[MESA (microsurgical epididymal sperm aspiration) and IVF (in vitro fertilization). A therapy concept in treatment of male infertility]. / MESA (mikrochirurgische epididymale Spermienaspiration) und IVF (In-vitro-Fertilisation). Ein Therapiekonzept zur Behandlung der männlichen Infertilität.

Microsurgical epididymal sperm aspiration (MESA), or sperm microaspiration retrieval technique (SMART), in conjunction with in vitro fertilization is a successful therapy in male infertility. From November 1991 to March 1994 a total of 29 attempts at MESA with subsequent IVF were made. Of 48 aspirations, 37 were successful and 13 attempts at IVF were possible, 6 of which were successful with 10 subsequent embryo transfers. In all, 3 pregnancies were achieved and 1 boy was born. In conclusion, microsurgical spermaspiration in conjunction with in vitro fertilization is a way of treating male infertility with a chance of achieving paternity with the partner's own sperm; the chances are probably better with intracytoplasmatic sperm injection.

[Cryopreservation of human spermatozoa--evaluation of 93 semen depots]. / Kryokonservierung humaner Spermatozoen--Auswertung von 93 Samenlangzeitdepots.

Cryopreserved semen depots from 93 patients who had terminated contract with the CRYO-BANK-KREFELD were evaluated for prefreeze quality and post-thaw motility after a few days and after long-term storage (mean = 44 month). 2/3 of these patients had onkological disease. Only 20% of the ejaculates were normozoospermic and showed motility after thawing of more than 20%. The average recovery rate after long term storage was only 28% and revealed a significant decrease with time of about 20%. The concentration of motile spermatozoa, especially in patients with testicular cancer (x = 2.1 mill/ml), was so much reduced one must assume that in the most cases of semen banking there was no chance to achieve pregnancy.

Prognostic value of the sperm-penetration-meter test according to Kremer.

An eighteen month follow up of 84 couples with unexplained infertility was analysed to determine the significance of the sperm-penetration-meter test (SPM-test). Pregnancy occurred independently of treatment in 16 couples (19%). The fertile and infertile group displayed no difference in the classical semen parameters such as volume, density and morphology. Only motility was slightly depressed in the infertile collective. The penetration quality was assessed at 2,4 and 24 hours incubation time according to Kremer (1980). Of all parameters only the two- and four hour values showed a significant, although poor correlation with the pregnancy rate (r = 0.32; p less than 0.01). Furthermore, the fertile group achieved twofold more positive results after two and four hours incubation (75% and 69%) than the group with persistent infertility (34% and 31%). In both collectives more than 90% of the 24 hour values were negative and thus insignificant. From all conventional semen parameters motility had the greatest prognostic point of view the results of the SPM-test bear the greatest significance during the first four hours.

[Crossed sperm-penetration-meter test. Prognostic value in male subfertility]. / Gekreuzter Sperm-Penetration-Meter Test. Prognostische Wertigkeit bei der androgenen Subfertilität.

57 infertile couples with "unknown" or "therapy resistant" infertility were investigated by the double crossed sperm-penetration-meter-test (SPM-test). The results of the SPM-test were compared with the results of the postcoital-tests, the duration of infertility, and the male and female diagnosis of infertility. Postcoital tests and SPM-tests showed only in 63% of cases corresponding results. The ability of the spermatozoa to penetrate cervical mucus was directly depending from the number of progressively motile spermatozoa per ml, and no connection could be found between the results of the SPM-test and the gynecological findings. The investigations showed that the primary diagnosis of infertility can be extended by the SPM-test respectively in many cases of so-called "unknown infertility" where a disturbed spermatozoa-mucus-interaction has to be postulated.

A new instant cryoprotectant for human sperm.

A new instant cryoprotectant, mainly containing culture medium, human serum albumin, and kallikrein, and its laboratory handling in the cryoprotection of semen are described. A comparative study distributed to three cryobiological centers using either egg yolk citrate medium or instant cryoprotectant showed that neither had a significantly better or worse mean percentage recovery rate. The advantage of the new instant cryoprotectant presented consists clinically in the absence of egg yolk and technically in its easy handling. Therefore, ICP is judged as an alternative to ECM in the cryopreservation of human sperm.

Further evaluation of the split ejaculate for artificial insemination.

The role of artificial insemination with husband semen (AIH) in solving fertility problems has been studied by many investigators. Different conclusions have been made due to the variety of indications for AIH. This communication attempts to evaluate the use of the best split ejaculate fraction for AIH in 55 couples in which the only pathological condition was oligozoospermia or oligoasthenozoospermia. Couples in which the women had a known history of anatomical abnormalities were excluded. Although the seminal parameters of spermatozoal density and/or motility in the best split fractions were frequently found to be in the fertile range, the pregnancy rate achieved was only 18% (n = 10). This was comparable to the results obtained by others using whole ejaculates for AIH. The spontaneous pregnancy rate in the present study after termination of treatment was 13% (n = 6). The observations suggest that men with a long-standing history of seminal deficiency produce spermatozoa with a pathology that cannot be microscopically recognized.

[Menstrual cycle changes during consecutive artificial insemination therapy and their treatment]. / Zyklusstörungen während konsekutiver artefizieller Inseminationstherapie und ihre Behandlung.

