Category Archives: Human Reproduction

BACKGROUND

Ovulation induction treatment with metformin, either alone or in combination with clomiphene citrate (CC), remains controversial even though previous randomized trials have examined this.

METHODS

A double blinded multi-centre randomized trial was undertaken including 171 women with anovulatory or oligo-ovulatory polycystic ovary syndrome. Women with high body mass index (BMI) > 32 kg/m² received placebo (‘standard care’) or metformin; women with BMI ≤ 32 kg/m² received CC (‘standard care’), metformin or both. Treatment continued for 6 months or until pregnancy was confirmed. Primary outcomes were clinical pregnancy and live birth.

RESULTS

For women with BMI > 32 kg/m², clinical pregnancy and live birth rates were 22% (7/32) and 16% (5/32) with metformin, 15% (5/33) and 6% (2/33) with placebo. For women with BMI ≤ 32 kg/m², clinical pregnancy and live birth rates were 40% (14/35) and 29% (10/35) with metformin, 39% (14/36) and 36% (13/36) with CC, 54% (19/35) and 43% (15/35) with combination metformin plus CC.

CONCLUSIONS

There is no evidence that adding metformin to ‘standard care’ is beneficial. Pregnancy and live birth rates are low in women with BMI > 32 kg/m² whatever treatment is used, with no evidence of benefit of metformin over placebo. For women with BMI ≤ 32 kg/m² there is no evidence of significant differences in outcomes whether treated with metformin, CC or both.

ClinicalTrials.gov number NCT00795808; trial protocol accepted for publication November 2005: Johnson, Aust N Z Journal Obstet Gynaecol 2006;46:141–145.

BACKGROUND

Although female genital mutilation (FGM) does not feature in Judeo–Christian populations, it is estimated that, 100–140 million women in the world have undergone some form of FGM. Given the increasing diversity of the western populations, a review of specific complications of FGM is of paramount importance to practicing clinicians. The objective of this study is to report a case series of epidermal clitoral inclusion cysts after FGM in a Muslim population primarily from the Middle East.

METHODS

Between January 1998 and July 2009, 32 females underwent surgical removal of epidermal clitoral inclusion cysts in a tertiary referral university hospital. Data regarding age, clinical presentation, operation time, estimated blood loss, presence of intraoperative and post-operative complications, duration of admission to the hospital and long-term follow-up were extracted from the records.

RESULTS

There were 15 women (46.9%) with a definitive history of FGM, 14 (43.8%) did not know whether they had FGM or not and 3 (9.3%) had no history of FGM and were excluded from the analysis. The mean age of subjects was 28.1 years (range 5–91 years). All presented with increasing clitoral mass over a mean duration of 5.2 ± 4.1 years. The mean diameter of the cyst was 4.2 ± 2 cm. Regarding treatment, 28 subjects underwent surgical excision, and one underwent incision and drainage of a clitoral abscess. No short- or long-term complications occurred.

CONCLUSIONS

Clitoral cysts appear to be a more common complication of FGM than previously thought. Publication of studies that highlight the medical complications of FGM should be encouraged to advocate abandonment of the procedure.

BACKGROUND

It has been identified that human epididymal protease inhibitor (EPPIN) plays a critical role in sperm function and male fertility. The aim of this study was to determine whether variants of the EPPIN gene are risk factors for idiopathic male infertility.

METHODS

All subjects, including 473 idiopathic infertile men and 198 fertile controls, underwent complete historical and physical examinations. Each subject donated 5 ml of peripheral blood for genomic DNA extraction and serum testosterone evaluation and an ejaculate for semen analysis. The semen analysis was performed by computer-assisted semen analysis system. The serum testosterone level was evaluated by radioimmunoassay. Four tagging single-nucleotide polymorphisms were analyzed by polymerase chain reaction–restriction fragment length polymorphism.

RESULTS

We have demonstrated a significant decreased risk of idiopathic infertility with abnormal semen parameters in association with the variant rs2231829, and an increased risk of idiopathic infertility with abnormal semen parameters in association with the variant rs11594. However, among men with normal semen parameters, there were no differences in risk for these genotypes. Furthermore, no significant differences were found for the other variants, rs6124715 and rs2227290, on the risk of male infertility with normal or abnormal semen parameters. Similar serum testosterone levels among different EPPIN genotypes were observed for each group.

CONCLUSIONS

These results suggest that different variants in the EPPIN gene may have different relationships with idiopathic male infertility and men carrying these variants have a decreased or increased risk of abnormal semen parameters associated with male infertility.

BACKGROUND

It has been suggested that the zona pellucida (ZP) may mediate species-specific fertilization. In human the ZP is composed of four glycoproteins: ZP1, ZP2, ZP3 and ZP4. In the present study, the expression profile of ZP1 in human oocytes and ovaries, and its role during fertilization, is presented.

