative stress in boars.

A 2×2 factorial experiment (10 boars per treatment) was conducted for 16 weeks to evaluate the effects of the dietary n-6:n-3 ratio (14:1 vs 6:1) and vitamin E (200 vs 400mg kg-1) on boar sperm morphology and oxidative stress. Sperm mitochondrial membrane potential (MMP), reactive oxygen species (ROS), DNA damage (8-hydroxydeoxyguanosine; 8-OHdG), seminal lipoperoxidation (malondialdehyde; MDA) and antioxidant capacity in the serum, spermatozoa and seminal plasma were assessed as indicators of oxidative stress. Sperm production was similar among groups but increased (P<0.05) throughout the 16 weeks of the study. Although sperm α-tocopherol content, ROS and seminal MDA did not differ between the two dietary n-6:n-3 ratio treatments, enhanced antioxidant enzyme activity and MMP, but decreased 8-OHdG, were found in spermatozoa from boars consuming the 6:1 diet. The diet with the 6:1 ratio positively affected sperm morphology at Weeks 12 and 16 (P<0.05). The α-tocopherol content and antioxidant capacity increased in boars with increasing levels of vitamin E supplementation. Compared with low-dose vitamin E, high-dose vitamin E supplementation improved sperm morphology. Overall, the results indicate that an n-6:n-3 ratio of 6:1 and 400 mg/kg vitamin E have beneficial effects on sperm morphology by improving antioxidative stress.

A novel antioxidant formulation designed to treat male infertility associated with oxidative stress: promising preclinical evidence from animal models.

Does a novel antioxidant formulation designed to restore redox balance within the male reproductive tract, reduce sperm DNA damage and increase pregnancy rates in mouse models of sperm oxidative stress? Oral administration of a novel antioxidant formulation significantly reduced sperm DNA damage in glutathione peroxidase 5 (GPX5), knockout mice and restored pregnancy rates to near-normal levels in mice subjected to scrotal heat stress. Animal and human studies have documented the adverse effect of sperm DNA damage on fertilization rates, embryo quality, miscarriage rates and the transfer of de novo mutations to offspring. Semen samples of infertile men are known to be deficient in several key antioxidants relative to their fertile counterparts. Antioxidants alone or in combination have demonstrated limited efficacy against sperm oxidative stress and DNA damage in numerous human clinical trials, however these studies have not been definitive and an optimum combination has remained elusive. The efficacy of the antioxidant formulation was evaluated in two well-established mouse models of oxidative stress, scrotal heating and Gpx5 knockout (KO) mice, (n = 12 per experimental group), by two independent laboratories. Mice were provided the antioxidant product in their drinking water for 2-8 weeks and compared with control groups for sperm DNA damage and pregnancy rates. In the Gpx5 KO model, oxidative DNA damage was monitored in spermatozoa by immunocytochemical detection of 8-hydroxy-2'-deoxyguanosine (8OHdG). In the scrotal heat stress model, male fertility was tested by partnering with three females for 5 days. The percentage of pregnant females, number of vaginal plugs, resorptions per litter, and litter size were recorded. Using immunocytochemical detection of 8OHdG as a biomarker of DNA oxidation, analysis of control mice revealed that around 30% of the sperm population was positively stained. This level increased to about 60% in transgenic mice deficient in the antioxidant enzyme, GPX5. Our results indicate that an 8 week pretreatment of Gpx5 KO mice with the antioxidant formulation provided complete protection of sperm DNA against oxidative damage. In mouse models of scrotal heat stress, only 35% (19/54) of female mice became pregnant resulting in 169 fetuses with 18% fetal resorption (30/169). This is in contrast to the antioxidant pretreated group where 74% (42/57) of female mice became pregnant, resulting in 427 fetuses with 9% fetal resorption (38/427). In both animal models the protection provided by the novel antioxidant was statistically significant (P < 0.01 for the reduction of 8OHdG in the spermatozoa of Gpx5 KO mice and P < 0.05 for increase in fertility in the scrotal heat stress model). It was not possible to determine the exact level of antioxidant consumption for each mouse during the treatment period. Recent clinical studies confirm moderate to severe sperm DNA damage in about 60% of all men visiting IVF centers and in about 80% of men diagnosed with idiopathic male infertility. Our results, if confirmed in humans, will impact clinical fertility practice because they support the concept of using an efficacious antioxidant supplementation as a preconception therapy, in order to optimize fertilization rates, help to maintain a healthy pregnancy and limit the mutational load carried by children. The study was funded by the Clermont Université and the University of Madrid. P.G. is the Managing Director of CellOxess LLC, which has a commercial interest in the detection and resolution of oxidative stress. A.M. and A.P. are employees of CellOxess, LLC. J.R.D., A.G.-A. and R.J.A. are honorary members of the CellOxess advisory board.

