Protective role of superoxide dismutase in human sperm motility: superoxide dismutase activity and lipid peroxide in human seminal plasma and spermatozoa.

The levels of superoxide dismutase (SOD), a highly specific scavenging enzyme for superoxide anion radicals (O2-), and lipid peroxide produced by oxygen free radicals were measured in human seminal plasma and spermatozoa. Seminal plasma contained 366.8 +/- 20.9 U/ml (mean +/- SE) of SOD activity. SOD activity in human spermatozoa showed a significant correlation to the number of motile spermatozoa, while the activity in seminal plasma did not relate to the sperm concentration or motility. The lipid peroxide concentration in seminal plasma was 6.22 +/- 0.46 nmol/ml and had no significant relationship to sperm concentration or motility. The malondialdehyde (MDA) concentration in spermatozoa was significantly related to the number of immotile spermatozoa. A decrease in the motility of spermatozoa incubated in medium without seminal plasma was observed after 120 min, while the MDA concentration of the spermatozoa increased. Addition of exogenous SOD (400 U/ml) to the sperm suspension significantly decreased this loss of motility and the increase of the MDA concentration. These data suggest a significant role for SOD in sperm motility. It seems that lipid peroxidation of human spermatozoa may cause loss of motility and that SOD may inhibit this lipid peroxidation. These results suggest that SOD may have a possible clinical application in the use of spermatozoa for in-vitro fertilization (IVF) or artificial insemination.     

Glutathione in spermatozoa and seminal plasma of infertile men

Glutathione has a central role in the defence against oxidative damage; however, the data on glutathione concentrations in the semen of infertile men are limited. To expand this knowledge the glutathione content of the ejaculates and blood plasmas of 77 infertile men and 11 controls were analysed. The concentrations of total glutathione were assessed in spermatozoa, seminal plasma and blood plasma using a coupled spectrophotometric assay. In the spermatozoa of patients with oligozoospermia the glutathione concentrations (2.57+/-0.96 nmol/10[8] spermatozoa; mean +/- SD) were significantly lower than in the controls (3.49+/-0.87 nmol/10[8] spermatozoa; P +/- 0.03). The glutathione content of spermatozoa from patients with normozoospermia showed large variations (3.04+/-1.37 nmol/10[8] spermatozoa). An association between the intracellular glutathione content and the ability to penetrate bovine cervical mucus was observed (r = 0.2, P = 0.04). The intracellular glutathione concentrations correlated with the glutathione levels in seminal plasma (r = 0.46, P < 0.0001). There was no correlation between glutathione concentrations in blood plasma and those in seminal plasma or in spermatozoa. The glutathione concentrations in seminal plasmas did not differ between the different groups, however, correlated with the serum follicle stimulating hormone concentrations (r = 0.53, P = 0.005). This study demonstrated that intracellular glutathione levels of spermatozoa are decreased in certain populations of infertile men.      

Pre-freezing sperm preparation does not impair thawed spermatozoa binding to the zona pellucida.

The present study was conducted to assess the fertilizing potential of frozen-thawed spermatozoa, which were cryopreserved after separation on a Percoll gradient, or washed out of seminal plasma. For this purpose, binding to the zona pellucida and other characteristics of the treated sperm cells were compared with those of cryopreserved spermatozoa from the same original sample which were not manipulated before freezing. Semen specimens were obtained from 80 candidates for sperm donation. Percoll-treated sperm samples compared with the sibling, unprocessed controls had significantly higher values of sperm motility characteristics and per cent of cells with normal morphology after freezing and thawing. Sperm binding ability to the zona pellucida was not statistically different (109 +/- 8.1% and 94 +/- 6.7% in unprocessed and Percoll-treated samples respectively). Sperm specimens processed by washing had significantly higher values for motility characteristics than untreated sibling samples, but no differences were found between the treated and untreated samples for morphology and binding to the zona pellucida (hemizona index of 75 +/- 7.0% and 76 +/- 6.7% in unprocessed and washed samples respectively). These findings suggest that, judged by the binding assay, the aforementioned pre-freezing separation processes have no adverse effect upon the fertilizing potential of the thawed sperm cells. These procedures make it possible to optimize the progressive motile sperm cell concentration of the frozen specimen, which facilitates the storage of samples with good quality, even when the features of the original semen are sub-optimal.     

