Porcine sperm vitrification II: Spheres method

Owing to current problems in boar sperm cryopreservation, this study proposes to evaluate vitrification in spheres as an alternative cryopreservation procedure, comparing the use or not of permeable cryoprotectants and two warming methods. Extended (n = 3; r = 4) and raw (n = 5; r = 2) porcine spermatozoa were diluted in media, in the absence or presence of either 4% dimethylformamide or 4% glycerol, to a final concentration of 5 × 10⁶ spermatozoa/ml and vitrified using the spheres method. Two warming procedures were evaluated: a rapid method (30 s at 37°C) and an ultrarapid method (7 s at 75°C, followed by 30 s at 37°C). Percentages of total motility (phase contrast), membrane function (hypo‐osmotic swelling test), acrosome integrity (phase contrast), sperm viability (6‐carboxyfluorescein diacetate and propidium iodide stain), chromatin condensation (toluidine blue stain) and chromatin susceptibility to acid denaturation (acridine orange stain) were evaluated in the samples before and after vitrification. Results, analysed using Friedman's test, suggest that rapid warming of raw porcine spermatozoa vitrified without permeable cryoprotectants may preserve DNA condensation and integrity better than the other processing methods studied in this work. Hence, porcine sperm vitrification using spheres could be used to produce embryos with ICSI to further validate this method.     

Conventional slow freezing cryopreserves mouflon spermatozoa better than vitrification

This work examines the effectiveness of a TCG (Tris, citric acid, glucose, 6% egg yolk and 5% glycerol) and a TEST (TES, Tris, glucose, 6% egg yolk and 5% glycerol) sperm extender in the freezing of mouflon spermatozoa at slow cooling rates, using different pre‐freezing equilibration times (2–3 hr). It also examines the tolerance of mouflon spermatozoa to different concentrations of cryoprotectants (5, 10, 20% glycerol; 5%, 10%, 20% dimethyl sulfoxide; 6% polyvinylpyrrolidone) and/or sucrose (100, 300, 500 mm ). The highest quality (p < .01) thawed spermatozoa were obtained when using the TEST extender and an equilibration time of 3 hr. Sperm motility and membrane integrity were strongly reduced when using rapid freezing rates (60–85°C min^?1), independent of the concentration of cryoprotectants. The lowest sucrose concentration (100 mm ) provided the highest (p < .05) percentage of motile spermatozoa and live spermatozoa with an intact acrosome. Vitrified–warmed sperm variables were at their best when the spermatozoa was diluted in TCG–6% egg yolk + 100 mm sucrose and warmed at 60°C. Slow warming at 37°C strongly reduced (p < .05) sperm motility and viability. However, sperm vitrification returned lower fertility, sperm motility and sperm viability values than conventional sperm freezing.     

Cryoprotectant-free vitrification of fish (Oncorhynchus mykiss) spermatozoa: first report

The aims of this investigation were to test a novel technology comprising cryoprotectant‐free vitrification of the spermatozoa of rainbow trout and to study the ability of sucrose and components of seminal plasma to protect these cells from cryo‐injuries. Spermatozoa were isolated and vitrified using three different media: Group 1: standard buffer for fish spermatozoa, Cortland^® medium (CM, control); Group 2: CM + 1% BSA + 40% seminal plasma; and Group 3: CM + 1% BSA + 40% seminal plasma + 0.125 m sucrose. For cooling, 20‐μl suspensions of cells from each group were dropped directly into liquid nitrogen. For warming, the spheres containing the cells were quickly submerged in CM + 1% BSA at 37 °C with gentle agitation. The quality of spermatozoa before and after vitrification was analysed by the evaluation of motility and cytoplasmic membrane integrity with SYBR‐14/propidium iodide staining technique. Motility (86%, 81% and 82% for groups 1, 2 and 3, respectively) (P > 0.1) was not decreased significantly. At the same time, cytoplasmic membrane integrity of spermatozoa of Groups 1, 2 and 3 was changed significantly (30%, 87% and 76% respectively) (P < 0.05). All tested solutions can be used for vitrification of fish spermatozoa with good post‐warming motility. However, cytoplasmic membrane integrity was maximal in Group 2 (CM + 1% BSA + 40% seminal plasma). In conclusion, this is the first report about successful cryoprotectant‐free cryopreservation of fish spermatozoa by direct plunging into liquid nitrogen (vitrification). Vitrification of fish spermatozoa without permeable cryoprotectants is a prospective direction for investigations: these cells can be successfully vitrified with 1% BSA + 40% seminal plasma.     

