Effects of pure human follicle-stimulating hormone (pFSH) on sperm quality correlate with the hypophyseal response to gonadotrophin-releasing hormone (GnRH)*

Summary. In 41 men with idiopathic infertility, the effect of pure follicle-stimulating hormone (pFSH) therapy on semen parameters was evaluated in relation to the results of the gonadotrophin-releasing hormone (GnRH) stimulation test in an open study. The patients showed a mean (± standard error) sperm concentration of 15.84 (±2.87) Mill, ml^?1, 41.02 (±4.19)% of the spermatozoa were motile and 46.16 (±2.36)% normo-morph before treatment. All patients were administered 150 IU pFSH subcutaneously three times weekly for 10 weeks. After therapy with pFSH, no significant differences compared with the pre-treatment sperm characteristics were observed (P>0.05) except for the velocity average path, VAP (P= 0.029). However, the stimulation factors of FSH in the GnRH test showed a significantly negative correlation with the pFSH mediated improvement of sperm concentration (r= ?0.662, P= 0.000002), of percentage of normo-morph spermatozoa (r= ?0.480, P=0.0015) and total count of motile spermatozoa per ejaculate (r=?0.567, P=0.00014). In consequence, the patients were divided into two groups depending on the stimulation factor of hypophyseal FSH secretion, FSH-SF. The cut-off point of the FSH-SF was set at 1.7 because the correlation analyses detected a 1.5–fold improvement of sperm parameters at this hypophyseal response on average. Pure FSH treatment increased the concentration of spermatozoa (P= 0.0007), the total count of motile spermatozoa in the ejaculate (P=0.015) as well as the computer-aided sperm motion parameters VAP (P=0.029) and velocity curve linear, VCL (P=0.049) in patients with FSH-SF<1.7, whereas in the patient group with FSH-SF>1.7 no improvement of semen parameters was found. Insufficient stimulation of hypophyseal FSH secretion may be a prerequisite but not a guarantee for responsiveness to pFSH treatment. The results of the present investigation suggest that FSH stimulation in the GnRH test should be taken into account in idiopathic infertile men before pFSH therapy.     

无精子症患者睾丸内精子存在的评估

目的　检测无精子症患者睾丸内精子存在情况。　方法　睾丸活检病例 5 0例 ,每例均作血内分泌激素检测、睾丸体积测量、睾丸组织学检查及睾丸精子提取 (TESE) ,分析促卵泡生成素(FSH)、睾丸体积和睾丸组织学与睾丸内精子存在的相关性。　结果　血FSH和睾丸体积预测睾丸精子是否存在准确性不强 ,而睾丸组织学结果与TESE一致 (敏感性 96 % ,特异性 10 0 % ,准确性10 0 % )。　结论　血FSH高和睾丸体积小的无精子症患者 ,应行睾丸活检并同时行TESE以明确睾丸内是否有精子。     

Open testicular mapping: A less invasive multiple biopsy approach for testicular sperm extraction

This study proposes a testicular sperm extraction technique that was inspired by testicular fine-needle aspiration. Here, we have described the technique of open testicular mapping (OTEM) and evaluated the successful sperm recovery in 92 patients with nonobstructive azoospermia (NOA). All patients underwent an OTEM biopsy. Patients were divided into two groups; group I included men with spermatozoa recovered and group 0 included men without spermatozoa recovered. Age, follicle-stimulating hormone (FSH) level and testicular volume were compared between the groups. In 50 of 92 men (54%), viable spermatozoa were found after OTEM. No differences were noted in age, FSH level or testicular volume. Using OTEM, it was possible to retrieve spermatozoa in 54% of the NOA men.     

