Semen quality of young men in Switzerland: a nationwide cross-sectional population-based study.

BACKGROUND: Sperm counts have been steadily decreasing over the past five decades with regional differences in the Western world. The reasons behind these trends are complex, but numerous insights indicate that environmental and lifestyle factors are important players. OBJECTIVE: To evaluate semen quality and male reproductive health in Switzerland. MATERIALS AND METHODS: A nationwide cross-sectional study was conducted on 2523 young men coming from all regions of Switzerland, recruited during military conscription. Semen volume, sperm concentration, motility, and morphology were analyzed. Anatomy of the genital area and testicular volume was recorded. Testicular cancer incidence rates in the general population were retrieved from Swiss regional registries. RESULTS: Median sperm concentration adjusted for period of sexual abstinence was 48 million/mL. Comparing with the 5th percentile of the WHO reference values for fertile men, 17% of men had sperm concentration below 15 million/mL, 25% had less than 40% motile spermatozoa, and 43% had less than 4% normal forms. Disparities in semen quality among geographic regions, urbanization rates, and linguistic areas were limited. A larger proportion of men with poor semen quality had been exposed in utero to maternal smoking. Furthermore, testicular cancer incidence rates in the Swiss general population increased significantly between 1980 and 2014. DISCUSSION: For the first time, a systematic sampling among young men has confirmed that semen quality is affected on a national level. The median sperm concentration measured is among the lowest observed in Europe. No specific geographical differences could be identified. Further studies are needed to determine to what extent the fertility of Swiss men is compromised and to evaluate the impact of environmental and lifestyle factors. CONCLUSION: A significant proportion of Swiss young men display suboptimal semen quality with only 38% having sperm concentration, motility, and morphology values that met WHO semen reference criteria.

LIF and sIL-2R plasma concentrations in IVF patients on the day of embryo transfer: predictive markers of IVF outcome.

Successful implantation is still the limiting step in IVF. We hypothesized that maternal plasma concentrations of certain cytokines at the time of embryo transfer could predict the likelihood of successful implantation and pregnancy. sIL-2R, IL-6, LIF, and MMP2 concentrations were measured in plasma from 160 IVF patients (natural and stimulated IVF cycles) on the morning of the embryo transfer (ET0) and 14 days later (ET+14). Patients were ultimately subdivided into four groups depending on the IVF treatment outcome (pregnancy failure, biochemical pregnancy, first-trimester miscarriage and normal term delivery). In natural and stimulated IVF cycles at ET0, sIL-2R concentrations were threefold higher in biochemical pregnancies than in pregnancy failures (P=0.020), and in natural cycles only, 2.5-fold higher in normal term deliveries than in pregnancy failures (P=0.023). Conversely, in natural and stimulated IVF cycles at ET0, LIF concentrations were one third lower in biochemical pregnancies/first-trimester miscarriages compared with pregnancy failures (P=0.042). We suggest that high sIL-2R and low LIF concentrations in maternal plasma on the morning of the embryo transfer might be associated with increased risks of early pregnancy loss, while a basal level of sIL-2R is necessary for normal term delivery outcome. Both cytokine measurements might therefore be useful in the management of IVF patients, and modulation of their concentrations could be investigated as a therapeutic alternative for women with abnormal concentrations at the time of embryo transfer.

A European multicentre prospective randomized study to assess the use of assisted hatching with a diode laser and the benefit of an immunosuppressive/antibiotic treatment in different patient populations.

BACKGROUND: Assisted hatching (AH) techniques, designed for facilitating the embryo escape out of the zona pellucida (ZP) have been used in IVF centres since 1992. The initial indications for AH were patient's age, ZP thickness, high basal FSH and repeated IVF failures. Several retrospective and prospective studies assessing AH in these indications have given disparate results. Our aims were to evaluate the benefits of AH and immunosuppressive/antibiotic treatment (IA) in patients with either a poor prognosis of success, previous implantation failures or transfers of cryopreserved embryos. METHODS: Four IVF centres allocated 426 patients, randomized for AH and IA, into four groups of AH indications between 1997 and 1999. AH was performed with a diode laser. ZP thickness, opening size and embryo score were recorded. Outcome measures were implantation and delivery rates. RESULTS: Patients coming for a first or third transfer of cryopreserved embryos and poor prognosis patients admitted for a first trial did not benefit from AH. Even patients with repeated implantation failures of fresh embryos did not gain significantly from AH. CONCLUSIONS: Among AH indications, absence of implantation after several transfers of good quality embryos remains the strongest patient selection criterion. Prescription of an immunosuppressive/antibiotic treatment is essential.