Vitrification of human ICSI/IVF spermatozoa without cryoprotectants: new capillary technology

The aim of this study was to develop and to test the standardized aseptic technology of permeable cryoprotectant-free vitrification of human spermatozoa in capillaries (for intracytoplasmic sperm injection [ICSI] or in vitro fertilization [IVF]). To test the effect of vitrification on basic sperm parameters, each of 68 swim-up-prepared ejaculates from oligo-astheno-terato-zoospermic patients were aliquoted and distributed into 3 groups: 1) nontreated control, 2) 10 μL of spermatozoa cryopreserved by slow conventional freezing with glycerol-contented medium, and 3) 10 μL of spermatozoa vitrified in 50-μL plastic capillaries in culture medium with 0.25 M sucrose. Spermatozoa motility (1, 24, and 48 hours after warming), plasma membrane integrity, acrosomal integrity, and spontaneous capacitation-like changes were determined after warming. Aseptic cryoprotectant-free vitrification showed a significantly stronger cryoprotective effect compared with conventional freezing. One hour after warming, motility, plasma membrane integrity, and acrosomal integrity were significantly higher than is observed for conventionally frozen spermatozoa (28% vs 18%, 56% vs 22%, and 55% vs 21%, respectively; P < .05), although lower than in fresh spermatozoa (35%, 96%, and 84%, respectively; P < .05). Capacitation-like changes did not differ significantly between vitrified and conventionally frozen samples (8% vs 9%, respectively; P > .1) (2% in fresh spermatozoa). The newly developed technology of aseptic vitrification of human spermatozoa in capillaries can effectively preserve these cells from cryo-injures. Spermatozoa, vitrified by this technology, are free from seminal plasma owing to swim-up preceding vitrification and are free from permeable cryoprotectants. They are ready for further use immediately after warming without any additional treatment. Therefore, the reported technology has a great potential for use in ICSI/IVF.     

Influence of extended incubation time on Human sperm chromatin condensation,sperm DNA strand breaks and their effect on fertilisation rate

The purpose of this study was to determine influence of extended incubation time on sperm chromatin condensation and DNA strand breaks and their effect on fertilisation rate. Forty couples undergoing ICSI therapy were included. Semen was prepared by PureSperm gradient centrifugation and divided into two parts. The first part (G1) was used immediately for ICSI, whereas the second part (G2) was kept in the incubator at 37°C, 5% and 90% Humidity for 5 hr, and thereafter, the capacitated spermatozoa were used for ICSI. The TUNEL test and chromomycin CMA3 were used to evaluate the DNA strand breaks and chromatin condensation respectively. The percentage of condensed chromatin was 73.92 ± 12.70 in the group 1 and 81.13 ± 10.31% in group 2 (p = .001). However, the double‐strand breaks were 11.15 ± 8.67% in G.1 and 16.30 ± 11.12% in G.2. (p = .001). Fertilisation rate in the (Group 1) was 62.45% and 69.17% in (Group 2). There was a positive correlation between condensed chromatin and fertilisation rate (r = 0.846, p = .001) and a negative correlation with DNA double‐strand breaks (r = ?0.802; p = .001). In conclusion, the prolonged sperm incubation (5 hr) leads to a higher chromatin condensation and to a significantly increased number of DNA strands double breaks with no influence on fertilisation rates.     