Pregnancy after intracytoplasmic sperm injection following extended sperm preparation and hormone therapy in an azoospermic man with maturation arrest and microlithiasis: a case report and literature review

In the management of azoospermia, a combination of testicular sperm extraction and intracytoplasmic sperm injection (ICSI) is usually the most successful option for fatherhood. However, an outstanding question remains: How can at least a few spermatozoa be obtained from the ejaculate, thus avoiding the need for a surgical procedure? A 36‐year‐old man presented to Assisted Reproduction Unit with his 26‐year‐old wife. The ultrasound assessment revealed bilateral microlithiasis. Two spermograms revealed absolute azoospermia. Levels of follicle‐stimulating hormone (FSH) and luteinising hormone were normal–low. The patient underwent 10 months of treatment with clomiphene citrate. A bilateral testicular sperm extraction failed to retrieve spermatozoa and revealed a maturation arrest at spermatocyte/spermatid stages depending on the tubules. Clomiphene citrate was replaced with recombinant FSH (rFSH). After 9‐month treatment with rFSH, motile spermatozoa from droplets of ejaculate pellet were cryopreserved as a single straw. Ovarian stimulation was provided using classic antagonist protocol, and five mature oocytes were collected. Two consecutive fresh semen samples on the day of ICSI yielded seven motile spermatozoa, and fertilisation was achieved in all five oocytes. On day 3, two embryos were transferred, yielding positive beta‐human chorionic gonadotropin and a healthy delivery of a boy and a girl.     

Predictive factors of a successful testicular biopsy and subsequent clinical pregnancy

Forecast of success with testicular sperm extraction and intracytoplasmic sperm injection (ICSI) remains unknown, as predictive factors have rarely been studied. We evaluated the association among possible predictive factors and a successful biopsy and clinical pregnancy. A consecutive sample of men submitted to a testicular open biopsy in S. João Hospital was used. Patient's age, medical history, testicular volume, spermogram, genetic testing, endocrinologic results, biopsy results and clinical pregnancy information were collected. From the 113 men included, it was possible to retrieve spermatozoa in 79.6% of the cases, which resulted in 58 fertilisations and 22 clinical pregnancies. Retrieving viable spermatozoa on biopsy was associated with the identification of spermatozoa in the spermogram (100.0% versus 74.4%; P = 0.010), diseases causing obstructive infertility (100.0% versus 79.2%; P = 0.036) and no genetic causes detected (82.4% versus 54.5%; P = 0.030). Successful clinical pregnancy was only associated with lower female partner age (31.7 versus 36.0 year; P = 0.001) but not the quality of the spermatozoa or the time until the reproduction cycle. Identification of spermatozoa in the spermogram, diseases causing obstructive infertility and lack of genetic causes for infertility were associated with higher probability of viable spermatozoa retrieval but the female partner age remained the principal determinant of a successful pregnancy.     

Can preoperative gonadotropin and testosterone levels predict the success of varicocelectomy?

We examined the effects of preoperative hormonal values on varicocelectomy success. A total of 136 patients who underwent varicocelectomy for infertility in our clinic were analysed retrospectively. Improvement in semen quality was defined as >50% increase in post-operative total motile sperm count (TMSC) in those with preoperative TMSC >5 million and at least 100% increase in those with <5 million. The patients were divided into two groups as benefiting from the treatment (Group A) and no benefits (Group B). The best cut-off value for follicle-stimulating hormone (FSH) and the luteinising hormone/testosterone ratio (LTR) that can predict varicocelectomy success were 7.01 and 0.016 with an area under the curve of 0.844 and 0.856 respectively. The highest sensitivities and specificities of FSH and LTR were 0.845 and 0.788 and 0.821 and 0.846 respectively. Binary logistic regression analysis showed FSH (odds ratio [OR]: 3.7; p < .001) and LTR (OR: 5.2; p < .001) as independent predictive factors in predicting varicocelectomy success. Our study demonstrated that low FSH (7.01 IU/L) and LTR (<.016) can be a useful preoperative predictive tool to help identify men who benefit most from varicocelectomy in infertile patients with varicocele.     