Lipid dynamics in the plasma membrane of fresh and cryopreserved human spermatozoa.

Preserving the integrity of the plasma membrane of spermatozoa is crucial for retention of their fertilizing capacity, especially after stressful procedures such as freezing and storage. In this investigation we have measured lipid diffusion in different regions of the plasma membrane of fresh and cryopreserved human spermatozoa using a sensitive, high resolution fluorescence photobleaching technique (FRAP) with 5-(N-octadecanyl)aminofluorescein as reporter probe. Results show that diffusion was significantly faster on the plasma membrane overlying the acrosome and decreased progressively in the postacrosome, midpiece and principal piece. The midpiece plasma contains a higher proportion of immobile lipids than other regions. In cryopreserved spermatozoa, lipid diffusion in the plasma membrane was significantly reduced on the acrosome, postacrosome and midpiece relative to fresh spermatozoa. Diffusion, however, could be restored to normal levels by washing spermatozoa in a medium containing 0.4% polyvinylpyrrolidine but not in medium alone or in medium containing 0.4% albumin. These results suggest that (i) lipid dynamics in the plasma membrane of human spermatozoa varies significantly between surface regions; (ii) in-plane diffusion is adversely affected by cryopreservation; and (iii) washing frozen spermatozoa in 0.4% polyvinylpyrrolidine restores membrane lipid fluidity to normal levels. The latter finding has important implications for improving the fertility of human spermatozoa following cryopreservation.     

Membrane fluidity predicts the outcome of cryopreservation of human spermatozoa

Semen cryopreservation is an important procedure in the treatment of human infertility. However, the ability of spermatozoa to survive freeze/thaw processes varies between patients. Cryopreservation-induced stress may result in membrane injury with consequent loss of sperm motility and viability. We investigated the relationship between the physico-chemical state of the human sperm membranes and their tolerance to cryopreservation. Conventional characteristics of 20 semen samples were analysed before and after cryopreservation as well as their membrane fluidity assessed by measuring the fluorescence polarization anisotropy, which is inversely proportional to the fluidity. Correlation between fluidity and post-thaw recoveries of motile and viable spermatozoa were examined. Results showed that membrane anisotropy markedly varies between patients. In cryopreserved spermatozoa, anisotropy values were significantly higher than in fresh spermatozoa. Furthermore, recovery of motile and viable spermatozoa after freeze/thaw was strongly correlated with anisotropy of fresh spermatozoa (P < 0.05). The higher the membrane fluidity was before freezing, the better was the response of spermatozoa to cryopreservation. The results indicate that the freeze/thaw process results in a rigidifying effect on the sperm membrane and suggest that sperm adaptability to freeze/thaw-induced stress could be dependent on their initial membrane fluidity. The latter finding has practical implications for predicting the response of spermatozoa following freezing and thawing and for improving the recovery of viable spermatozoa.     

Cryopreservation of spermatozoa: a 1996 review

Both the ability to freeze human spermatozoa and the possibilityof pregnancy following intrauterine insemination have existedfor >40 years. There have been a number of improvements duringthat time concerning the methods of freezing and thawing humanspermatozoa. Initially, the use of the cryoprotective propertiesof glycerol allowed a major improvement; subsequently, changeswere mainly empirical. It was a long time before specific cryobiologicalstudies were undertaken. However, the necessity for these becameapparent with the partial recovery or sometimes loss of motilityafter freezing either subfertile semen before chemotherapy orradiotherapy, or spermatozoa collected from non-physiologicalsituations (epididymal or testicular spermatozoa). The maintrends in improvement have defined end-points other than thepercentage of motility recovery or the assessment of ultrastructuraldamage. More sensitive criteria of the objective assessmentof motility, energy status, damage to the plasma membrane orto subcellular elements, chromatin stability and chromosomaldamage have been proposed as complementary end-points to betterassess sperm cryopreservation. A different approach was relatedto the biochemical environment and physical conditions imposedon spermatozoa during the freezing and thawing process. Biochemicalchanges were assessed following different combinations of variousextenders which attempted either to better preserve some parameteror to avoid the tendency towards drastic increase in osmoticpressure. Analysis of physical conditions was linked to therate of cooling, freezing ad warming, and was based on cryobiologicalstudies. Finally, even though such improvements are not negligible,many questions remained unanswered. The extensive use of frozenspermatozoa during assisted reproductive techniques, togetherwith the development of assisted fertilization using surgicallycollected spermatozoa, creates the need for additional studiesto improve the cryopreservation of human spermatozoa. Keywords: cryopreservation/review/spermatozoa     