Vitrified sperm banks: the new aseptic technique for human spermatozoa allows cryopreservation at −86 °C

The vitrification technique is simple, quick, cost‐effective and has showed a significantly stronger cryoprotective effect in contrast to conventional freezing. The method is based on the rapid cooling of the cell by direct immersion in liquid nitrogen (LN ₂), thereby avoiding the formation of ice crystals, due to the lower risk of water thawing, which impairs cell function. The aim of this study was to evaluate the effect of storage at ?86 °C compared to the conventional ?196 °C (under LN ₂) on essential parameters of the functioning of aseptically vitrified human sperm. Sperm motility, integrity of mitochondrial membrane potential and the rate of DNA fragmentation were determined. The comparison of ?86 °C and ?196 °C demonstrated no statistical difference in sperm progressive motility (73% vs. 77%), integrity of mitochondrial membrane potential (71% vs. 74%) or DNA fragmentation (3.1% vs. 2.9%). In conclusion, aseptically vitrified sperm can be preserved at ?86 °C; eliminating the use of LN ₂ simplifies and significantly reduces the costs associated with storage in sperm banks by decreasing the time and space needed for storage, the effort in finding stored samples, and by improving safety for the operator. However, for prolonged storage further studies are needed.     

Sperm telomere length in motile sperm selection techniques: A qFISH approach

Several studies have associated telomere shortening with alterations in reproductive function. The objective of the present study was to determine telomere length (TL) in spermatozoa selected by either density‐gradient centrifugation (DGC) or swim‐up. The analysis of TL was performed using quantitative fluorescent in situ hybridisation (qFISH) using PNA probes in combination with a chromatin decompaction protocol in sperm cells. Results of TL were 24.64 ± 5.00 Kb and 24.95 ± 4.60 Kb before and after DGC, respectively, and 19.59 ± 8.02 Kb and 20.22 ± 5.18 Kb before and after swim‐up respectively. Sperm selected by DGC or swim‐up did not show any significant differences in TL as compared to nonselected sperm (p > .05). Negative correlations between TL and sperm motility (r = ?.308; p = .049) and concentration (r = ?.353; p = .028) were found. Furthermore, exposure of sperm to increasing concentrations of hydrogen peroxide during incubation resulted in a reduction in TL. These data indicate that oxidative stress may be one of the main factors involved in the reduction of TL in sperm. Preliminary clinical results from patients included in this study indicate that TL was shorter in spermatozoa from couples who never achieved a pregnancy compared to couples who did achieve at least one natural pregnancy (p < .05); however, the clinical utility of this biomarker still needs to be confirmed in further studies.     

High temperature is essential for preserved human sperm function during the devitrification process

Sperm vitrification is a cryopreservation method based on high‐speed freezing by direct exposure of cells in liquid nitrogen (N₂L), thereby avoiding the traditional cooling curves of freezing. The objective of this work was to determine the optimal warming temperature for vitrified human spermatozoa in order to maintain their fertilisation potential. Spermatozoa were cryopreserved by direct plunging into N₂L and warmed at different temperatures for 5 and 10 s at 38, 40 and 42 °C. Sperm motility was evaluated by the CASA system and the sperm membrane function by HOST test. It was detected that progressive motility of sperm warmed at 38, 40 and 42 °C was 26.4 ± 8.4%; 56.6 ± 16.3% and 65.4 ± 15%, respectively, with statistically significant differences between the temperatures of 38 and 40 °C and 38 and 42 °C (P < 0.05). The plasma membrane function evaluated by HOST test was better preserved at 42 °C (76.3 ± 2.0%) compared to 40 °C (43 ± 2%) and 38 °C (65.6 ± 1.5%). The temperature in the thawing process can affect the motility and plasma membrane integrity and function. The warming at 42 °C for thawed vitrified sperm is the optimum temperature to preserve the sperm physiological parameters.     