Sperm Oxidative Stress Is Detrimental to Embryo Development: A Dose-Dependent Study Model and a New and More Sensitive Oxidative Status Evaluation.

Our study aimed to assess the impact of sperm oxidative stress on embryo development by means of a dose-dependent model. In experiment 1, straws from five bulls were subjected to incubation with increasing H2O2 doses (0, 12.5, 25, and 50 μM). Motility parameters were evaluated by Computed Assisted System Analysis (CASA). Experiment 2 was designed to study a high (50 μM) and low dose (12.5 μM) of H2O2 compared to a control (0 μM). Samples were incubated and further used for in vitro fertilization. Analyses of motility (CASA), oxidative status (CellROX green and 2'-7' dichlorofluorescein diacetate), mitochondrial potential (JC-1), chromatin integrity (AO), and sperm capacitation status (chlortetracycline) were performed. Embryos were evaluated based on fast cleavage (30 h.p.i.), cleavage (D = 3), development (D = 5), and blastocyst rates (D = 8). We observed a dose-dependent deleterious effect of H2O2 on motility and increase on the percentages of positive cells for CellROX green, capacitated sperm, and AO. A decrease on cleavage and blastocyst rates was observed as H2O2 increased. Also, we detected a blockage on embryo development. We concluded that sperm when exposed to oxidative environment presents impaired motility traits, prooxidative status, and premature capacitation; such alterations resulting in embryo development fail.

Causes and consequences of oxidative stress in spermatozoa.

Spermatozoa are highly vulnerable to oxidative attack because they lack significant antioxidant protection due to the limited volume and restricted distribution of cytoplasmic space in which to house an appropriate armoury of defensive enzymes. In particular, sperm membrane lipids are susceptible to oxidative stress because they abound in significant amounts of polyunsaturated fatty acids. Susceptibility to oxidative attack is further exacerbated by the fact that these cells actively generate reactive oxygen species (ROS) in order to drive the increase in tyrosine phosphorylation associated with sperm capacitation. However, this positive role for ROS is reversed when spermatozoa are stressed. Under these conditions, they default to an intrinsic apoptotic pathway characterised by mitochondrial ROS generation, loss of mitochondrial membrane potential, caspase activation, phosphatidylserine exposure and oxidative DNA damage. In responding to oxidative stress, spermatozoa only possess the first enzyme in the base excision repair pathway, 8-oxoguanine DNA glycosylase. This enzyme catalyses the formation of abasic sites, thereby destabilising the DNA backbone and generating strand breaks. Because oxidative damage to sperm DNA is associated with both miscarriage and developmental abnormalities in the offspring, strategies for the amelioration of such stress, including the development of effective antioxidant formulations, are becoming increasingly urgent.

Fertilix, a novel antioxidant formulation designed to treat male infertility emanating from sperm oxidative DNA damage: promising preclinical evidence from mouse models

Does Fertilix, a novel antioxidant formulation designed to alleviate oxidative stress in the male reproductive tract, reduce Sperm DNA Damage (SDD) and increase pregnancy rates in mouse models of Sperm Oxidative Stress (SOS)?

Antioxidative effects of magnetized extender containing bovine serum albumin on sperm oxidative stress during long-term liquid preservation of boar semen

Magnetized water is defined as water that has passed through a magnet and shows increased permeability into cells and electron-donating characteristics. These attributes can protect against membrane damage and remove reactive oxygen species (ROS) in mammalian cells. We explored the effects of improved magnetized semen extenders containing bovine serum albumin (BSA) as antioxidants on apoptosis in boar sperm. Ejaculated semen was diluted in magnetized extender (0G and 6000G) with or without BSA (0G + BSA and 6000G + BSA), and sperm were analyzed based on viability, acrosome reaction, and H2O2 level of live sperm using flow cytometry. Sperm were then preserved for 11 days at 18 °C. We found that viability was significantly higher in 6000G + BSA than under the other treatments (P < 0.05). The acrosome reaction was significantly lower in the 6000G + BSA group compared with the other treatments (P < 0.05). Live sperm with high intracellular H2O2 level were significantly lower in the 6000G + BSA group than under other treatments (P < 0.05). Based on our results, magnetized extenders have antioxidative effects on the liquid preservation of boar sperm.