Selection of normal spermatozoa with a vacuole‐free head (x6300) improves selection of spermatozoa with intact DNA in patients with high sperm DNA fragmentation rates

Intracytoplasmic morphologically selected sperm injection (IMSI, 6300× magnification with Nomarski contrast) of a normal spermatozoon with a vacuole‐free head could improve the embryo's ability to grow to the blastocyst stage and then implant. However, the most relevant indications for IMSI remain to be determined. To evaluate the potential value of IMSI for patients with a high degree of sperm DNA fragmentation (n = 8), different types of spermatozoa were analysed in terms of DNA fragmentation. Motile normal spermatozoa with a vacuole‐free head selected at 6300× magnification had a significantly lower mean DNA fragmentation rate (4.1 ± 1.1%, n = 191) than all other types of spermatozoa: non‐selected spermatozoa (n = 8000; 26.1 ± 1.5% versus 4.1 ± 1.1%; P < 0.005), motile spermatozoa (n = 444; 20.8 ± 2.7% versus 4.1 ± 1.1%; P < 0.001) and motile, normal spermatozoa selected at 200× magnification (n = 370; 18.7 ± 2.7% versus 4.1 ± 1.1%; P < 0.001) and then motile, morphometrically normal spermatozoa with anterior vacuoles (n = 368; 15.9 ± 2.9% versus 4.1 ± 1.1%; P < 0.05) or posterior vacuoles (n = 402; 22.5 ± 3.6% versus 4.1 ± 1.1%; P < 0.001) selected at 6300× magnification. For patients with high sperm DNA fragmentation rates, selection of normal spermatozoa with a vacuole‐free head (6300×) yields the greatest likelihood of obtaining spermatozoa with non‐fragmented DNA.     

COVID-19 pneumonia causes lower testosterone levels

To evaluate the testicular damage caused by COVID-19, we prospectively evaluated 44 patients who applied to the COVID-19 outpatient clinic between March 2020 and July 2020. Patients' ages, COVID-19 PCR results, presence of pneumonia, total testosterone, luteinising hormone (LH) and follicle-stimulating hormone (FSH) values were recorded. It was evaluated whether there were significant differences between people who were positive for COVID-19 and those who were not. Any differences between those who had COVID-19 pneumonia and those who did not were also recorded. There was no difference between the FSH, LH and testosterone values of the COVID-19 PCR positive and negative patients (p = 0.80, vp = 0.62, p = 0.56 respectively). However when LH values were separated as low, normal and high, LH values were statistically significantly higher in the COVID-19 PCR positive group (p = 0.04). Thoracic computed tomography was performed in 42 patients. Testosterone levels were significantly lower in patients with COVID-19 pneumonia (p = 0.01). When FSH, LH and testosterone values were separated as low, normal and high, there was no difference in FSH and LH values (p = 1, p = 0.2). Testosterone levels were found significantly lower in patients with COVID-19 pneumonia (p < .001). Testosterone levels seem to decrease during acute COVID-19 infection, especially in the patient group with viral pneumonia.     

Impact of cell phone use on men's semen parameters

The objective of the present retrospective study was to report our experience concerning the effects of cell phone usage on semen parameters. We examined 2110 men attending our infertility clinic from 1993 to October 2007. Semen analysis was performed in all patients. Serum free testosterone (T), follicle stimulating hormone (FSH), luteinising hormone (LH) and prolactin (PRL) were collected from all patients. The information on cell phone use of the patients was recorded and the subjects were divided into two groups according to their cell phone use: group A: cell phone use (n = 991); group B: no use (n = 1119). Significant difference was observed in sperm morphology between the two groups. In the patients of group A, 68.0% of the spermatozoa featured a pathological morphology compared to only 58.1% in the subjects of group B. Patients with cell phone usage showed significantly higher T and lower LH levels than those who did not use cell phone. No significant difference between the two groups was observed regarding FSH and PRL values. Our results showed that cell phone use negatively affects sperm quality in men. Further studies with a careful design are needed to determine the effect of cell phone use on male fertility.     