Sperm selection based on motility in polyvinylpyrrolidone is associated with successful pregnancy and embryo development

The aim of this study was to investigate whether spermatozoon motility in polyvinylpyrrolidone (PVP) is associated with better embryo development and pregnancy rates in ICSI cycles. A total of 123 primary ICSI treatment cycles were included in this study. Semen samples were tested for motility before ICSI procedure in PVP. Within 3 min, the presence or absence of motility was recorded. Sperm functions were examined by the aniline blue (AB) chromatin condensation test and the hypoosmotic swelling test, and the chromatin stability was evaluated by inducing its decondensation with sodium dodecyl sulphate and ethylenediaminetetraacetic acid (EDTA). Fertilisation and embryo scoring were evaluated. Fifty (64%) of 78 women conceived in the PVP (+) group; and 12 (26%) of 45 women conceived in the PVP (?) group; the pregnancy rate was significantly higher in the PVP (+) group (P = 0.003). Semen parameters were observed to be similar in both groups. The mean number of total embryos obtained in ICSI procedure and transferred grade 1 embryos were significantly higher in PVP (+) group (P = 0.01 and P = 0.003 respectively). The presence of sperm motility in PVP is associated with increased pregnancy rate, higher percentage of good quality embryos, sperm chromatin condensation and decondensation.     

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.     

Human sperm DNA integrity in normal and abnormal semen samples and its correlation with sperm characteristics

Reports indicate an increase in the incidence of DNA fragmentation in male factor infertility and its role in the outcome of assisted reproductive techniques (ART). However, reports are conflicting between the relationships of sperm DNA integrity with conventional semen parameters. We examined the relationship between conventional sperm parameters and DNA integrity using acridine orange (AO) test. The study included 373 patients and 28 fertile volunteers. DNA normality was compared with semen parameters between the patient and donor populations. Significant correlations were noted between DNA normality and sperm concentration ( r  = 0.18, P  = 0.000), motility ( r =  0.21, P  = 0.0001), rapid motility (0.19, P  = 0.000), normal morphology by World Health Organization ( r =  0.15, P  = 0.019) and head defects ( r =  −0.15, P  = 0.023). A significant difference was noted in AO levels between donors and patients with asthenozoospermia ( P  = 0.002) and oligoasthenozoospermia ( P  = 0.001). A significant difference in DNA integrity was noted in samples having <30% and >30% normal morphology. A wide range of % DNA normality was observed in the patient group. Sperm assessment for DNA status using AO is reliable and shows good correlation with sperm count, motility and morphology. Assessment of sperm DNA status with AO staining may be helpful prior to ART.     

Sperm impairments in adult vesper mice (Calomys laucha) caused by in utero exposure to bisphenol A

This study aimed to evaluate the effects of in utero administration of bisphenol A (BPA) on semen parameters of vesper mice. Sixty female Calomys laucha were divided into six groups and received by gavage during gestation the following substances: Water (negative control), Olive Oil (vehicle control), Diethylstilbestrol (DES – positive control – 6.5 μg kg^?1 bw) and BPA (40, 80 and 200 μg kg^?1 bw). Male offspring were euthanised at 70 days of age, and sperm parameters were analysed. BPA reduced normal sperm morphology (water = 96.1 ± 0.65; BPA200 = 96.8 ± 2.3%), sperm membrane integrity (water = 88.8 ± 1,65; BPA200 = 70.6 ± 4,15%), sperm motility (water = 87.5 ± 1.71; BPA200 = 51.3 ±9.9%) and in vitro penetration rates (water = 55.0 ± 7.14; BPA200 = 7.47 ±2.96%), but it did not affect body weight, anogenital distance, sperm DNA integrity and acrosome integrity. In conclusion, in utero exposure to BPA caused a reduction in sperm parameters of adult C. laucha. Natural mating studies should be conducted to verify the effects of BPA on fertility of the animals.     