Involvement of connexin 43 in meiotic maturation of bovine oocytes.

In ovarian follicles, cumulus cells provide the oocyte with small molecules that permit growth and control maturation. These nutrients reach the germinal cell through gap junction channels, which are present between the cumulus cells and the oocyte, and between the cumulus cells. In this study the involvement of intercellular communication mediated by gap junction channels on oocyte maturation of in vitro cultured bovine cumulus-oocyte complexes (COCs) was investigated. The stages of oocyte maturation were determined by Hoechst 33342 staining, which showed that 90% of COCs placed in the maturation medium for 24 h progress to the metaphase II stage. Bovine COC gap junction communication was disrupted initially using n-alkanols, which inhibit any passage through gap junctions. In the presence of 1-heptanol (3 mmol l(-1)) or octanol (3.0 mmol l(-1) and 0.3 mmol l(-1)), only 29% of the COCs reached metaphase II. Removal of the uncoupling agent was associated with restoration of oocyte maturation, indicating that treatment with n-alkanols was neither cytotoxic nor irreversible. Concentrations of connexin 43 (Cx43), the major gap junction protein expressed in the COCs, were decreased specifically using a recombinant adenovirus expressing the antisense Cx43 cDNA (Ad-asCx43). The efficacy of adenoviral infection was > 95% in cumulus cells evaluated after infection with recombinant adenoviruses expressing the green fluorescence protein. RT-PCR performed on total RNA isolated from Ad-asCx43-infected COCs showed that the rat Cx43 cDNA was transcribed. Western blot analysis revealed a three-fold decrease in Cx43 expression in COCs expressing the antisense RNA for Cx43. Injection of cumulus cells with Lucifer yellow demonstrated further that the resulting lower amount of Cx43 in infected COCs is associated with a two-fold decrease in the extent of coupling between cumulus cells. In addition, oocyte maturation was decreased by 50% in the infected COC cultures. These results indicate that Cx43-mediated communication between cumulus cells plays a crucial role in maturation of bovine oocytes.

A low-dose stimulation protocol using highly purified follicle-stimulating hormone can lead to high pregnancy rates in in vitro fertilization patients with polycystic ovaries who are at risk of a high ovarian response to gonadotropins.

OBJECTIVE: To study the benefits of a low-dose stimulation (LDS) protocol with purified urinary follicle-stimulating hormone in patients with polycystic ovaries who have presented previously with a very high ovarian response to a standard hMG stimulation. DESIGN: Cohort study. SETTING: Fertility center in a university hospital. PATIENT(S): Sixty-one patients involved in an IVF/ICSI program from January 1995 to December 1996. INTERVENTION(S): The patients were first stimulated with a standard protocol using hMG and presented with a very high ovarian response. These patients were then stimulated a second time using a low-dose protocol. Cryopreserved embryos were transferred in later artificial or natural cycles until to December 1999. MAIN OUTCOME MEASURE(S): Number of gonadotropin ampules; estradiol level on the day of ovulation induction; follicles, oocytes, and cryopreserved zygotes; fertilization, implantation, and pregnancy rates; and number of ovarian hyperstimulation syndromes (OHSS). RESULT(S): The number of ampules used, the estradiol level reached, and the number of oocytes obtained were significantly lower under the LDS than the standard protocol. High implantation (21.8%) and clinical pregnancy (38.4%) rates were obtained after LDS. The cumulated deliveries per cycle started and per patient were, respectively, 41.6% and 52.5%. Five patients suffered OHSS with the standard protocol, and none with the LDS. CONCLUSION(S): The LDS protocol offers a safe and efficient treatment for patients who present with echographic polycystic ovaries and are at risk of an excessive ovarian response to standard IVF stimulation protocols.

Molar pregnancy with a coexistent fetus after intracytoplasmic sperm injection. A case report.

BACKGROUND: Normal fertilization is usually considered to have occurred when two pronculei (2PN) and two polar bodies are observed. Exceptions are the single pronucleated zygote resulting from asynchronous pronuclei. CASE: A 29-year-old woman entered a program of intracytoplasmic sperm injection and embryo transfer because of her husband's oligoasthenoteratozoospermia. Two cleavage-stage embryos (four blastomeres, grade 1 and 2) were obtained from one fertilized oocyte containing distinct 2PN and the other a single pronucleus (1PN). At 15 weeks' gestation the patient developed severe preeclampsia requiring termination of the pregnancy. Histopathologic examination and DNA ploidy by image analysis were consistent with a twin pregnancy combining a complete hydatidiform mole and normal pregnancy. CONCLUSION: We hypothesize that this 1PN was at the origin of the hydatidiform mole. This case highlights the danger of transferring an embryo having 1PN.