An Efficient Technique to Detect Sperm Reactive Oxygen Species: The CellRox Deep Red® Fluorescent Probe

Heat stress and testicular traumas increase sperm Reactive Oxygen Species (ROS) resulting in Oxidative Stress (OS) and injury of sperm. The fluorescent probe CellROX Deep Red® is not already used to detect ROS in spermatozoa. On this way, this study aims to evaluate its efficiency in detecting ROS in ram sperm. For that, it was used ejaculates from rams performing the in vitro induction of sperm OS in T0, T50 and T100. T0 was semen sample that was not submitted to OS induction, T50 was 50% induced to OS induction and T100 was entire submitted to OS induction. The production of ROS was detected by CellROX Deep Red®. Data polynomial regression analysis was performed and the result of determination coefficient was 0.728. Besides, sixteen rams were submitted to scrotal insulation during 72 hours performing the in vivo induction of OS sperm. Analyses of sperm motility, morphology, DNA fragmentation and ROS (detected by CellROX Deep Red®) were done two times before the scrotal insulation and two times afterwards. Data analysis of variance was performed from the periods (before x after) and it is observed that after scrotal insulation the total and progressive motility decrease (p<0.05) and total and major sperm defects and sperm DNA fragmentation increase (p<0.05). Moreover, after insulation, it was observed increase (p<0.05) in sperm showing moderate and intense OS. Thus, it is possible to conclude that CellROX® fluorescent probe is able to detect ROS production in ram sperm by in vitro and in vivo induction being a new technique to detect sperm OS.

Maternal obesity and overnutrition increase oxidative stress in male rat offspring reproductive system and decrease fertility.

Increasing evidence exists that maternal obesity (MO) and overnutrition during pregnancy and lactation have long-lasting consequences for progeny metabolism, cardiovascular and endocrine function. Data on effects of MO on offspring reproduction are limited. We hypothesized that MO during pregnancy and lactation in founder F(0) rat mothers would increase testicular and sperm oxidative stress (OS) and adversely impact male fertility in their F(1) offspring. We induced pre-pregnancy MO by feeding F(0) females a high-fat diet from weaning through pregnancy and lactation. After weaning, all F(1) rats ate control (C) diet. We determined serum testosterone, malondialdehyde (MDA), reactive oxygen species (ROS) and superoxide dismutase (SOD) and glutathione peroxidase (GPx) activity in F(1) testes and sperm at postnatal days (PNDs) 110, 450 and 650. At PNDs 450 and 650, MO offspring had lower luteinizing hormone while testosterone levels were lower at all ages. Testicular MDA and ROS concentrations and SOD and GPx activity were higher in MO F(1) at all ages. Nitrotyrosine immunostaining was higher at all ages in MO F(1) testes than C F(1). At PNDs 450 and 650, MO F(1) spermatozoa showed higher MDA concentrations and lower SOD and GPx activity with reduced sperm concentration, viability and motility, and more sperm abnormalities. Fertility rate was not affected at PND 110 but was lower in MO F(1) at PNDs 450 and 650. We conclude that MO during pregnancy and lactation increases F(1) testicular and sperm OS leading to premature aging of reproductive capacity.

Engaging practicing gynecologists in the management of infertile men.