Impact of using a fast‐freezing technique and different thawing protocols on viability and fertility of frozen equine spermatozoa

The effects of freezing technique and thawing protocol on thawed semen viability and fertility were studied. Ejaculates from 5 stallions (n = 25) were frozen by conventional or a fast‐freezing technique. Frozen semen was thawed by two thawing protocols (37 °C 30 s^?1 or 75 °C 7 s^?1). Thawed semen was evaluated by progressive motility, vigour, morphology and plasma membrane integrity. Mares (n = 25) were inseminated with 300 (n = 11) or 150 (n = 14) million spermatozoa. A greater (P < 0.05) vigour and progressively motile spermatozoa were detected, respectively, at thawing and after 20 min post‐thawing in the fast‐freezing technique than in the conventional one. Plasma membrane integrity was also greater (P < 0.05) in semen frozen with the fast‐freezing technique. Semen viability was not affected by thawing protocol. Pregnancy rate using the fast‐freezing technique was 76% (19/25), and did not differ (P > 0.05) between insemination doses. We concluded that the 150 million progressively motile spermatozoa per dose using a deep‐horn insemination maximises the use of equine semen. The fast‐freezing technique, as compared to the conventional one, efficiently preserves the viability and fertilising capacity of spermatozoa, indicating a new method to improve the fertility of frozen equine semen.     

In vitro equine embryo production using air‐dried spermatozoa,with different activation protocols and culture systems

The aim of this work was to evaluate the use of air‐dried spermatozoa for in vitro production of equine embryos and verify if sperm extract activation and in vivo culture improve in vitro embryo production. Cooled spermatozoa (control) and air‐dried spermatozoa stored for 2, 14 or 28 days were used for ICSI sperm extract, or ionomycin was used for oocyte activation, and embryos were in vitro or in vivo (in mare′s oviduct) cultured for 7 days. With in vitro culture, cleavage rate was higher when activating with sperm extract (P < 0.05). No differences in embryo development were seen between the two activation treatments nor between storage periods (P > 0.05). Blastocysts were obtained with cooled spermatozoa, and morulae were achieved using in vivo culture with 28‐day storage spermatozoa and ionomycin‐activated oocytes. When in vivo culture was performed, sperm DNA fragmentation was assessed using the sperm chromatin dispersion test and did not show statistical correlation with cleavage nor embryo recovery rates. In conclusion, equine embryos can be produced using air‐dried spermatozoa stored for several weeks. Sperm extract activation increased cleavage rates but did not improve embryo development. In vivo culture allowed intrauterine stage embryos to be achieved.     

Age as only predictive factor for successful sperm recovery in patients with Klinefelter’s syndrome

The study was performed to determine factors affecting successful sperm retrieval by testicular sperm extraction in patients with nonmosaic Klinefelter’s syndrome (KS). From May 2001 to February 2007, 27 azoospermic patients were diagnosed as having nonmosaic KS. All patients underwent sperm testicular extraction. Patient’s age, testicular volume, serum follicle‐stimulating hormone (FSH) and inhibin B were assessed as predictive factors for successful sperm recovery. Of the 27 Klinefelter’s patients examined, eight (29.6%) had successful sperm recovery. The comparisons of serum FSH, inhibin B and testicular volume between patients with and without successful sperm retrieval did not show any statistical significance. The patients with successful sperm recovery were significantly younger (28.6 ± 3.11 years) than those with failed attempts (33.9 ± 4.5 years, P = 0.002). The rate of positive sperm retrieval was significantly higher in patients younger than 32 years compared with patients older than 32 years (P = 0.01, chi‐squared test). The study showed that clinical parameters such as FSH, inhibin B and testicular volume do not have predictive value for sperm recovery in patients with KS. The mean age of our patients with successful sperm recovery was significantly lower than that of men with unsuccessful results. Testicular sperm extraction or testicular sperm aspiration should be performed before the critical age of 32 years.     