[Assisted fertilization in azoospermic men: results of CHUV from 1994 to 1997]. / Fécondation assistée chez les hommes azoospermiques: résultats du CHUV de 1994 à 1997.

In vitro fertilisation (IVF) and intracytoplasmic sperm injection (ICSI) have revolutionised the treatment and prognosis of oligo-terato-asthenozoospermia (OTA). Sperm extraction in the vas deferens, the epididymis (MESA: with epididymal sperm aspiration) or the testicles (TESE: with testicular sperm extraction), associated with ICSI, can achieve pregnancy in cases of excretory or secretory azoospermia. We report the results of the use of MESA and TESE in 42 patients with an average age of 37 (age range 24 to 58). Of these, 26 have excretory azoospermia, 11 secretory azoospermia and 5 a problem linked to ejaculation. Of the 506 oocytes that were inseminated, 270 zygotes were obtained, giving a fertilisation rate of 53.4%. 85 embryo transfers were carried out (55 with fresh embryos and 30 with cryo-preserved embryos). Three spontaneous abortions and one extrauterine pregnancy were reported. Six pregnancies are developing normally. To date, 13 children have been born (9 boys and 4 girls) in 10 deliveries (7 single children and 3 sets of twins). The limits of male infertility need to be revised to take these new forms of therapy into account and patients should be advised on the new possibilities available.

Follow-up of 32 hypothalamo-hypopituitary patients treated with pulsatile gonadotropin-releasing hormone or human menopausal gonadotropin.

In a clinical retrospective study, a follow-up of hypothalamo-amenorrheic patients treated firstly with gonadotropin-releasing hormone (GnRH) pump stimulation and secondly with human menopausal gonadotropin (hMG) was performed. Thirty-two hypothalamo-amenorrheic patients, 24-38 years old, were submitted to 103 GnRH stimulation cycles. Seven, with polycystic ovaries (PCO) on ultrasound, were stimulated with hMG after one or several unsuccessful pump cycles. Ovulation was confirmed by a luteinizing hormone (LH) surge or triggered by human chorionic gonadotropin in 80 out of 103 cycles (77.7%/cycle) leading to 62 timed sexual intercourses and 17 intrauterine inseminations (IUI). Twenty-one pregnancies (26.3%/cycle) terminated in eight abortions (38.1%/pregnancy) and 13 deliveries (40.6%/patient). hMG stimulation, in the seven PCO patients (six IVF, one IUI), led to four additional deliveries in three patients. Five patients became pregnant spontaneously after pump failure (n = 2) or unsuccessful IVF (n = 3). Combining all cycles, 17 deliveries were obtained in 16 patients. No case of ovarian hyperstimulation syndrome (OHSS) was observed. GnRH is an efficient and safe treatment of hypothalamo-amenorrheic-induced anovulation. Following GnRH or hMG ovarian stimulation, spontaneous ovulation and conception may be restored in certain hypothalamo-amenorrheic patients.

Aorto-subclavian thromboembolism: a rare complication associated with moderate ovarian hyperstimulation syndrome.

The case of an arterial aorto-subclavian thromboembolism associated with a moderate ovarian hyperstimulation syndrome (OHSS) and following ovulation induction for in-vitro fertilization in a young woman is reported. Because of the lack of response to systemic thrombolysis, a left postero-lateral thoracotomy was performed on day 8 after embryo transfer. A fibrinocruoric embolus situated at the junction of the left subclavian artery from the aorta was removed through a left subclavian arteriotomy. The distal axillary embolus was removed by a retrograde balloon catheter embolectomy. A moderate OHSS was observed. The ovarian stimulation and OHSS-related risks of thromboembolism are discussed. We conclude that, in the absence of risk factors, counselling about possible complications resulting from stimulation must be emphasized.

Normogonadotropic primary amenorrhea in a growth hormone-deficient woman with ectopic posterior pituitary: gonadotropin pulsatility and follicle-stimulating hormone bioactivity.