圆头精子的冷冻保存

目的:探讨圆头精子的冷冻保存效果及其精子功能改变。方法:6例圆头精子症患者各提供1-3份精液标本,应用不含卵黄冷冻保护剂和二步降温方法冷冻保存圆头精子症标本,检测冷冻精子活动率、头-尾膜完整率、存活时间和顶体蛋白酶活性。结果:圆头精子标本冷冻保存后,精子活动率显著减少,膜完整率显著下降,头膜损伤-尾膜完整的精子率显著升高(n=13,P＜0.01)。体外存活时间缩短。精子冷冻前后无顶体蛋白酶活性。不含卵黄冷冻保护剂与含卵黄冷冻保护剂,以及两步冷冻与多步冷冻的效果比较未见显著差异(n=7,P＞0.05)。结论:圆头精子可经历冷冻保存后存活,但冷冻复苏率低。冷冻-解冻过程显著降低精子功能。不含卵黄冷冻保护剂和两步降温可应用于冷冻保存圆头精子症标本。     

Glutathione and free sulphydryl content of seminal plasma in healthy medical students during and after exam stress

BACKGROUND: It has been reported that there is a relationship between stress and infertility. The mechanisms of stress-related semen quality alterations have not been fully elucidated. In the present study, we investigated the effect of examination stress on seminal glutathione and free sulphydryl content and sperm quality. METHODS: Semen samples were collected from 34 healthy volunteers who were students of medical school in the fourth semester just before (stress period) and 3 months after (non-stress period) their final examinations. Their psychological examination stress was measured by the State Trait Anxiety Inventory (STAI) questionnaire. After standard semen analysis, semen samples were centrifuged at 10 000g for 15 min. Glutathione and free sulphydryl concentration of seminal plasma were measured. RESULTS: During the period of examination stress, the glutathione and free sulphydryl content of seminal plasma and the motility index of spermatozoa were significantly lower, whereas the percentage of morphologically abnormal spermatozoa was higher, than during the non-stress period (P < 0.001, for all). An association between seminal plasma glutathione and motility index was observed at both periods (P < 0.05 and P < 0.01, respectively). CONCLUSIONS: This study demonstrated that glutathione and free sulphydryl levels in seminal plasma decreased in subjects undergoing examination stress. Furthermore, poor sperm quality may be due to loss of glutathione and free sulphydryl content of seminal plasma.     

Detection of fractalkine in human seminal plasma and its role in infertile patients

BACKGROUND: Fractalkine is a relatively newly discovered CX(3)C chemokine, which is a chemoattractant for T cells, monocytes and natural killer cells. Several reports have demonstrated the association between chemokine levels in seminal plasma and semen quality. The fractalkine levels in ejaculates from normal donors and infertile male patients with or without asthenozoospermia, were examined and correlated with sperm motility and morphology. METHODS AND RESULTS: Western blot analysis showed fractalkine protein to be present in the seminal plasma. Fractalkine titres in the seminal plasma of infertile men with asthenozoospermia (0.64 +/- 0.04 microg/ml; n = 58) were lower than those in patients without asthenozoospermia (0.94 +/- 0.10 microg/ml; n = 22, P < 0.01) and fertile donors (1.04 +/- 0.07 microg/ml; n = 10, P < 0.001). There was no significant difference between fractalkine levels in patients with and without leukospermia. No significant correlation was found between fractalkine and interleukin-8 levels in seminal plasma. Sperm motility was positively correlated (R(2) = 0.14, P < 0.001) with fractalkine concentration. The existence of CX(3)CR-positive leukocytes in semen was confirmed using specific primers for CX(3)CR. CONCLUSIONS: These results suggest that fractalkine is a chemokine associated with sperm motility and the migration of CX(3)CR-positive leukocytes into semen.     