METHODS

Human ZP1 (amino acid residues 26–551) was cloned and expressed in both non-glycosylated and glycosylated forms and its ability to bind to the capacitated human spermatozoa and to induce acrosomal exocytosis was studied. Monoclonal antibodies (MAbs), specific for human ZP1 and devoid of reactivity with ZP2, ZP3 and ZP4 were generated and used to localize native ZP1 in oocytes and ovarian tissues.

RESULTS

The MAbs generated against ZP1 recognized specifically the zona matrix of secondary and antral follicles, ovulated oocytes, atretic follicles and degenerating intravascular oocytes, but failed to react with the Fallopian tube, endometrium, ectocervix and kidney. Escherichia coli and baculovirus-expressed recombinant human ZP1 revealed bands of ~75 and ~85 kDa, respectively, in western blot. Lectin binding studies revealed the presence of both N- and O-linked glycosylation in baculovirus-expressed ZP1. Fluorescein isothiocyanate-labelled E. coli- and baculovirus-expressed recombinant ZP1 bound to the anterior head of capacitated spermatozoa, however, only baculovirus-expressed ZP1 induced acrosomal exocytosis in capacitated sperm suggesting the importance of glycosylation in mediating the acrosome reaction. The human ZP1-mediated acrosome reaction involved the activation of both T- and L-type voltage-operated calcium channels, but does not activate the G_i-coupled receptor pathway. Inhibition of protein kinase A and C significantly also reduced the ZP1-mediated induction of the acrosome reaction.

CONCLUSION

These studies revealed for the first time that in humans ZP1, in addition to ZP3 and ZP4, binds to capacitated spermatozoa and induces acrosomal exocytosis.

BACKGROUND

To better understand the infertility of patients with Robertsonian translocation, the biochemical and ultrastructural apoptotic characteristics of apoptosis in the sperm of patients and fertile donors were studied.

METHODS

Ejaculated sperm samples of seven Robertsonian translocation carriers and seven fertile donors were analyzed after cryopreservation. The proportion of both viable and dead spermatozoa expressing activated caspases was detected by flow cytometry through the use of different specific carboxyfluorescein-labeled caspase inhibitors. Sperm DNA fragmentation was evaluated by the TUNEL method. The percentages of intact spermatozoa or spermatozoa with ultrastructural features of apoptosis, immaturity or necrosis were estimated by electron microscopy. Meiotic segregation analysis was performed by FISH.

RESULTS

Significantly lower concentration, forward motility and normal morphology of spermatozoa were found in ejaculated samples of the Robertsonian patients than fertile donors. Compared with the control group, in Robertsonian translocation carriers: (i) the caspase assays showed a significantly increased (P < 0.05) proportion of viable spermatozoa with activated poly-caspases (57.4 versus 25.8%), caspase-3 (43.5 versus 13.4%), caspase-8 (44.4 versus 17.1%) and caspase-9 (42.4 versus 10.0%); (ii) the rate of DNA fragmentation was higher (26.3 versus 12.8%); and (iii) sperm ultrastructural examination highlighted a higher percentage of immature (28.0 versus 10.0%) and apoptotic (24.5 versus 18.5%) spermatozoa. FISH study showed predominant normal/balanced spermatozoa (78.34–85.53%).

CONCLUSIONS

These results show a predominant proportion of balanced and normal gametes and higher numbers of spermatozoa showing apoptosis and immaturity features in oligoasthenozoospermic Robertsonian translocation carriers than in fertile donors. This suggests defects in spermatogenesis and especially spermiogenesis of these infertile patients.

BACKGROUND

Semenogelin (Sg), the main protein of human semen coagulum, prevents sperm capacitation. The objective of this study was to examine the role of Sg and its mechanism of action.

METHODS AND RESULTS

Sg blocked sperm capacitation triggered by various stimuli, via inhibition of superoxide anion (O₂•–; luminescence assay) and nitric oxide (NO•; tested using diaminofluorescein) generation. Triton-soluble and -insoluble sperm fractions contained Sg and Sg peptides (immunoblotting), the level of which decreased with initiation of capacitation. This drop was prevented by superoxide dismutase and NO• synthase inhibitor and was reproduced by addition of O₂•– and NO•. Zinc (Zn²⁺) blocked and a zinc chelator (TPEN) promoted the decline in Sg levels. There was a decreased labelling of Sg on the head in capacitating spermatozoa with the two fixation techniques tested (immunocytochemistry). Reactive oxygen species (ROS) (O₂•– and NO•) caused, these changes, and zinc prevented them. Spermatozoa quickly internalized Sg upon incubation and Sg was then rapidly degraded in a zinc-inhibitable manner.

CONCLUSIONS

Sg blocked capacitation mainly via inhibition of ROS generation. Spermatozoa appeared permeable to Sg and processed Sg in a zinc-inhibitable fashion. ROS themselves could promote sperm disposal of Sg which maybe one of the mechanisms that allows initiation of capacitation.