AIMTo evaluate the long-term stability of the fluorescence signals of new fluorescence-based semen analysis assays for clinical application.

METHODSSemen samples from 87 unselected infertile patients were used to perform the following assays: (i) detection of active caspase-3 (n=17); (ii) integrity of the mitochondrial membrane potential (MMP) (n=17); (iii) externalization of phosphatidylserine (EPS) (n=16); and (iv) detection of intact acrosomes via CD46 (n=37). After the assays, 4% paraformaldehyde was added to all aliquots. The fluorescence intensity of each sample was evaluated by flow cytometry on days 0, 3, 7, 10 and 14.

RESULTSDifferences of up to +/-5% positive spermatozoa from the value measured at day 0 were estimated as acceptable deviation. The Caspase-3 FLICA showed mean differences<5% at day 3, 7 and 10. At day 14 the mean difference was 7.6%. In contrast, the disrupted MMP and the EPS detection showed differences>5% at day 3. The CD46-FITC labeling displayed absolute differences<5% CD46-positive spermatozoa at days 3, 7, 10 and 14.

CONCLUSIONAlthough immediate analysis of the fluorescence signals is recommended, it is possible to evaluate caspase-3 activation up to 10 days and CD46 up to 14 days after staining of sperm. The FACS evaluation of MMP and EPS detection should be conducted on the same day.

OBJECTIVESTo evaluate the effects of cryopreservation on human sperm chromosome.

METHODSperm chromosome were acquired using in vitro fertilization of zona-free hamster oocytes and human sperm. The frequency of sperm chromosome anomalies and sex chromosomes ratio before and after freezing and with different freezing methods were compared.

RESULTSThere were no significant differences of frequency of sperm chromosome anomalies among fresh, fast frozen and slow frozen sperm (9.40%, 7.48% and 8.74%) (P > 0.75) or ratios of sex chromosomes (P > 0.90).

CONCLUSIONSThese studies indicate that cryopreservation does not exert effects on human sperm chromosome.

Sperm cryopreservation is essential link of assisted reproductive techniques. Because of widespread application for AID and establish of human sperm storeroom, the effects of cryopreservation on human sperm genetic substance have been paid more and more attention. This report summarizes the effects of cryopreservation on human sperm genetic substance and preventive measures.
Chromosomes ; Cryopreservation ; Humans ; Male ; Spermatozoa ; physiology

Recent studies show that long exposure to high altitude and hypoxia can seriously affect men's reproductive health by reducing their sperm concentration, which decreases with the increase of altitude. High altitude and hypoxia are strongly associated with spermatogenic reduction, sperm DNA damage, sperm apoptosis, and decreased level of sex hormones. This article reviews the mechanisms of high altitude and hypoxia affecting sperm concentration.
Altitude ; Humans ; Hypoxia ; Male ; Sperm Count ; Spermatogenesis ; Spermatozoa ; physiology

OBJECTIVETo evaluate the application of distilled water in sperm-counting and hypoosmotic swelling test.

METHODSThirty-seven semen samples were collected and each was diluted by distilled water and sodium acid carbonate-formaldehyde solution, respectively. Then the hemacytometer was used for sperm counting. Meanwhile, the percentage of swelled sperm diluted by distilled water was compared with the result of hypoosmotic swelling test recommended by WHO. Another 26 semen samples were diluted by distilled water and hypoosmotic swelling solution respectively, and the percentages of the swelled sperm were compared.

RESULTSThere was no significant difference either between the sperm concentrations obtained by distilled water and sodium acid carbonate-formaldehyde solution (P > 0.05) or between the percentages of the swelled sperm diluted by distilled water and hypoosmotic swelling solution.

CONCLUSIONDistilled water can not only replace sodium acid carbonate-formaldehyde solution for sperm-counting dilution but also be used as a hypoosmotic swelling solution.

To investigate the influences of sperm quality on the zygotes and embryos development, as the role of the paternal factor in early human embryogenesis is gaining more attention because of the application of techniques such as intracytoplasmic sperm injection (ICSI) for the treatment of men infertility. 136 infertility couples with men factors (Group I ) were included from May 2002 to January 2001. One hundred and seventy-two infertility couples with tube factors (Group II) served as controls. The sperm parameters, gemmates and embryos quality, implantation rate and pregnant rate in both groups were analyzed. It was found that there was no significant differences in the number of oocytes retrieved, the fertilization rate and number of embryos transferred between two groups. Sperm concentration, percentage of motile sperm and percentage of sperm with normal morphology were significantly lower in group I than in group II (P < 0.01). The proportion of good quality zygotes and good quality embryos were significantly lower in the male infertility group than in the tubal disease group (P < 0.05). Implantation rate and pregnancy rate were similar in two groups. It was concluded that spermatozoa is involved in the embryo quality, even in the early stages of development, which limited the treatment potency of IVF procedure.
Embryo Transfer ; Embryonic Development ; Fertilization in Vitro ; Infertility, Male ; Sperm Count ; Sperm Injections, Intracytoplasmic ; Sperm Motility/*physiology ; *Spermatozoa/cytology ; *Spermatozoa/physiology