The efficacy of combined l-carnitine and l-acetyl carnitine in men with idiopathic oligoasthenoteratozoospermia: A systematic review and meta-analysis

The purpose of our analysis is to identify the effect of l -carnitine (LC) and l -acetyl carnitine (LAC) on the semen parameters of men with idiopathic oligoasthenoteratozoospermia (iOAT). We performed a comprehensive search to ascertain all the trials about LC and LAC in the treatment of iOAT and compared the results, including percentage of total sperm motility, sperm concentration, percentage of forward sperm motility, semen volume, percentage of atypical forms, total motile spermatozoa, forward motile spermatozoa and the number of pregnancies between the two groups that treated with LC + LAC or placebo respectively. Seven randomised controlled trials (RCTs) involving 693 patients were included in our analysis. We found that patients who treated with LC and LAC had significantly increased the percentage of forward sperm motility (MD 6.98; 95% CI 1.06–12.90; p = .02), total motile spermatozoa (MD 16.45; 95% CI 8.10–24.79; p = .0001), forward motile spermatozoa (MD 13.01; 95% CI 11.08–14.94; p < .00001) and the number of pregnancies (OR 3.76; 95% CI 1.66–8.50; p = .002). However, no significant differences were found in other semen indicators between the two groups. LC and LAC can significantly increase part of the semen parameters. The combination therapy of LC and LAC is effective in the men with iOAT.     

Body mass index has no impact on sperm quality but on reproductive hormones levels

The influence of overweight and obesity on sperm quality and reproductive hormone levels is under discussion. The aim of the present retrospective study was to evaluate the influence of body mass index (BMI) on sperm quality and reproductive hormones. We analysed semen samples and serum levels of FSH, LH, T and PRL of a total of 2110 men attending our andrology unit from 1994 to 2010 due to infertility work‐up. Patients were stratified according to their BMI in four groups. Main outcome measures were sperm motility, morphology and concentration. Serum levels of FSH, LH, T and PRL were evaluated as well. No statistically significant difference was found for sperm quality and BMI between patients categorised according to the four BMI levels. T (P < 0.001) and LH (P = 0.006) significantly differed between the four groups. In multivariable analysis, BMI did not have significantly independent influence on all assessed sperm quality parameters, whereas BMI significantly influenced hormone values for LH (P = 0.001), T (P = <0.001) and PRL (P = 0.044). We therefore conclude that BMI has no significant impact on sperm quality parameters. However, serum levels of LH, T and PRL were significantly influenced by BMI.     

The number of spermatozoa collected with testicular sperm extraction is a novel predictor of intracytoplasmic sperm injection outcome in non-obstructive azoospermic patients

The purpose of this study was to determine the relationships between monitors of spermatogenesis and predictors of the intracytoplasmic sperm injection (ICSI) outcome in patients with non-obstructive azoospermia (NOA) undergoing testicular sperm extraction (TESE). Seventy-nine patients with NOA (mean age: 43.6±5.2 years), each of whom yielded (97 000±3040) spermatozoa with conventional TESE, were considered in our analysis. Their partners (mean age: 35.8±5.1 years) underwent a total of 184 ICSI cycles; 632 oocytes were collected, 221 oocytes were injected, 141 oocytes were fertilized, 121 embryos were obtained, 110 embryos were transferred, 14 clinical pregnancies were achieved and only one miscarriage occurred. Multivariate regression analysis indicated relationships between the percentage of fertilized oocytes, transferred embryos and clinical pregnancies with the following variable values: female partner''s age, number of spermatozoa collected, testicular volume, male partner''s levels of follicle stimulating hormone (FSH), number of oocytes collected, number of oocytes injected and number of ICSI cycles. A significant inverse relationship was found between female partner''s age or male partner''s FSH levels and biochemical pregnancies. A significant direct relationship emerged between the number of ICSI cycles and the percentage of oocytes fertilized, embryos transferred and biochemical pregnancies, and between the number of spermatozoa collected per testicular biopsy and biochemical pregnancies. The number of spermatozoa was positively linked to the number of clinical pregnancies, independent of the number of ICSI cycles and the number of oocytes collected/injected. The number of spermatozoa collected, FSH level and testicular volume are monitors of spermatogenesis linked to ICSI success.     

