Chromomycin A3 staining, sperm chromatin structure assay and hyaluronic acid binding assay as predictors for assisted reproductive outcome.

Functional sperm tests such as the sperm chromatin structure assay (SCSA), chromomycin A3 staining (CMA(3)) and hyaluronic acid binding assay (HBA) have been suggested as predictive tests of fertility in vitro. This study aimed to define the clinical role of these functional parameters in assisted reproduction in a prospective cohort study. Conventional sperm diagnosis (motility, morphology and concentration) as well as SCSA, CMA(3) and HBA tests were performed on 205 semen samples [74 IVF, 94 ICSI and 37 combined IVF/intracytoplasmic sperm injection (ICSI)]. Main outcome parameters were fertilization rate, clinical pregnancy rate and take-home baby rate. The study showed that each of the three functional sperm tests was related to one or more conventional and one or more functional sperm tests, indicating that spermatozoa from patients with abnormal conventional semen parameters have a higher likelihood for multiple functional abnormalities. Only SCSA and CMA(3) staining were shown to have a limited predictive value when IVF or combined IVF/ICSI was applied. The proposed threshold value of

Influence of freeze-thawing on hyaluronic acid binding of human spermatozoa.

Mature human spermatozoa have at least three specific hyaluronic acid (HA) binding proteins present on their sperm membrane. These receptors play a role in the acrosome reaction, hyaluronidase activity, hyaluronan-mediated motility and sperm-zona and sperm-oolemmal binding. Cryopreservation of spermatozoa can cause ultrastructural and even molecular damage. The aim of this study was to investigate if HA binding receptors of human spermatozoa remain functional after freeze-thawing. Forty patients were enrolled in the study. Semen samples were analysed before and after cryopreservation. Parameters analysed included concentration, motility, morphology and hyaluronan binding. Samples were frozen in CBS straws using a glycerol-glucose-based cryoprotectant. HA binding was studied using the sperm-hyaluronan binding assay. Freeze-thawing resulted in a significant decline in motility: the percentage of motile spermatozoa reduced from 50.6 to 30.3% (P < 0.001). HA binding properties of frozen-thawed spermatozoa remained unchanged after the freeze-thawing process: 68.5 +/- 17.1% spermatozoa of the neat sample were bound to HA, as were 71.3 +/- 20.4 of the frozen-thawed sample. This study indicates that freeze-thawing did not alter the functional hyaluronan binding sites of mature motile spermatozoa, and therefore will not alter their fertilizing potential.

Reprotoxicity of intrauterine insemination and in vitro fertilization-embryo transfer disposables and products: a 4-year survey.

OBJECTIVE: To test consumables and other products used during oocyte collection, sperm preparation, IUI, embryo culture, and IVF-embryo transfer for their possible reprotoxic properties. DESIGN: A prospective 4-year survey of reprotoxicity testing of consumables and other products used in assisted reproductive technologies (ART). SETTING: Private infertility center in a university-affiliated teaching hospital. INTERVENTION(S): Thirty-six products of 72 different brands, including plastics, syringes, tubing, and surgical gloves were analyzed for their reprotoxicity in 350 human sperm survival tests (SpST). MAIN OUTCOME MEASURE(S): The SpST index: percentage progressive motility of test sample/percentage progressive motility of control sample after 24 and 96 hours. RESULT(S): Thirteen of 36 products were found to be reprotoxic: an SpST index <0.85 was noted 24 and 96 hours after exposure. These products included eight brands of unpowdered surgical gloves, two types of hysterometers and one type of tubing attached to the oocyte collection needle, one type of ovum pickup procedure needle, and one type of embryo transfer catheter. One type of condom used for ultrasound, one type of sterile Pasteur pipette and petri dish, as well as the cover of a specimen container, were reprotoxic. CONCLUSION(S): The SpST is an inexpensive, easy, and reliable method to identify potential reprotoxic products and consumables used in ART procedures. These data underline the importance of the inclusion of the SpST in a continuous quality control (QC) program of ART outcome.

Influence of the abstinence period on human sperm quality.

OBJECTIVE: To determine the influence of ejaculatory abstinence on within-subject semen parameters and DNA fragmentation. DESIGN: Prospective study. SETTING: Private infertility institute and university-based research laboratory. PATIENT(S): Sixteen consenting male volunteers undergoing infertility investigation. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Within-subject analysis of World Health Organization semen parameters and sperm DNA fragmentation and chromatin packaging after 1, 3, 5, and 8 days' abstinence. RESULT(S): Of 16 men recruited, data for 11 men were included for statistical analysis because 5 men did not strictly comply with abstinence criteria. Duration of abstinence had a statistically significant positive influence on sperm concentration and semen volume. Abstinence had no statistically significant influence on pH, viability, total and grade A motility, or morphology. The percentage of DNA fragmentation remained unchanged relative to abstinence. The percentage of sperm with immature chromatin was statistically significantly increased with 1 day of abstinence. CONCLUSION(S): This is the first study to report on within-subject semen parameter, DNA fragmentation, and chromatin packaging variations after specified target days of abstinence. Sperm numbers and semen volume increased with duration of abstinence. Abstinence did not influence pH, viability, morphology, total or grade A motility, or sperm DNA fragmentation. A short (24-hour) abstinence period negatively influenced chromatin quality.

Semen quality and intrauterine insemination.

There is good evidence in literature that intrauterine insemination (IUI) is the best first line treatment and most cost-effective procedure for moderate male factor subfertility. It seems very difficult to identify individual semen parameters predicting the likelihood of pregnancy after IUI. This can be explained by a lack of standardization of semen analysis, but many other methodological variables may also influence IUI success rates such as the patient selection, type of ovarian stimulation and number of inseminations per cycle. A review of the literature confirmed that sperm morphology using strict criteria and the inseminating motile sperm count (IMC) after sperm preparation are the two most important sperm parameters to assess the real impact of semen quality on IUI outcome. A universal threshold level above which IUI can be performed with acceptable pregnancy rates has not been determined yet, although IUI success seems to be impaired with <5% normal spermatozoa and an IMC of <1 x 10(6). Until now, no method of sperm preparation has been shown to be superior with regard to pregnancy rate after IUI. Whether supplementation of culture media with substances such as antioxidants and platelet activating factor may improve the results remains the subject of further research.