How accurate is sperm morphology as an indicator of sperm function?

Sperm morphology has been consistently correlated with fertilisation success or failure. The clinical relevance of the percentage normal spermatozoa has been a widely discussed topic amongst infertility specialists and scientists. This study aimed to evaluate the role of sperm morphology as an indicator of additional sperm functions among 114 andrology referrals. The sperm functions that were investigated included chromatin packaging quality (CMA3 test (n = 109), zona‐induced acrosome reaction (ZIAR test; n = 36), hemizona assay (HZI; n = 36) and progressive motility (n = 47). Chromatin packaging quality had a negative and significant (P = 0.0001, r = ?0.74) correlation with the percentage normal spermatozoa, while progressive motility had a significant and positive correlation (P = 0.0001, 0.59). Accurate sperm morphology scoring as described by the WHO 2010 manual can therefore be used as an indicator of specific sperm functions.     

The micronutrient supplements,zinc sulphate and folic acid,did not ameliorate sperm functional parameters in oligoasthenoteratozoospermic men

We investigated the effects of folic acid and zinc sulphate supplementation on the improvement of sperm function in subfertile oligoasthenoteratozoospermic (OAT) men. Eighty‐three OAT men participated in a 16‐week intervention randomised, double‐blind clinical trial with daily treatment of folic acid (5 mg day^?1) and zinc sulphate (220 mg day^?1), or placebo. Before and after treatment, semen and blood samples were obtained for determining sperm concentration, motility, and morphology, sperm viability, sperm mitochondrial function, sperm chromatin status using toluidine blue, aniline blue, acridine orange and chromomycin A₃ staining; and semen and blood folate, zinc, B₁₂, total antioxidant capacity ( TAC) and malondialdehyde (MDA) concentrations. Sperm concentration (×10⁶ ml^?1) increased in subfertile men receiving the combined treatment of folic acid and zinc sulphate and also in the group receiving only folic acid treatment; however, it was not statistically significant (P = 0.056 and P = 0.05, respectively). Sperm chromatin integrity (%) increased significantly in subfertile men receiving only zinc sulphate treatment (P = 0.048). However, this improvement in sperm quality was not significant after adjusting placebo effect. This study showed that zinc sulphate and folic acid supplementation did not ameliorate sperm quality in infertile men with severely compromised sperm parameters, OAT. Male infertility is a multifactorial disorder, and also nutritional factors play an important role in results of administration of supplementation on sperm parameters. However, these results should be confirmed by multiple studies in larger populations of OAT men.     

Correlation of biochemical constituents of seminal plasma with semen quality in Teddy goat (Capra hircus) bucks

This study was planned to determine the relationship between semen quality parameters and the levels of biochemical constituents of seminal plasma of Teddy (Capra hircus) buck semen. For this purpose, semen ejaculates were collected from five mature healthy Teddy bucks. All the experimental bucks were kept under natural environmental conditions. Semen was collected twice in a week for the duration of 6 weeks by Artificial Vagina (AV) in the breeding season (February‐April). Two successive ejaculates of single buck were pooled at time of collection, and a total of 60 semen samples were processed for semen analysis. Sperm per cent motility, sperm concentration, dead sperm percentage, morphological abnormal spermatozoa, plasma membrane integrity were correlated with biochemical constituents of seminal plasma. The mean per cent motility (89.18% ± 0.37%), sperm concentration (1.86 ± 0.04 × 10⁹/ml), dead sperm percentage (8.08% ± 0.29%), morphological abnormal spermatozoa (6.05% ± 0.29%) and plasma membrane integrity (88.22% ± 0.34%) were recorded. The seminal plasma contained Na⁺ (144.12 ± 1.59 mEq/L), K⁺ (27.38 ± 0.49 mEq/L), Cl^? (65.73 ± 0.45 mEq/L), Ca⁺⁺ (9.34 ± 0.22 mg/dl), P (19.32 ± 0.97 mg/dl), aspartate aminotransferase (AST; 26.48 ± 1.30 IU/L), alanine aminotransferase (ALT; 168.47 ± 5.18 IU/L), lactate dehydrogenase (LDH; 215.98 ± 6.06 IU/L), albumin (1.90 ± 0.10 g/dl), globulins (2.08 ± 0.11 g/dl) and total protein (3.98 ± 0.20 g/dl). The collected data were analysed by applying Pearson's correlation coefficients. Dead sperm percentage had negative correlation with sodium (r = ?.278, p < .05), albumin (r = ?.294, p < .05), globulin (r = ?.266, p < .05) and total protein (r = ?.295, p < .05). Phosphorus was negatively associated with sperm concentration (r = ?.262, p < .05). AST was negatively correlated with plasma membrane integrity (r = ?.292, p < .05). It was concluded that most of the semen quality parameters of Teddy bucks were positively correlated with biochemical constituents, but opposite trends were found in case of dead sperm percentage. The seminal biochemical constituents dynamically interact with each other.     