In the modern era, contemporary management of male infertility has undergone groundbreaking changes with the introduction of new concepts, advanced testing, and therapeutic interventions. As practicing gynecologists are often the first physicians who encounter an infertile couple, it is essential that these clinicians are continuously updated about the new pearls and pitfalls of male infertility management. Semen analysis is commonly ordered by gynecologists. In 2010, the WHO released new cutoff reference values for the semen parameters adopting novel methodology, which has incited much debate. Reference values have been lowered in comparison with previous standards, with a direct clinical implication in decision-making strategies. Specialized sperm-function tests, such as sperm oxidative stress and sperm chromatin integrity assessments, became clinically available, thus offering an opportunity to better understand sperm dysfunctions concealed during routine semen analysis. Furthermore, the initial counseling of azoospermic men by an andrologically well educated gynecologist may alleviate the misconception and distress surrounding the false belief of sterility, and will clarify the available options of percutaneous and microsurgical sperm-retrieval techniques and assisted conception outcome. Regarding varicocele, which is commonly seen in infertile males, it is now clear that the best treatment option for infertile men with clinical varicocele is the microsurgical vein ligation. Natural conception is significantly improved after varicocelectomy, and recent data suggest that such treatment optimizes reproductive outcome of couples undergoing ICSI or micro-TESE sperm retrieval. Lastly, new therapeutic interventions, including oral antioxidant therapy and lifestyle modifications, have gained increasing attention, as they aid in alleviating male infertility.

Protective effects of coenzyme Q10 and aspartic acid on oxidative stress and DNA damage in subjects affected by idiopathic asthenozoospermia.

Male infertility is negatively influenced by reactive oxygen species and by other oxidant radicals [1]. In this regard, the relevant role of Coenzyme Q10 (CoQ10) in contributing to the total antioxidant buffer capacity of semen has been extensively investigated [1]. Two distinct studies conducted by our group, showed that in idiopathic asthenozoospermic patients undergoing CoQ10 therapy a significant increase in spermatozoa motility was observed [1, 2], suggesting its potential therapeutic role. On the other hand, Aspartic acid (D-Asp) supplementation was found to increase the concentration and motility of spermatozoa in oligo-asthenozoospermic and asthenozoospermic subjects [3]. Some evidence suggests a protective effect against oxidative stress for this molecule [4]. Only a few interventional studies have assessed oxidative stress in sperm of idiopathic infertile men treated with CoQ10 and D-Asp [5–9]. However, they just examined some of the markers of oxidative stress, focusing, in the great majority of cases, only on subjects who were administered CoQ10 [6–9]. Given these premises, we conducted a study in infertile idiopathic subjects who underwent co-administration of CoQ10 and D-Asp in order to evaluate some previously untested parameters of sperm oxidative stress, like superoxide dismutase (SOD) activity, nitric oxide (NO) and peroxynitrite levels, and analyze the effects of these substances on sperm DNA damage markers.

Abnormal early cleavage events predict early embryo demise: sperm oxidative stress and early abnormal cleavage.

Human embryos resulting from abnormal early cleavage can result in aneuploidy and failure to develop normally to the blastocyst stage. The nature of paternal influence on early embryo development has not been directly demonstrated although many studies have suggested effects from spermatozoal chromatin packaging, DNA damage, centriolar and mitotic spindle integrity, and plasma membrane integrity. The goal of this study was to determine whether early developmental events were affected by oxidative damage to the fertilizing sperm. Survival analysis was used to compare patterns of blastocyst formation based on P2 duration. Kaplan-Meier survival curves demonstrate that relatively few embryos with short (<1 hr) P2 times reached blastocysts, and the two curves diverged beginning on day 4, with nearly all of the embryos with longer P2 times reaching blastocysts by day 6 (p < .01). We determined that duration of the 2nd to 3rd mitoses were sensitive periods in the presence of spermatozoal oxidative stress. Embryos that displayed either too long or too short cytokineses demonstrated an increased failure to reach blastocyst stage and therefore survive for further development. Although paternal-derived gene expression occurs later in development, this study suggests a specific role in early mitosis that is highly influenced by paternal factors.

Effect of varicocelectomy on sperm functional characteristics and DNA methylation.

In individuals with varicocele, DNA is damaged due to high level of oxidative stress, and varicocelectomy can overcome this effect. Damaged DNA is less liable to DNA methylation, and antioxidant therapy appears to have the potential to reduce sperm oxidative stress and DNA damage and thereby maintain DNA methylation, while effect of varicocelectomy on DNA methylation patterns has remained unclear. In the light of these considerations, we aimed to examine the effect of varicocelectomy on sperm DNA methylation and functional characteristics. Fifty-two men with left-sided varicocele (grade II &III) were included. Sperm parameters, DNA fragmentation, protamine deficiency, oxidative stress and global DNA methylation were evaluated before and 3months after surgery. Our data show that sperm concentration, percentages of spermatozoon with abnormal morphology, DNA fragmentation, protamine deficiency and oxidative stress significantly improved after surgery. Percentage of sperm motility, global DNA methylation and intensity of DNA methylation also improved after surgery, although the differences were not significant when compared with before surgery. Categorisation of individuals to subgroups revealed that improvement of DNA methylation appears to take place in oligozoospermic individuals, which are more severely affected by state of varicocele. However, this is a preliminary study, and further studies are required to solidify this conclusion.