Andrology: Effect of freezing method, thawing temperature and post-thaw dilution/washing on motility (CASA) and morphology characteristics of high-quality human sperm

Sixteen semen samples, 12 donor and four patient samples ofhigh initial quality, were processed to compare the effect oftwo freezing methods, two thawing temperatures and the effectof dilution and washing on sperm motility and morphology characteristics.Sperm samples were divided in two equal parts and frozen eitherby fast vapour freezing or by slow computer-controlled freezing.For each freezing method, half of the straws were thawed atroom temperature (22°C), the other half were thawed at 37°C.From each freeze—thawing treatment, one straw was evaluatedimmediately post-thawing; another straw was washed to removethe cryoprotectant solution. In this way, each semen samplewas subjected to eight freeze—thawing treatments. No effectof the freezing method and thawing temperature was observedon motility characteristics evaluated by computer-assisted semenanalysis, nor on light-microscopical morphology parameters.Post-thaw dilution and washing, however, exerted a deleteriouseffect on sperm motility, by reducing percentage motility by50% compared to unwashed thawed specimens. Linearity and percentageof morphologically normal spermatozoa were obviously impaired,while percentage of abnormal tails and beat cross frequencyincreased significantly. In general, freeze-thawing was mostsuccessful when rapid vapour freezing was followed by 37°Cthawing, and when slower computer-controlled freezing was combinedwith 22°C thawing, causing significant interactions betweenthe freezing method and the thawing temperature. For semen samplesof high initial quality, vapour and computer-controlled freezingwere equally effective in terms of recovery of morphologicallynormal, motile spermatozoa.     

Detection of human immunodeficiency virus-1 RNA and DNA by extractive and in situ PCR in unprocessed semen and seminal fractions isolated by semen-washing procedure

BACKGROUND: To determine the presence of human immunodeficiency virus-1 (HIV-1) viral RNA/DNA in whole semen, in properly isolated seminal fractions and in spermatozoa after swim-up, by extractive nested PCR and to compare the detection of HIV DNA by in situ PCR (IS-PCR) with the results of nested PCR. METHODS: We tested HIV-1 RNA and DNA by nested PCR in semen and in seminal fractions from 55 patients. Non-spermatic cells and spermatozoa pellet fractions from 10 HIV-1-positive and five HIV-1-negative men were tested for proviral DNA by IS-PCR. RESULTS: All samples of spermatozoa recovered after sperm washing were free of HIV RNA. HIV RNA tested positive in seven (13%) seminal plasma samples and only in two (4.2%) whole semen of these same samples. Of the seven seminal plasma samples testing positive for HIV RNA, four men had elevated blood viral load and three an undetectable viraemia. HIV DNA by IS-PCR turned positive in three of five samples in semen of HIV-noninfected men. CONCLUSION: HIV RNA/DNA detection in the semen of HIV-infected men proves the efficacy of sperm washing with swim-up of spermatozoa. It is recommended that nested PCR be conducted on purified seminal compartments. IS-PCR is inadequate for detecting HIV in semen.     

Light and electron microscopic analysis of human testicular spermatozoa and spermatids from frozen and thawed testicular biopsies.

