Marco Balerna

Effect of peritoneal fluid on sperm motility and velocity distribution using objective measurements

: The aim of this study was to analyze in vitro the effect(s) of peritoneal fluid (PF) on sperm motility. Seventy PFs obtained during laparoscopy were tested on motile-rich sperm suspensions. Proportion of motile sperm and velocity distribution were evaluated by multiple-exposure photography technique. At time (t) = 0, PFs increased both sperm parameters as compared with control (P less than 0.01). Maximum effect was observed at t = 5 hours: 32 (45%) PFs increased and 5 (7%) decreased the proportion of motile sperm, while 8 (11%) PFs increased and 4 (6%) decreased sperm velocity. No correlation was found between a particular infertile group and a definite negative effect. However, 70% of PFs from fertile women maintained or increased the proportion of motile sperm at t = 5 hours, compared with 36% in the total infertile group. Comparison of the sperm motility effect(s) of a given PF on different ejaculates revealed that the effects observed also were influenced by the sperm sample tested. In conclusion, PFs can maintain or increase the motility of spermatozoa as function of time. However, some PFs can inhibit sperm motility and these effect(s) can be influenced by the sperm sample.

Oocyte penetration and acrosome reactions of human spermatozoa. I: Influence of incubation time and medium composition

Summary:  Washed sperm suspensions were evaluated for their ability to penetrate zona‐free hamster oocytes and to exhibit an acrosome reaction in vitro. The percentage of acrosome reactions (AR%) was found to increase with incubation time and increased bovine serum albumin (BSA) concentration. We also found, however, that a remarkably high number of live, unreacted spermatozoa persisted during prolonged incubation in sperm samples obtained from some fertile men. After 22 hrs incubation, the motility of the spermatozoa was higher when the medium was supplemented with 10% human cord serum instead of 3.0% BSA. Penetration rates in hamster oocytes were not significantly different with 10% serum or 3.0% BSA as medium‐supplementation. The addition of human instead of bovine serum albumin did not apparently affect the oocyte penetration rate of the spermatozoa.

Oocyte Penetration and Acrosome Reactions of Human Sperms II: Correlation with other Seminal Parameters

Summary:  Washed sperm suspensions of fertile men and men consulting for infertility were evaluated for their ability to penetrate zona‐free hamster eggs and for their ability to exhibit an acrosome reaction in vitro. Furthermore, the swelling of the spermatozoa under hypoosmotic conditions as indication of their membrane integrity was determined. In the group of fertile men and in the group of patients with normal spermiogram, significantly more acrosome reactions were observed than in the group of infertile men with abnormal spermiogram parameters (p < 0.05). This difference was still more significant when men with a positive hamster penetration test (H.O.P. test) and men with a negative H.O.P. test were compared (p < 0.005). However, within the groups the level of acrosome reactions after incubation appeared to be highly variable.

Bacterqides ureolyticus in men consulting for infertility

Summary: A screening of 3196 semen analyses performed in our clinic from January 1986 to December 1990 revealed 314 (9.8%) patients whose semen was infected with Bacteroides ureolyticus. Investigating the relationship between the presence of B. ureolyticus, the seminal microflora and the conventional semen parameters, we observed that the presence of this micro‐organism in the semen was coupled (1) to an increased presence of Enterococcus species, (2) to an increased number of short‐tailed spermatozoa and epithelial cells, and (3) to a decreased total fructose concentration (mg ejaculate−1). These results suggest that B. ureolyticus or its toxins may influence sperm morphology and function by yet unknown mechanisms and may also increase the number of epithelial cells by soft tissue infection in vivo. The decreased fructose levels suggest that this anaerobic micro‐organism might specifically colonize the seminal vesicles, while the normal zinc values recorded suggest a normal prostatic function. Overall, our data support the hypothesis that the presence of B. ureolyticus is not associated with nongonococcal urethritis.