Antioxidant effect of hydroxytyrosol on human sperm quality during in vitro incubation

The aim of the study was to evaluate the antioxidant potential of hydroxytyrosol (HT) on human sperm quality during incubation in vitro. Semen samples collected from men attending the Laboratory of Histology‐Embryology of Sfax Faculty of Medicine (Tunisia) for infertility investigations were evaluated for initial sperm parameters. Only normal selected ejaculates (n = 15) were centrifuged and incubated further with or without HT (200ug ml^?1) at room temperature for 45 min. After incubation, sperm motility and viability, DNA oxidation and reactive oxygen species (ROS) production were assessed. The results showed that centrifugation significantly influenced sperm motility and viability. The supplementation of HT in incubating media improved (P = 0.01) significantly sperm viability and decreased sperm DNA oxidation (P < 0.001) and ROS levels (P = 0.03) following centrifugation. It can be concluded that supplementation of HT might be helpful to maintain the human spermatozoon after centrifugation.     

A comparative study of two cooling protocols on stallion sperm cryosurvival

There are many protocols for horse sperm cryopreservation, but results are inconsistent; sperm survival after freeze‐thawing is usually poor; in consequence, fertility is low. The objective of this work was to see whether slow cooling before freezing to minus 3 °C instead of +5 °C, the traditional target temperature, could improve horse sperm cryosurvival, capability to carry out capacitation and the acrosome reaction induced by progesterone. Spermatozoa from five stallions were packaged in straws and slowly cooled to +5 °C. Half of the straws were frozen directly and the other half was further cooled to ?3 °C before freezing. Progressive motility, viability, plasma membrane integrity, acrosome integrity and capacitation status were assessed. After thawing, there were no differences between cooling treatments on motility, viability, acrosome integrity and capacitation status; however, there was difference (P < 0.05) regarding plasma membrane integrity. Acrosome integrity decreased as incubation, without or with progesterone (2 μg ml^?1), progressed, but there were no differences between cooling treatments regardless of progesterone. Both capacitated and acrosome‐reacted spermatozoa increased as incubation progressed, but there were no differences between cooling treatments regardless of progesterone. Slow cooling to ?3 °C before freezing did not improve horse sperm cryosurvival or capability to undergo the acrosome reaction.     