The morphological changes caused by freezing and thawing human testicular spermatozoa have been assessed here. Retrieval of testicular biopsies was carried out on six patients with obstructive azoospermia preparatory to intracytoplasmic sperm injection (ICSI). Light microscope analysis was carried out on testicular cells and ultrastructural analysis was carried out on spermatozoa and different spermatid stages before and after the freezing procedure. Upon examination under light microscopy, all germ cells presented increased vacuolization in their cytoplasm and shrinkage or swelling of the nuclei and cytoplasmic membranes. These altered structures were accentuated in the spermatocyte I cell which often presented disrupted membranes. The ultrastructural findings under transmission electron microscopy demonstrated that after freezing and thawing the major types of cryoinjury were the swelling and rupture of inner and outer acrosomal and plasma membranes. The acrosome material often appeared as dispersed material or as condensed spots or was even lost. Such damage was observed mainly at the spermatozoa and late spermatid stages. We conclude that the freezing and thawing of testicular biopsies causes similar morphological damage to testicular spermatozoa and frozen-thawed ejaculated spermatozoa. It is still unclear whether these changes in testicular spermatozoa after freezing and thawing may compromise its use in the ICSI procedure.     

HE1/NPC2 status in human reproductive tract and ejaculated spermatozoa: consequence of vasectomy

We have previously demonstrated that the amount of HE1/NPC2 mRNA and protein expressed in the human epididymis is decreased under vasectomy. In this study, western blot analyses showed that many vasovasostomized men are characterized by high HE1/NPC2 levels in spermatozoa when compared with fertile donors. HE1/NPC2 association with sperm from vasovasostomized men was not related to low motility per se as spermatozoa from asthenospermic men have HE1/NPC2 levels similar to those in normal fertile semen samples. Spermatozoa from vasovasostomized men with high amount of HE1/NPC2 are characterized by higher concentration of cholesterol and more lipid raft domains. HE1/NPC2 is secreted in different glycoforms by different tissues of human male reproductive tract. These forms are due to variation in N-glycosylation, and only the deglycosylated form is associated with spermatozoa from some vasovasostomized men. Compared with normal men, seminal plasma of vasectomized men is characterized by a major decrease in immunodetectable HE1/NPC2 without change in the glycosylation pattern. Following surgical vasectomy reversal, seminal plasma HE1/NPC2 was found in similar amounts to the ones characterizing normal men. Considering the potential role of HE1/NPC2 in cholesterol transport during sperm maturation, unusual high levels of this protein associated with spermatozoa of vasovasostomized men may reflect epididymal sequelae occurring when the vas deferens is obstructed.     

Andrology: Prevention of osmotic injury to human spermatozoa during addition and removal of glycerol

Use of a cryoprotective agent is indispensable to prevent injuryto human spermatozoa during the cryopreservation process. However,addition of cryoprotective agents to spermatozoa before coolingand their removal after warming may create severe osmotic stressfor the cells, resulting in injury. The objective of this studywas to test the hypothesis that the degree (or magnitude) ofhuman sperm volume excursion can be used as an independent indicatorto evaluate and predict possible osmotic injury to spermatozoaduring the addition and removal of cryoprotective agents. Glycerolwas used as a model cryoprotective agent in the present study.To test this hypothesis, first the tolerance limits of spermatozoato swelling in hypoosmotic solutions (iso-osmotic medium dilutedwith water) and to shrinkage in hyperosmotic solutions (iso-osmoticmedium with sucrose) were determined. Sperm plasma membraneintegrity was measured by fluorescent staining, and sperm motilitywas assessed by computer-assisted semen analysis before, duringand after the anisosmotic exposure. The results indicate firstlythat motility was much more sensitive to anisosmotic conditionsthan membrane integrity, and secondly that motility was substantiallymore sensitive to hypotonic than to hypertonic conditions. Basedon the experimental data, osmotic injury as a function of spermvolume excursion (swelling or shrinking) was determined. Thesecond step, using these sperm volume excursion limits and previouslymeasured glycerol and water permeability coefficients of humanspermatozoa, was to predict, by computer simulation, the cellosmotic injury caused by different procedures for the additionand removal of glycerol. The predicted sperm injury was confirmedby experiment. Based on this study, an analytical methodologyhas been developed for predicting optimal protocols to reduceosmotic injury associated with the addition and removal of hypertonicconcentrations of glycerol in human spermatozoa.     

Successful fertilization and pregnancy outcome in in-vitro fertilization using cryopreserved/thawed spermatozoa from patients with malignant diseases.