Interference in thyroid‐stimulating hormone determination

Eur J Clin Invest 2010; 40 (8): 756–758

Computer-aided semen analysis: sperm concentration assessment by the Strömberg-Mika system

Summary The scope of this study was to evaluate the accuracy, precision and specificity of the sperm concentration measurements by the Strömberg‐Mika Cell Motion Analyser (SM‐CMA). Our data show that the instrument generally underscores the sperm concentration and therefore the uncorrected measurements must be corrected by the operator using the ‘mouse’‐driven option. In terms of precision, the system appears to have an excellent internal precision whereas its repeatability is influenced by the sperm concentration, the samples homogeneity and the correction of the raw data. In order to increase the systems repeatability, we suggest that sperm counts should be carried out in various fields of the counting chamber, and the mean of the corrected values be taken as representative of the sperm concentration in the ejaculate if the various measurements show a homogenous (poissonian) distribution. The correction of the raw data with the ‘mouse’‐driven correction option was also shown to improve the systems reproducibility. Concerning specificity, our data evidenced that, without technical correction, the instrument failed to correctly classify certain spermatozoa as such, thereby grossly underscoring sperm counts. This finding was more evident at low sperm counts. Overall, the SM‐CMA requires additional laboratory time but the corrected sperm counts are comparable to manual counts and semi‐automated counts with the added option that it provides the andrologists with various motility characteristics not possible with the latter methodologies.

Etiology of severe asthenozoospermia and fertility prognosis. A screening of 5216 semen analyses

Summary. A review of n = 5216 semen analyses performed in our two Clinics from January 1986 to December 1989 allowed to identify n = 35 patients whose sperm had constantly very low motility (< 5% progressive motile gametes in three subsequent analyses; necrozoospermia cases were excluded from this study). This apparently rare but severe anomaly was found to be associated not only with ultra‐structural anomalies (n = 18), but also with positive seminal bacteriology (n = 8) or the presence of antisperm antibodies (n = 2). In eight cases the cause(s) for this constant asthenozoospermia remained obscure. The fertility potential of the men affected was followed‐up and is discussed in relation to their anamnesis, physical exam and seminal characteristics.

Influence of peritoneal fluid from spontaneous and stimulated cycles on sperm motility in vitro.

Summary. Peritoneal fluids (PFs) from spontaneous (n= 14) and gonadotrophin‐stimulated cycles (n= 20) were obtained during diagnostic laparoscopy and gamete intrafallopian transfer (GIFT) procedures, respectively. The effects of these fluids on the linear component of sperm motility and on the percentage of motile spermatozoa were studied in vitro by objective motility assessments and compared to a control medium (B2‐Ménézo).

Removal of sperm‐coat from human spermatozoa by interaction with cervical mucus or a capacitating medium: Entfernung des Sperm‐Coats bei menschlichen Spermien durch Interaktion mit Zervixschleim oder mit einem kapazitierenden Medium

Summary Human spermatozoa were labelled with cationized ferritin and their interaction with cervical mucus or a capacitating medium enriched with 3% bovine serum albumin (BSA) was investigated by transmission electron microscopy. It was found that the spermcoat was removed from the surface of spermatozoa after incubation in the BSA‐enriched capacitating medium, as well as after crossing a column filled with cervical mucus. These results suggest that both media have a quantitatively similar action in removing the spermcoat.

Preincubation in peritoneal fluid decreases the follicular fluid‐induced acrosomal reactivity of human spermatozoa

Summary. The aim of this study was to determine the effects of preincubation in peritoneal fluid on the follicular fluid‐induced acrosomal reactivity of human spermatozoa in vitro. Thirty women participating in our IVF‐EL program were given a GnRH‐analogue, highly purified FSH and hCG in order to induce superovulation. Peritoneal and follicular fluids were aspirated during pick‐up laparoscopy, centrifuged, filtered and frozen until use. An aliquot of swim‐up suspension from nor‐mospermic semen specimens (n=30) was incubated with peritoneal fluid or HAM‐F10 for 30–180 min, and follicular fluid (in volumetric proportion approximately 50/50 with peritoneal fluid) was subsequently added. The percentage of acrosomally‐reacted spermatozoa was assessed using the FITC‐conjugated Pisum sativum lectin before and after incubation in peritoneal fluid or control medium, as well as after follicular fluid addition. Peritoneal fluid was not able to stimulate acrosomal reactivity; further, preincubation in peritoneal fluid decreased, but not abolished, the follicular fluid‐induced acrosomal reactivity. A longer pre‐incubation in peritoneal fluid was associated with a lower percentage of reacted spermatozoa in response to the addition of follicular fluid. In conclusion, our data suggest that peritoneal fluid acts maintaining spermatozoa in an unreacted status in the upper female genital tract. After mixing with follicular fluid, a phenomenon that is likely to occur at ovulation, peritoneal fluid reduces, but does not abolish, the stimulating effect of follicular fluid on acrosomal reactivity.