Additional deleterious effects of alcohol consumption on sperm parameters and DNA integrity in diabetic mice

The aim of this study was to survey the impact of alcohol consumption on sperm parameters and DNA integrity in experimentally induced diabetic mice. A total of 32 adult male mice were divided into four groups: mice of group 1 served as control fed on basal diet, group 2 received streptozotocin (STZ) (200 mg kg^?1, single dose, intraperitoneal) and basal diet, group 3 received alcohol (10 mg kg^?1, water soluble) and basal diet, and group 4 received STZ and alcohol for 35 days. The cauda epididymidis of each mouse was dissected and placed in 1 ml of pre‐warm Ham's F10 culture medium for 30 min. The swim‐out spermatozoa were analysed for count, motility, morphology and viability. Sperm chromatin quality was evaluated with aniline blue, toluidine blue, acridine orange and chromomycin A3 staining. The results showed that all sperm parameters had significant differences (P < 0.05), also when sperm chromatin was assessed with cytochemical tests. There were significant differences (P < 0.001) between the groups. According to our results, alcohol and diabetes can cause abnormalities in sperm parameters and chromatin quality. In addition, alcohol consumption in diabetic mice can intensify sperm chromatin/DNA damage.     

Role of ADAMTS1 and ADAMTS5 in male infertility

The aim of this study was to investigate the relationship of infertility with metalloproteinases ADAMTS1 and ADAMTS5, which are known to be responsible for the degradation of extracellular matrix (ECM) proteins associated with many diseases. ECM is the noncellular component that provides structural and biochemical support to the surrounding cells required for tissue morphogenesis, differentiation and homoeostasis. Sixty infertile individuals and 10 healthy semen donors were included in this study. The infertile individuals were classified as normozoospermia (NS; n = 20), oligozoospermia (OS; n = 20), azoospermia (AS; n = 20) groups. ADAMTS1 and ADAMTS5 protein levels in semen were analysed by Western blot. ADAMTS1 protein level was 3.0‐, 3.3‐ and 1.6‐fold lower in the OS, AS and NS groups, respectively, than in the control group (P < 0.001). ADAMTS5 protein level was 3.2‐, 2.7‐ and 1.4‐fold lower in the OS, AS and NS groups, respectively, than in the control group (P < 0.001). Sperm count and sperm motility showed a negative correlation with the levels of ADAMTS1 and ADAMTS5 protein expression: r = ?0.477, r = ?0.470; and r = ?0.332, r = ?0.275 respectively (P < 0.001). In conclusion, ADAMTS1 and ADAMTS5 protein expressions in semen are significantly related with sperm production. It is very important to understand molecular function and organisation of ADAMTSs which will be significant in enlightening the process of spermatogenesis in male infertility.     

Effect of etodolac hydrazone,a new compound synthesised from etodolac,on spermatozoon quality,testicular lipid peroxidation,apoptosis and spermatozoon DNA integrity

The aim of this study was to investigate the effect of etodolac hydrazone (EH), a new compound synthesised from etodolac, on spermatozoon quality, testicular lipid peroxidation, apoptosis and spermatozoon DNA integrity in rats. Group 1 (n = 8) received 1 ml dimethyl sulfoxide (DMSO) daily (Control); group 2 (n = 8) was treated with 5 mg kg^?1 day^?1 EH, dissolved in 1 ml DMSO (EH‐5); and group 3 (n = 8) was treated with 10 mg kg^?1 day^?1 EH, dissolved in 1 ml DMSO (EH‐10). All administrations were performed by gavage and maintained for 8 weeks. Both doses of EH administration caused significant decreases in absolute and relative weights of testis, whole epididymis, right cauda epididymis, and spermatozoon motility, spermatozoon count in comparison with the control group. Only 10 mg kg^?1 day^?1 EH administration caused significant decreases in absolute and relative weights of seminal vesicles and serum testosterone level, and significant increases in testicular lipid peroxidation level, and numbers of TUNEL+ apoptotic germ cells and spermatozoa with damaged DNA along with some histopathological damages when compared to the control group. However, body and ventral prostate weight, and testicular antioxidant markers (glutathione, glutathione‐peroxidase and catalase), were unaffected significantly by both doses of EH administration. In conclusion, two different doses of EH, in particular its high dose, damage to testicular spermatogenic cells and spermatozoon DNA and, it decreases spermatozoon motility, count and testosterone level in healthy rats.