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PubMed1997

Enhancement of motility by treating spermatozoa with an antioxidant solution (Sperm-Fit) following ejaculation.

Oxygen radical generation is known to be detrimental to sperm function, especially motility, through the lipid peroxidation of the membranes. Generation of reactive oxygen species can be induced by leukocyte contamination, sperm centrifugation and the presence of abnormal spermatozoa with excess residual cytoplasm. This study aims to evaluate the effect on sperm motility of incubation in an antioxidant-containing solution, during liquefaction and centrifugation. Thirty semen samples were each divided into two equal parts: one mixed with Tyrode's solution, the other with a salt solution containing antioxidants (Sperm-Fit; Ellios Bio-Media, Paris, France). All the procedures were identical in the two groups. The ratio of leukocytes to spermatozoa was significantly correlated with the motility after liquefaction and after a 24 h incubation in routine in-vitro fertilization (IVF) medium and with the number of motile spermatozoa recovered after Percoll preparation. Moreover, when this ratio was > or = 0.2, all motility parameters were lowered. Incubation with Sperm-Fit allowed a higher percentage of motility after Percoll preparation when the ratio was > or = 0.2 (48 +/- 5% versus 41 +/- 6% for Sperm-Fit and Tyrode's solution respectively; P < 0.05) and a greater number of motile spermatozoa recovered after Percoll preparation, whatever the ratio (3.2 +/- 1.0 x 10(6) versus 2.4 +/- 0.7 x 10(6) for Sperm-Fit and Tyrode's solution respectively when ratio > or = 0.2; 18.1 +/- 3.4 x 10(6) versus 14.4 +/- 2.9 x 10(6) for Sperm-Fit and Tyrode's solution respectively when ratio < 0.2; P < 0.05). These results show that incubation with antioxidants during liquefaction and centrifugation increases recovery of motile spermatozoa.

Antioxidants↗
PubMed1997

Assisted ejaculation and in-vitro fertilization in the treatment of infertile spinal cord-injured men: the role of intracytoplasmic sperm injection.

The objective of the present longitudinal descriptive study was to extend previous observations on the benefit of in-vitro fertilization (IVF) in cases of anejaculatory infertility due to spinal cord injuries (SCI) and to report results achieved by intracytoplasmic sperm injection (ICSI). The study was performed in a national referral unit for SCI, Spinalis SCI Research Unit, the Karolinska Institute. The patient material consisted of couples with SCI men seeking treatment for their infertility. The inclusion criteria were: stable relationship, motile spermatozoa in a diagnostic sample and no female contraindications. Spermatozoa were retrieved through electroejaculation or vibratory stimulation. If the sperm quality was judged to be sufficient, standard IVF was performed. ICSI was employed if the semen quality was extremely poor. We have treated 25 couples in 52 cycles, leading to 81 ovum retrievals and 47 embryo transfers. Total sperm counts were very variable (0.01-978 x 10(6)). Before the introduction of ICSI the fertilization rate was 30%. ICSI increased the fertilization rate to 88%. There was no association between the pregnancy rate and the sperm count, level of injury or fertilization technique. A total of 16 clinical pregnancies was established, leading to 11 deliveries. This gives a cumulative pregnancy rate per couple of 56%.

Adult↗
PubMed2001

Chromatin status in human ejaculated spermatozoa from infertile patients and relationship to seminal parameters.

The aim of this study was to evaluate the chromatin status in different groups of patients. Five groups of men were selected: pre-vasectomy; male factor infertility; varicocele; immunological male infertility; and idiopathic infertility. Chromatin status was evaluated using flow cytometry after staining the DNA with the fluorochrome propidium iodide. Differences were observed in the state of sperm chromatin between the male factor and varicocele groups with respect to the others. These two groups presented poorer quality chromatin, as evidenced fundamentally by a lower degree of condensation. These deficiencies in chromatin status were usually accompanied by alterations in the other standard parameters of semen analysis. Individuals who are infertile due to male factor and those presenting varicocele have spermatozoa with less condensed chromatin which might, in part, explain their sterility.

Chromatin↗
PubMed2005

Apoptosis of ejaculated human sperm is induced by co-incubation with Chlamydia trachomatis lipopolysaccharide.