Emotional stress-induced problems generated by the application of artificial insemination therapy by donor (AID) are considerable. After 6 months of frustrane insemination treatment, 54% of the women exhibited irregular or anovulatory cycles. Especially the persistent late ovulation often leads to a failure of the mechanisms of conception. An early ovulation timing with high dosages of synthetic LH-releasing hormone or a stimulating LH-RH-analogue (HOE 766) lead to an increasing pregnancy rate after application in the third insemination cycle from 34,0% untreated to 51,1% treated. The early application of ovulation inducing or timing substances is underlined in a program of artificial insemination therapy to eliminate possible stress-induced failures at an early point of treatment. As the spontaneous pregnancy rate is high in the first treatment cycles, LH-RH or its analogue should be administered after two frustrane insemination cycles.

Results of donor artificial insemination (AID) in 415 couples.

During a 4.5-year period 415 spouses of infertile males were treated by artificial insemination with donor semen, resulting in 210 pregnancies (51%). An average of 7.1 insemination cycles and 11.5 inseminations was required to achieve a pregnancy. Thirty-six percent of the pregnancies occurred in the first cycle of treatment, 71.5% after three cycles of treatment, and 91.5% in six cycles. The abortion rate was 15%, comparable to that in the fertile population. The sex ratio of deliveries at time of reporting was 60 males to 62 females. For selected couples suffering from male sterility, artificial insemination with donor semen provides a reliable and therapeutically effective alternative to adoption.

Purification of plasminogen activators from human seminal plasma.

Two plasminogen activators (1 and 2) were isolated from human seminal plasma by hiigh-speed centrifugation, Sephadex-gel filtration and ion-exchange chromatography. The activators were shown to be homogeneous by polyacrylamide-disc -gel electrophoresis at pH 8.3 and 4.5, and by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. The molecular weights of activators 1 and 2 were estimated as 69 000 and 74 000. Their amino acid compositions are very similar, both being high in aspartic acid, glutamic acid, serine, glycine and leucine, and low in methionine, tryptophan, tyrosine, isoleucine and histidine. Activators 1 and 2 each possess 16 cysteine residues. Both activators have isoelectric points of approx. 7.0, are stable over a wide pH range at temperatures up to 60 degrees C, but lose activity at higher temperatures, particularly under very basic or acidic conditions. They are not inhibited by EDTA, Mg2+ and Ca2+ at 10 mM concentrations, but their activity decreases on addition of 10 mM-cysteine or Fe2+ and 6-aminohexanoate or sera from pregnant women. The precipitin band formed between urokinase and its antiserum is continuous with the precipitin bands formed between the seminal plasminogen activators and the urokinase antiserum. Antisera to urokinase inhibit both the activity of urokinase and the seminal plasminogen activators.

An improved assay technique for the proteolytic activity of individual human spermatozoa.

An improved substrate-film technique has been developed for the assay of released proteinase from individual human spermatozoa. During the preparation of the thin gelatin membrane, it is pretreated with formaldehyde and NaOH. These agents alter the plasma membrane and the outer acrosomal membrane of the spermatozoon, facilitating the release of acrosomal enzymes. This effect is further enhanced by the addition of albumin to the incubation mixture. More than 90% of the spermatozoa in a normal ejaculate give a reaction by this method. It reaches a maximum after 4 hours of incubation and does not increase further even up to 12 hours. No difference in the reaction between washed and unwashed ejaculated spermatozoa can be found. The nature of the proteolytic activity and its possible significance in infertility are discussed.

Components of human split ejaculates. II. Enzymes and proteinase inhibitors.

Lysozyme, alpha-amylase, neutral proteinase and plasminogen activator were most concentrated in the initial portion of the ejaculate that consists mostly of Cowper's gland and prostate gland fluids as well as spermatozoa. The concentration of the high molecular weight proteinase inhibitors, alpha1-antitrypsin and alpha1X-antichymotrypsin, was essentially unaltered throughout the ejaculate fractions, although their absolute amounts showed an increase towards the final fraction. By contrast, the total inhibitory activity towards pancreatic trypsin was highest both in concentration and amount in the last fraction, thus indicating that the seminal vesicles are its primary source. Plasminogen, prothrombin, Factor XIII, and the proteinase inhibitors antithrombin III, alpha2-macroglobulin, inter-alpha-trypsin inhibitor and C1S-inactivator could not be detected immunochemically in whole ejaculates, and indicates the dissimilarity between the coagulation/liquefaction processes of semen and blood.

Scanning electron microscopy of cervical mucus crystallization.

The crystallization phenomenon of human cervical mucus was studied with the scanning electron microscope. Changes in the conformation of the crystals, crystal conglomerates, and ferning patterns occurred almost daily. During the very early and late stages of the cycle two types of crystals could be found, one columnar and the other flat. At the other times, the columnar crystals were absent and the flat crystals changed their shape to cuboidal. During the follicular phase, these cuboidal crystals were initially located in small groups, then in parallel rows, and finally in three types of ferning patterns, one seemingly typical for the time of ovulation. These patterns reversed during the luteal phase of the cycle. The scanning electron microscope appears to be an extremely useful instrument for the evaluation of these phenomena.