Chlorophytum borivilianum (Safed Musli) root extract prevents impairment in characteristics and elevation of oxidative stress in sperm of streptozotocin-induced adult male diabetic Wistar rats.

BackgroundWe hypothesized that C. borivilianum root, known to improve male reproductive performance, prevents impairment in characteristics, morphology and elevation of oxidative stress in sperm of diabetics. We therefore investigated the effect of aqueous root extract of C. borivilianum on these parameters in diabetic rat model.MethodsC. borivilianum root aqueous extract (250 and 500 mg/kg/day) or glibenclamide (600 μg/kg/day) were administered to streptozotocin (STZ)-induced diabetic male rats for 28 consecutive days. At the end of treatment, animals were sacrificed and sperm were collected. Sperm count and percentages of forward motility, viability, hypoosmotic swelling (HOS) tail-coiled and abnormal sperm were evaluated. Sperm lipid peroxidation product (LPO), hydrogen peroxide (H2O2) and nitric oxide (NO) levels, total antioxidant capacity (TAC), activity levels of endogenous antioxidant enzymes (superoxide dismutase-SOD, catalase-CAT and glutathione peroxidase-GPx), epididymal sperm density, serum fasting blood glucose (FBG) and HbA1c levels were measured. The expression of sperm caspase-3 was assessed. Meanwhile, in-vitro free radical scavenging activity of C. borivilianum root extract was determined and the root extract was analyzed for the presence of bioactive compounds by FTIR spectroscopy.ResultsC. borivilianum root aqueous extract prevents the decrease in sperm count, percentages of forward motility, viability, HOS and the increase in abnormal sperm percentage and caspase-3 level in diabetic rats. Sperm LPO, H2O2 and NO levels, FBG and HbA1c were lower while TAC, SOD, CAT, GPx and epididymal sperm density were higher in diabetic rats receiving C. borivilianum root extract treatment. C. borivilianum root exhibited strong in-vitro free radical scavenging activity which may be due to the phenolic compound.ConclusionsC. borivilianum root extract prevents impairment in sperm characteristics and morphology via preventing elevation of oxidative stress, apoptosis and free radicals levels of the sperm in diabetes. These effects may be achieved through maintaining sperm antioxidant level which could be related to free radical scavenging activity of the root extract by phenolic compounds. These effects could also be due to ability of the extract to maintain near normal serum FBG and HBA1c levels in diabetes.

Lipid Peroxidation, Sperm DNA Fragmentation Total Antioxidant Capacity and Semen Quality in Male Infertility

Infertility is a major clinical problem, affecting people medically and psychosocially. Male factor plays a significant role in about 50% of infertile couples. Recent reports indicate that increasing male infertility could be due to genomic abnormalities. The etiology of sperm DNA damage is multi-factorial but compromised due to nuclear defects, protamine deficiency and oxidative stress. The present study was aimed to evaluate sperm DNA integrity and oxidative stress in infertile men. The study is prospective, comprises 96 infertile patients and 30 fertile controls. Sperm DNA integrity was assessed by flowcytometry. MDA and TAC were evaluated spectrophotometrically. The percentage of DNA Fragmentation Index and MDA were found to be significantly increased while TAC was significantly decreased in infertile men as compared to control. DFI and MDA were negatively correlated with TAC levels. Present study indicates significant increases in seminal MDA and sperm DNA damage in infertile men. Seminal MDA was significantly correlated with sperm DNA damage, TAC and standard sperm parameters. The elevated levels of seminal OS observed in these infertile patients could be responsible for poor sperm quality and sperm DNA fragmentation. Hence evaluation of DFI, MDA and TAC can be used for diagnosis, prognosis of male infertility in addition to routine semen parameters to decide the treatment strategies.