We studied the gonadotropic function in a 25-year-old woman suffering from congenital GH deficiency, complaining of primary amenorrhea and wishing to become pregnant. She disclosed a hypoplasic anterior pituitary within a small sella turcica and an ectopic posterior pituitary lobe located below the median eminence. Immunoreactive LH and FSH plasma levels were normal, basal and in response to a GnRH iv bolus but estradiol was low. LH pulse frequency was elevated and FSH bioactivity was low in a granulosa cell aromatase bioassay. Pulsatile administration of iv GnRH at a slower, normal pace, failed to induce ovulation or to increase FSH bioactivity, with or without concomitant GH replacement. However treatment with exogenous im gonadotropins, when preceeded by GH replacement, succeeded in inducing mature oocytes and pregnancy. We concluded that the hypogonadism observed in this patient was due to rapid GnRH pulsatility and poor biological activity of endogenous FSH.

Non-contact microdrilling of mouse zona pellucida with an objective-delivered 1.48-microns diode laser.

BACKGROUND AND OBJECTIVE: A non-touch laser-induced microdrilling procedure is studied on mouse zona pellucida (ZP). STUDY DESIGN/MATERIALS AND METHODS: A 1.48-microns diode laser beam is focused in a 8-microns spot through a 45x objective of an inverted microscope. Mouse zygotes, suspended in a culture medium, are microdrilled by exposing their ZP to a short laser irradiation and allowed to develop in vitro. RESULTS: Various sharp-edged holes can be generated in the ZP with a single laser irradiation. Sizes can be varied by changing irradiation time (3-100 ms) or laser power (22-55 mW). Drilled zygotes present no signs of thermal damage under light and scanning electron microscopy and develop as expected in vitro, except for a distinct eight-shaped hatching behavior. CONCLUSION: The microdrilling procedure can generate standardized holes in mouse ZP, without any visible side effects. The hole formation can be explained by a local photothermolysis of the protein matrix.

Expression of serum albumin and of alphafetoprotein in murine normal and neoplastic primitive embryonic structures.

Alphafetoprotein (AFP), a major serum protein synthesized during the embryo-fetal and postnatal period (in the yolk sac, then in the liver), is also an oncoprotein. The intracellular presence of AFP and of serum albumin (SA) in normal and neoplastic neural crest and neural tube derivatives was previously demonstrated. In this work we have studied the comparative expression of AFP and SA in primitive neuroectoblastic structures of mouse embryos (6 and 7 days "post coitum") and mouse teratocarcinomas (derived from the PCC4 cell line). Using immunofluorescence technique, antibodies to SA gave a positive reaction in embryos of 7 days, while AFP was not detected during this period. By mRNA in situ hybridization, SA mRNA gave a strong signal in both 6 and 7 day embryos, whereas AFP mRNA gave a weak signal only in 7-day embryos. The distribution of SA and AFP and their mRNAs was investigated in primitive neuroectoblastic structures of the teratocarcinomas by in situ hybridization and immunostaining. Only SA protein was detectable by immunostaining. SA mRNA gave a strong signal in differentiating structures as well as in undifferentiated cell clusters. AFP mRNA was observed only in differentiating structure. Dot-blot hybridization indicated that the level of SA transcripts was at least 6-fold higher than that of AFP transcripts in the teratocarcinomas investigated. In teratocarcinoma-bearing mice injected intraperitoneally with 125I-radiolabeled SA and AFP, significant accumulations of both SA and AFP were demonstrated in the tumors, SA being about 3-fold higher than that of AFP after normalization to quantity of uptake in liver. External in vivo photoscanning confirmed this relationship of accumulated radiolabeled proteins. The last observation could be useful in vivo for diagnosis of teratocarcinoma. We conclude that the expression of SA relative to AFP and the external cellular uptake of SA relative to AFP are similar in normal embryonic developing tissues and in the corresponding morphologically neoplastic tissues of the teratocarcinomas. The same SA:AFP relationship constitutes an oncofetal marker of primitive neuroectoblastic structures.

Timing of hCG administration in cycles stimulated for in vitro fertilization: specific impact of heterogeneous follicle sizes and steroid concentrations in plasma and follicle fluid on decision procedures.