Cryopreservation of spermatozoa before treatment is the only proven effective method available to circumvent the sterilizing effect of therapy in some patients with malignant diseases. Because of impaired sperm quality after freezing and thawing in-vitro fertilization/embryo transfer (IVF/ET) was indicated in 10 patients (12 cycles) during 1986-1990. The patient's mean age was 33.4 +/- 1.6 years. The following diagnoses were made: seminoma (1), testicular carcinoma (3), leiomyosarcoma of the prostate (1), Wegener's granulomatosis (1), non-Hodgkin's (1) and Hodgkin's lymphoma (3). When motile spermatozoa could be recovered after thawing, the total fraction of motile spermatozoa after swim-up separation ranged from 0.2 to 4.2 x 10(6) spermatozoa/ml (eight patients, nine cycles). In all these cases, insemination was performed with multiple oocytes per dish. Fertilization was achieved when swim-up recovered a mean of 1.8 +/- 0.5 x 10(6) spermatozoa/ml and when insemination was performed with at least a calculated concentration of motile spermatozoa of 1 x 10(5) spermatozoa/oocyte. The fertilization rate of preovulatory oocytes was 60%. Four patients achieved a pregnancy: two of them delivered a single healthy baby, one delivered triplet healthy babies and one had a preclinical abortion. In two patients (three cycles), no motile spermatozoa were recovered after thawing, and micromanipulation of oocytes for assisted fertilization was performed. Although fertilized oocytes were transferred, those couples did not achieve a pregnancy. Patients with lymphopathies had the best results, whilst those with testicular neoplasms had the poorest outcome, thus suggesting a poor gametogenic function in the non-affected testis. These results give hope to some patients with malignant diseases to maintain their reproductive capacity through sperm banking and IVF/ET.     

Andrology: Density adjustment of software settings minimizes bias in automated sperm motility estimation

To minimize overestimation of motility, it is recommended thatfresh semen be diluted with seminal plasma prior to automatedanalysis. However, for glycerolated or cryo-preserved sementhis is impractical, and alternative methods are needed to minimizeautomated motility bias. In the present study, the proportionof motile spermatozoa was determined in fresh, diluted and cryopreservedsemen (n = 25 ejaculates) using visual and automated methods.The effect of software settings on motility was investigatedby assessing samples at a range of modified settings. At standardsettings, automated motility was biased in fresh semen (+7.2%)after dilution with cryopreservative (–2.9%) and aftercryopreservation (–7.8%) (P < 0.0001 versus visual).Automated motility was inversely related to the minimum numberof frames for motility sampling (P < 0.0001), with mean estimatesof 41.0, 46.1, 52.0 and 58.2% generated at settings of 8, 4,2 and 1 frame(s) respectively (n = 15 fresh, diluted and cryopreservedsamples). Based on an arbitrary ordinal scale, a method wasdeveloped whereby motility sampling was adjusted prior to analysisaccording to sperm density. Analysis of an independent set ofsemen samples with density-adjusted software settings reducedbias in automated estimates (n = 30) before and after freezing(P < 0.0001). In addition, bias was no longer related tosperm density. In conclusion, modification of software settingsis an effective alternative to dilution to minimize bias inautomated motility estimates in fresh, diluted and cryopreservedhuman semen.     

Acrosomal status in fresh and frozen/thawed stallion spermatozoa evaluated by scanning electron microscopy

Semen from stallions with equal fertility at natural services, but yielding semen with either satisfactory or poor fertilizing capability after freezing and thawing, was processed for scanning electron microscopy before and after freezing and thawing. In all fresh semen samples the following three categories of acrosomal defect were noticed: (1) minor fenestrations of the plasma membrane (PM) and outer acrosomal membrane (OAM), (2) complete vesiculation and loss of PM and OAM and (3) lack of a large circular part of PM and OAM. The frequency of these defects ranged from 15% to 27%. All frozen and thawed samples displayed defects of categories 1–3 at similar frequencies as the fresh ones. However, additional defects categorized as: (4) major fenestrations of PM and OAM and (5) complete vesiculation of PM and OAM without loss of the vesicles were noted upon freezing and thawing. The total frequency of defects categorized as 4 and 5 ranged from 8% to 21%, and they seemed to be more frequent in stallions with poor fertility after freezing and thawing although the difference was not significant. Moreover, a particular defect categorized as (6) loosening of the whole acrosomal cap was found exclusively in stallions yielding semen with poor freezability.     