Recombinant human FSH reduces sperm DNA fragmentation in men with idiopathic oligoasthenoteratozoospermia

A prospective randomized controlled study was designed to evaluate the effects of recombinant human follicle-stimulating hormone (rFSH) treatment on sperm DNA fragmentation in men with idiopathic oligoasthenoteratozoospermia (iOAT). One hundred twenty-nine men with sperm count less than 10 × 10(6) spermatozoa/mL and forward motility <25% were included; normal serum levels of FSH, luteinizing hormone (LH), and testosterone, and no other causes of infertility were enrolled. The patients were randomized into 2 groups: 65 men were treated on alternate days for 90 days with injections of 150 IU rFSH, and 64 subjects received nonantioxidant vitamin supplements. Main outcome measures were serum levels of FSH, LH, testosterone, and inhibin B and DNA fragmentation index (DFI) at baseline and after 90 days. No significant differences were observed between the 2 groups with regard to sperm parameters and hormone values. The DFI was similar between the 2 groups at the time of the enrollment but reduced significantly (P < .05) after rFSH therapy in study group, whereas no significant variation occurred in the control group. In the subgroup of patients with high basal DFI values (>15%), rFSH treatment significantly increased DFI (P < .01), whereas no significant variation occurred after 90 days of vitamin supplements. We conclude that rFSH administration improves sperm DNA integrity in iOAT men with increased DFI values. The degree of sperm DFI might be useful to identify those iOAT patients in which rFSH treatment can be advantageous.     

Effect of different types of textile fabric on spermatogenesis: an experimental study

Summary The effect of different types of textile fabric on spermatogenesis was studied. Twenty-four dogs were divided into two equal groups, one of which wore cotton underpants and the other polyester ones. Seven dogs wearing nothing were used as controls. The underwear was fashioned to fit loosely in the scrotal area so as to avoid its insulating effect. It was worn continuously for 24 months during which the semen character, testicular temperature, hormones (serum testosterone, follicle stimulating hormone, luteinizing hormone, prolactin) and testicular biopsy were examined. The garment was then removed, and the same investigations repeated through another 12 months. The results were analysed statistically. In the polyester group the testicular temperature showed insignificant changes during the period when the pants were worn (P>0.05). By the end of the 24 months there was a significant decrease in sperm count and motile sperms, with an increase in abnormal forms (P<0.001); the testicular biopsy showed degenerative changes. After garment removal the semen character improved gradually to normal in 10 dogs; two remained oligozoospermic. There were insignificant changes (P>0.05) in hormones during the study. In contrast, the cotton and control groups showed insignificant changes (P>0.05) in all the study. The polyester pants thus had a deleterious effect on spermatogenesis in the dogs which was, however, reversible in the majority of cases. The cause of this effect is unknown, but it may be assumed that the electrostatic potentials generated by the polyester fabric play a role in it.     

Clinical and genetic analysis in males with 46,XX disorders of sex development: A reproductive centre experience of 144 cases