The timing of ovulation induction is usually decided according to estradiol plasma concentrations and follicle size. We administered human chorionic gonadotropin (hCG) when at least three follicles of 16 mm or more in diameter and adequate estradiol plasma concentrations were detected. We studied the percentage of mature oocyte-cumulus-corona radiata complexes, estradiol and progesterone concentrations in a heterogeneous sized follicle population (range 10-20 mm, n = 90) to perform a retrospective analysis of the adequacy of criteria adopted for the timing of ovulation induction. Plasma and follicular fluid were obtained from 20 normo-ovulating women (aged 28-37 years) treated with gonadotropin releasing hormone analogs (GnRH-a) and human menopausal gonadotropin (hMG) for in vitro fertilization (IVF). No correlation was found between the mean individual follicular fluid estradiol concentration (500-5640 nmol/l) and the respective maximum concentration in plasma (2-16 nmol/l). The estradiol concentration was similar in all follicles. Total follicular fluid estradiol concentration was found to be correlated with follicular fluid volume (r = 0.771, p < 0.01). On the day of hCG administration, the concentration of estradiol in the plasma but not the follicular fluid was correlated with the number of oocyte-cumulus-corona radiata complexes collected (p < 0.01) and the number of mature complexes (p < 0.01). At oocyte pick-up, the plasma concentration of progesterone was correlated (p < 0.01) with number of complexes collected and the number of mature complexes. The percentage of mature complexes collected (77.5%) was higher than suggested by the number of leading follicles. This indicates that our criteria for administering hCG were adequate and that heterogeneous follicle size does not exclude a high rate of mature oocyte-cumulus-corona radiata complexes.

[Midazolam versus placebo before spinal anesthesia]. / Midazolam versus placebo avant rachianesthésie.

The goal of this randomized, double blind and multicentric study was to compare the effects of midazolam (M) and placebo (P) administered by titration before puncture for spinal anaesthesia on the comfort of 211 patients scheduled for elective surgery after oral premedication with hydroxyzine. The administered dose of midazolam was 3.4 +/- 1.3 mg (mean +/- SD). Anxiety was nil in 92% of the patients of the M group and in 64% of the patients of the P group (p < 0.001) and memorization of the pain of the puncture was reported in 34% of the patients of the M group and in 66% of the patients of the P group (p < 0.001). However cooperation of the patient and easiness of the puncture were similar in both groups. In conclusion titrated sedation with midazolam before puncture for spinal anaesthesia increases the comfort of the patient.

[Ovulation induction by pulsatile intravenous administration of Gn-RH to patients with hypothalamic amenorrhea]. / Induzione dell'ovulazione con somministrazione pulsatile intravenosa di Gn-RH in pazienti con amenorrea ipotalamica.

Intravenous pump administration (Zyklomat, Ferring) of GnRH (Lutrelef, Ferring) has been used to induce ovulation in 16 patients suffering from hypothalamic amenorrhea between January 1989 and December 1991. One to 6 cycles were performed for a total of 47 cycles. GnRH pulses were set at 20 micrograms/pulse every 90 min up during the follicular phase and decreased to 1 pulse/120 min during the 3-5 days following ovulation. Thereafter, the luteal phase was supported by 1000 IU HCG/48 h up to the end of the 5th week. Response to the treatment was monitored with seriC E2, P and LH and vaginal echography. Cycles (N = 3) were cancelled when E2 did not reach at least 0.25 nmol/l after 30 days. An E2 level > 0.8 nmol/l, associated with a mature follicle (> 20 mm), and the presence of a LH peak or an increased progesterone level were used to determine the day of ovulation. In case of normozoospermy (N = 9), sexual intercourse was recommended to the patients (27 cycles), whereas an intrauterine insemination was performed in case of a male factor (N = 7, 20 cycles). Results can be summarized as follows: 39 (83%) cycles were ovulatory and led to 9 (19%) clinical and 6 (13%) biochemical pregnancies. Five (8.5%) pregnancies aborted and 5 (11%) came to full term. After exclusion of the male factors, the pregnancy rate reaches 26%/initiated cycle, 28%/ovulatory cycle and 56%/patient. No patient developed a hyperstimulation syndrome or a multiple pregnancy. Three cycles (6.3%) were interrupted due to 2 superficial phlebitis and 1 pump failure.(ABSTRACT TRUNCATED AT 250 WORDS)

[A traditional anesthesia consultation or a "telephone interview" within the framework of ambulatory surgery?]. / Consultation d'anesthésie traditionnelle ou "interview téléphonique" dans le cadre de la chirurgie ambulatoire?