Improvement in motion characteristics and acrosome status in cryopreserved human spermatozoa by swim-up processing before freezing

The purpose of this study was to examine if selecting a sperm population with improved motion characteristics before freezing reduces the deleterious effects of cryopreservation. Semen specimens from 15 normal donors were divided into two equal aliquots. The first aliquot received no treatment (control), and the second was processed by swim-up from a washed sperm preparation to select a sperm population with better motility and motion characteristics (swim-up). Both aliquots were cryopreserved by the liquid nitrogen vapour method. Percentage motility and motion characteristics were evaluated by computer-assisted semen analysis. Acrosome integrity as well as spontaneous and calcium ionophore-induced acrosome reactions before freezing and after thawing were assessed by fluorescein isothiocyanate conjugated peanut agglutinin combined with a supra vital dye (Hoechst-33258). Swim-up processing enabled selection of a sperm population with better motion characteristics, percentage motility and viability before freezing (P < 0.001), but with no difference in percentage of acrosome-intact spermatozoa (P = 0.63). After thawing, the swim-up specimens exhibited faster velocity and progression than untreated specimens (P < 0.001). They also had higher percentages of spermatozoa with intact acrosomes and spermatozoa able to undergo acrosome reaction in response to calcium ionophore (P < 0.05). Selecting a highly motile sperm population before freezing enhances overall post-thaw spermatozoa quality.     

Dexamethasone therapy of infertile men with sperm autoantibodies: immunological and sperm follow-up.

A group of 14 infertile men with significant titres of anti-sperm antibodies in their sera and seminal plasma were treated with dexamethasone acetate at 2 of 3 mg/day for s13 or 9 weeks respectively. Response to the treatment was evaluated by the evolution of seminal and circulating anti-sperm antibodies and semen characteristics. An overall decline in the anti-sperm antibodies was observed. Serum spermotoxicity and seminal agglutinins decreased below detectable levels in 67 and 58% of the men respectively. The decrease in both kinds of antibodies was closely correlated. Serum sperm agglutination titres fell slightly (2 to 3 log2 dilutions) in most cases. The disappearance of antibodies from the semen was accompanied by decreased autoagglutination and increased percentages of progressively motile spermatozoa. In two oligozoospermic men a striking increase in the sperm count was observed. Pregnancies occurred in three couples at times when serum spermotoxic and seminal agglutinating antibodies were undetectable and semen characteristics were normal.     

The births of five Spanish babies from cryopreserved donated oocytes

BACKGROUND: The technique of freezing oocytes is still not widely used. Reasons cited for this include the technique's low efficacy and the risk of aneuploidy. However, the introduction of technical changes (the type and concentration of cryoprotective substances; slow freezing and rapid thawing; and fertilization by ICSI) has led to improved results. We present four pregnancies obtained using mature oocytes (in metaphase II) that had been frozen and thawed. The oocytes were donated by young women who were not patients. METHODS: The frozen oocytes (n = 88) came from seven donors aged 18-25 years. The metaphase II oocytes, morphologically normal in appearance, were denuded of their cumulus-corona complex. The cryoprotective freezing solution contained 1,2-propanediol (1.5 mol/l) and sucrose (0.3 mol/l). Freezing was slow and thawing rapid. The oocytes were fertilized by ICSI. RESULTS: Seventy-nine of the 88 thawed oocytes survived (89.8%); 58 were fertilized (73.4% of all those microinjected); and 26 were transferred (44.8% of all those fertilized). Four pregnancies were produced after seven transfers (57.1%). Five children were born from four pregnancies. CONCLUSIONS: With the freezing/thawing technique used, oocyte survival was high ( approximately 90%). The pregnancy rate with frozen oocytes was similar to that obtained using fresh oocytes from donors ( approximately 50%).