To explore the clinical features and assisted reproductive technology (ART) outcomes of 46,XX disorders of sex development (DSD) males, 144 males with 46,XX DSD were recruited in this retrospective study. The baseline information, clinical characteristics and ART outcomes of the participants were collected and analysed. The mean age was 29.06 ± 4.50 years. The mean volumes (95% CI) of left and right testicles were 2.16 (1.82–2.49) ml and 2.16 (1.83–2.49) ml, respectively. Cryptorchidism and/or hypospadias appeared in 19 patients (13.19%). Elevated levels of follicle‐stimulating hormone (FSH) were found in 136 patients (95.10%) and increased luteinising hormone (LH) values were detected in 125 patients (92.59%). Eighty subjects (62.99%) had low testosterone values. Among 86 patients with status of sex‐determining region Y (SRY)—gene and azoospermia factor (AZF) region available, fifteen (17.44%) patients were SRY‐negative and AZF region was absent in every patient without exception. Additionally, fertility achieved in 87 patients through ART using donor spermatozoa. In conclusion, hypergonadotropic hypogonadism appeared as the main presentation of 46,XX DSD males regardless of the SRY status. The available fertility option proved to achieve live birth was limited to ART using donor spermatozoa.     

Anti‐oxidative Status and Semen Quality during Cooled Storage in Stallions

Activity of the anti‐oxidative enzymes glutathione peroxidase (GSH‐Px), superoxide dismutase (SOD) and catalase (CAT), content of thiobarbituric acid reactive substances (TBARS) and SH‐groups were determined in native stallion semen (n = 8 stallions). Semen was then diluted in Kenney extender, EquiPro^® extender either with or without addition of N‐acetyl cysteine or phosphate‐buffered saline (PBS) and stored for 72 h at 5°C. Correlations between initial activity of enzymes and development of semen motility and membrane integrity were calculated. Activities of GSH‐Px, SOD and CAT immediately after semen collections were 10.0 ± 0.6 picokatals, 0.40 ± 0.03 SOD units and 0.70 ± 0.05 nanokatals/10⁶ spermatozoa respectively. TBARS content was 0.06 ± 0.01 nmol and SH‐group content 1.7 ± 0.5 mmol/10⁶ spermatozoa. The loss of motile spermatozoa during storage did not differ between extenders. N‐acetyl cysteine had no effect on semen motility and membrane integrity. The loss in membrane‐intact spermatozoa was highest (P < 0.05) in semen diluted in PBS. Motility and membrane integrity after addition of extender were positively correlated with GSH‐Px and CAT, indicating that anti‐oxidative mechanisms contribute to the initial high percentage of motile and membrane‐intact spermatozoa. However, in these samples the decrease in semen quality was most pronounced. No correlations existed between initial activity of anti‐oxidative enzymes, peroxidation products and semen quality during storage. This indicates that once extender has been added, peroxidative damage to sperm membranes is not the predominant cause of losses in semen quality.     

Canine sperm vitrification with sucrose: effect on sperm function

The ability of sucrose to protect spermatozoa against mitochondrial damage, artificial acrosome reaction and DNA fragmentation during ultra‐rapid cryopreservation in canine sperm was investigated. Swim‐up selected spermatozoa of second‐fraction semen were vitrified with different concentrations of sucrose (0.1, 0.25 and 0.4 m ) in proportion 1 : 1 v/v with HTF–BSA 1%. From each group, 30‐μl suspensions of cells were dropped directly into liquid nitrogen and stored for at least 24 h. Cells were thawed by submerging the spheres in HTF with 1% BSA at 37 °C. The number of progressively motile spermatozoa was significantly higher in the sucrose 0.25 m + HTF–BSA 1% (42.5 ± 2.3%, P < 0.01) than in HTF only (1.66 ± 0.3%). The same combination of sucrose 0.25 m + HTF–BSA 1% (42.7 ± 1.5%) had a stronger cryoprotective effect on the integrity of mitochondrial membrane potential (P < 0.05) and decreased the DNA fragmentation (2.8 ± 0.5%) as compared with HTF only (1.93 ± 0.6% and 5.6 ± 0.6% respectively). With respect to acrosome‐reacted spermatozoa, no significant difference was found between the groups investigated (P > 0.05). It is concluded that sucrose, a nonpermeable cryoprotectant, can effectively preserve important physiological parameters such as mitochondrial membrane potential and DNA integrity during ultra‐rapid cryopreservation.