In order to explore the effects of testicular infection of murine cytomegalovirus (MCMV) on mature sperm viability at different periods following MCMV inoculation in mice, 91 BALB/c mice without MCMV infection were randomly divided into two groups: an experimental group (n = 56) and a control group (n = 35). The mice in the experimental group were treated by inoculating MCMV intratesticularly, while those in the controlled group were directly inoculated with DMEM without MCMV. The mice in both groups were sacrificed separately on the day 1, 1. 5, 2, 4, 6, 9 and 14 post-inoculation (D1) 1. 5, 2, 4, 6, 9 and 14 PI). The MCMV M83 mRNA gene was detected in the testis by in situ hybridization (ISH) with MCMV late-mRNA probe labeled with digoxin. Sperm viability of mature sperm in the epididymis cauda was measured. The results demonstrated the positive signal of ISH of MCMV was found mainly in the cytoplasm of the testicular interstitial cells and spermatogenic cells in the experimental group. Compared with that in the controlled group, the sperm viability in the experimental group was decreased significantly on D1 PI and D1.5 PI (P < 0.05). No statistically significant difference in the sperm viability was found after D2 PI between two groups (P > 0.05). This suggested that sperm viability in mice might be descended significantly shortly after MCMV infection and might return to normal with time, indicating that MCMV acute infection might temporarily degrade sperm quality and influence procreation transiently.
Cytomegalovirus Infections/*physiopathology ; Mice, Inbred BALB C ; Orchitis/*virology ; Random Allocation ; Sperm Motility/*physiology ; Spermatozoa/cytology ; Spermatozoa/*physiology

Progesterone, as a female hormone, plays an important role in the physiological function and pregnancy maintenance in women. Recent studies show that progesterone and its receptor are also involved in male reproduction, and its receptor mRNA exists in male sexual glands. It is believed that progesterone, binding to its receptor, can regulate spermatogenesis and improve the fertilization of sperm, while the sperm from those with oligospermia, asthenozoospermia, teratospermia or unexplained infertility exhibit a low fertility due to the deficient expression of the progesterone receptor and insensitive reaction to progesterone. This review focuses on the progress in the studies of progesterone and its receptor in male reproduction.
Fertility ; Humans ; Male ; Progesterone ; physiology ; Receptors, Progesterone ; physiology ; Spermatogenesis ; Spermatozoa ; physiology ; Testis ; metabolism

The epididymal protease inhibitor (Eppin) has recently been cloned in human and mice, which is specifically expressed in the epididymis and testis. Eppin is a cystine-rich secretory protein which contains signal peptide, WAP and BPTI motifs. Eppin is involved in sperm maturation and fertilization, and in the innate immune system of human epididymis. Immunocontraception with Eppin is effective and reliable, but its safety is to be further proved. This paper summarizes the effects of Eppin of fertilization and immunity, as well as its utilization in immunocontraception.
Animals ; Contraception, Immunologic ; Epididymis ; immunology ; Fertility ; physiology ; Haplorhini ; Male ; Mice ; Proteins ; immunology ; physiology ; Spermatozoa ; physiology

Protein kinase C (PKC) is localized in the equatorial segment and the principal piece of the tail of spermatozoa. Activator of PKC results in increasing flagellar motility of sperm that is blocked by PKC inhibitors such as staurosporine. A good correlation (r = 0.9, P < 0.001) is found between the content of PKC in sperm and sperm motility. Zona pellucida (ZP) stimulates the spermatozoa binding the acrosome reaction resulting in the release of hydrolytic enzymes and in the exposure of new membrane domains. ZP binding to receptors in the plasma membrane can regulate adenyl cyclase (AC) leading to elevation of cAMP and protein kinase A (PKA) activation. The PKA activates a voltage-dependent Ca2+ channel in the outer acrosomal membrane which releases Ca2+ from the interior of the acrosome to the cytosol. Activation of the PLC resulted from the rise in Ca2+ hydrolyze phosphatidyl inositol bisphosphate. The product activate PCK to open a voltage-dependent Ca2+ channel (L) in the plasma membrane, leading to the second (II) Ca2+ higher increase which result in membrane fusion and acrosome reaction. It is proposed that PKC would be involved in the regulation of motility and acrosome reaction of sperm.
Acrosome Reaction ; physiology ; Humans ; Male ; Protein Kinase C ; metabolism ; Sperm Motility ; physiology ; Spermatozoa ; enzymology ; physiology