BACKGROUND: Previous work has shown that co-incubation of human sperm with Chlamydia trachomatis serovars E and LGV leads to premature sperm death and that this is due primarily to chlamydial lipopolysaccharide (LPS). Here, we investigated the possible involvement of apoptosis in this premature sperm death. METHODS: Highly motile preparations of sperm from normozoospermic patients were co-incubated for 6 h with extracted LPS from C. trachomatis serovars E and LGV. Three different methods were used to determine if LPS-treated sperm underwent apoptosis, including: (i) flow cytometry; (ii) measurement of ADP:ATP ratios; and (iii) measurement of mono- and oligonucleosomal DNA fragments. Caspase activity was also investigated by fluorimetry and by use of a pan-caspase inhibitor and caspase-3 inhibitor. RESULTS: All three methods used for detection indicated that C. trachomatis LPS induced some apoptosis in sperm after 6 h when compared with a staurosporine (apoptosis-positive) control. Moreover, a greater degree of apoptosis was seen with C. trachomatis serovar E than with serovar LGV. It was also shown that C. trachomatis LPS-induced apoptosis of sperm could be blocked with a pan-caspase inhibitor and a caspase-3 inhibitor. Moreover, by using a fluorogenic substrate, apoptosis was shown to be caspase-mediated. CONCLUSIONS: In general it is believed that apoptosis does not occur in C. trachomatis-infected host cells. However, using three different methods, our findings clearly indicate that co-incubation of sperm with C. trachomatis LPS results in cellular death which is in part due to apoptosis and is caspase-mediated. These findings provide an explanation as to how C. trachomatis can mediate premature death in human sperm.

Amino Acid Chloromethyl Ketones↗
PubMed2004

Analysis and significance of mRNA in human ejaculated sperm from normozoospermic donors: relationship to sperm motility and capacitation.

The existence of a complex population of mRNA in human sperm is well documented but their role is not yet elucidated. Using discontinuous density gradients, we have isolated high and low motile sperm from the same semen sample. The levels of different transcripts coding for molecules either involved in nuclear condensation (protamines 1 and 2) or in capacitation [endothelial nitric oxide synthase (eNOS), neuronal nitric oxide synthase (nNOS) and c-myc] were then assessed in the two populations using semi-quantitative RT-PCR. Sperm viability was estimated by eosin-nigrosin staining and by hypo-osmotic swelling test; apoptosis percentage was measured by the TdT (terminal deoxynucleotidyl transferase)-mediated dUDP nick-end labelling technique. The contamination by somatic and germ cells was assessed by looking for specific molecular markers of these cells, respectively CD-45 and E-cadherin for somatic cells and c-kit for germ cells. The viability of sperm was unchanged in high and low motile fractions, as well as DNA fragmentation percentage. The amount of Prm-1 mRNA was significantly higher in low density motile than in the high motile fraction. In most of high motile sperm samples eNOS and nNOS transcripts were undetectable whereas they were present in the low motile sperm. In contrast, no significant variation was found in the c-myc/Prm-2 mRNA ratio between the two populations. Moreover, a partial or complete disappearance of c-myc transcripts was observed after capacitation. Thus analysing mRNA profiles could be helpful as a diagnostic tool and prognosis value for fertilization.

Adult↗
PubMed2004

Cryopreservation and thawing is associated with varying extent of activation of apoptotic machinery in subsets of ejaculated human spermatozoa.

We investigated the impact of cryopreservation and thawing on levels of caspases-3, -8, and -9 activity, intact mitochondrial membrane potential (Deltapsim), and DNA fragmentation in human spermatozoa. Eleven pools of cryopreserved and eight pools of fresh semen samples were examined. Mature and immature fractions were separated on a two-layer density gradient (47% and 90%) and further subdivided based on the externalization of phosphatidylserine and its binding to annexin V-labeled superparamagnetic microbeads (ANMB). Levels of activated caspases were assessed using fluorescein-labeled inhibitors of caspases (FLICA), Deltapsim using a lipophilic cationic dye, and DNA fragmentation by the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assay. Cryopreservation was significantly associated with activation of caspases-3, -8, and -9, as well as disruption of the mitochondrial membrane potential but no significant changes were observed in DNA fragmentation. In mature sperm, caspase activation was only detected in the ANMB+ fraction, whereas in immature sperm, both ANMB+ and ANMB- fractions showed activated caspase levels. In ANMB+ immature sperm, apoptosis seemed to be triggered by a surface ligand-receptor mechanism as well as by disruption of mitochondria, whereas in ANMB- immature sperm, apoptosis was induced by activation of caspase-9 following loss of intact Deltapsim. These results demonstrate that selection of annexin V-negative mature spermatozoa might be of clinical relevance for fertility preservation, as this sperm fraction shows no activated apoptosis during the cryopreservation process.

Annexin A5↗
PubMed1982

A new proteinaceous factor participating in the rat ejaculate coagulation--identification, isolation and function.

A protein substance reacting with basic protein of rat seminal vesicle secretion, present in serum, testicular and epididymal homogenates and on unwashed spermatozoa, has been identified and isolated. The protein is partly of testicular origin, its concentration in male sera, testes and epididymides being age dependent and highest in sexually mature males. After castration, its concentration in male sera fails about 15%, to a comparable value present in female sera of the same age. The presence of this, as yet unrecognized, protein (molecular weight estimated at 163000) on epididymal spermatozoa is necessary for the in vitro coagulation of rat seminal vesicle proteins.

Aging↗