UNLABELLED: Preoperative assessment for anaesthesia must detect in the setting of ambulatory surgery both medical and sociopsychologic problems which may contraindicate the ambulatory aspect of the procedure. The goal of our prospective study was to assess the efficacy of a preoperative telephone interview in screening the medical and sociopsychologic problems of ambulatory patients. 1,000 consenting consecutive outpatients participated to the study and were randomly allocated to one of two groups: patients in the standard group (STD) had a consultation a few days before surgery whereas those in the telephonic group (TEL) called the anesthetist a few days before, the physical exam being performed on the day of surgery. The cancellation of the ambulatory procedure was decided upon either medical criterias (MC) or sociopsychologic criterias (from Waetchler). RESULTS: both groups were comparable regarding SPC and MC but prescription of preoperative tests was more frequent in the STD group (result statistically significant). In conclusion, this new approach is as reliable as the standard consultation in screening the SPC which remain the most frequent cause of cancellation in ambulatory surgery. Therefore this telephonic interview which is simple and reliable for screening outpatients may represent a suitable alternative to a standard consultation.

Immunofluorescence study of actin, acrosin, dynein, tubulin and hyaluronidase and their impact on in-vitro fertilization.

Using polyclonal antibodies, the distribution of actin, acrosin, dynein, tubulin and hyaluronidase has been examined by indirect immunofluorescence in sperm preparations from fertile donors and in-vitro fertilization (IVF) patients. After recording sperm parameters in native semen, spermatozoa were washed free of seminal plasma using either the swim-up or the Percoll filtration technique. Prior to insemination, aliquots of the washed sperm suspensions were prepared for antibody staining. Spermatozoa from fertile donors were analysed in order to establish the specific fluorescence patterns of each antibody and the threshold scores of normality. Immunofluorescence scores obtained from IVF patients were then analysed with respect to IVF outcome. For each tested protein, the number of normal samples were significantly lower in the group which did not fertilize and fertilization rates were significantly reduced when any of the tested proteins were scored as pathological. Normal fluorescence scores were correlated with morphology, motility, velocity and to a lesser extent with sperm concentration in native semen. On the basis of receiver-operating characteristic curves, likelihood ratios and Cohen's kappa values, the presence of acrosin and tubulin yields the most useful information on sperm functional and structural status and on its fertilizing ability.

Follicle-stimulating hormone bioactivity in idiopathic normogonadotropic oligoasthenozoospermia: double-blind trial with gonadotropin-releasing hormone.

OBJECTIVE: To identify, among patients with idiopathic normogonadotropic oligoasthenozoospermia, those with low bioactive follicle-stimulating hormone (FSH), possibly because of inadequate gonadotropin-releasing hormone (GnRH) pulsatility, whose bioactive FSH and sperm could be improved by GnRH treatment. DESIGN: Randomized, double-blind, placebo-controlled trial with intranasal (IN) GnRH, followed by open GnRH treatment. SETTING: Outpatient endocrinology clinic. PATIENTS: Twenty-eight infertile men with idiopathic normogonadotropic oligoasthenozoospermia. INTERVENTIONS: Gonadotropin-releasing hormone or placebo was self-administered IN every 2 hours. MAIN OUTCOME MEASURES: Serum immunoreactive and bioactive FSH and semen analyses. RESULTS: Ten men showed a low basal FSH bioactive/immunoreactive ratio, which increased in 5 of them under GnRH without parallel sperm modification. Sperm improvements were observed in 10 patients with no parallel evolution of FSH bioactive/immunoreactive ratio. Unpredicted by sperm changes, three pregnancies developed on placebo and 5 on GnRH. CONCLUSIONS: Low bioactive FSH was not the cause of idiopathic normogonadotropic oligoasthenozoospermia in our patients and could not predict response to GnRH. Pulsatile GnRH did not improve sperm beyond random fluctuations.

Endogenous and exogenous alpha-fetoproteins as differential markers of cultured neonatal mouse Schwann cells and fibroblasts.

alpha-Fetoprotein (AFP) and AFP-gene transcripts were demonstrated in vitro in Schwann (S) cell and fibroblast (F) cell cultures of neonatal mouse origin. All S and F cells of primary cultures and of established cell lines expressed the AFP gene. AFP mRNA was detected by an in situ hybridization technique using a 35S-AFP-cDNA probe. AFP was localized by immunocytoperoxidase labelling using purified anti-AFP antibodies. The amounts of stained endogenous AFP, estimated semiquantitatively, were about 3-fold higher in S cells than in F cells. After incubating the cultures with exogenous mouse AFP, both S and F cells showed significant ability to take up the protein; the amount of internalized protein was found to be higher in F cells than in S cells. Moreover, the uptake of AFP fluorescein conjugates (FITC-AFP), estimated quantitatively by fluorometry, also gave higher values for F cells. The cytoplasm of F cells exhibited a characteristic fluorescence pattern, strongly illuminated and dispersed grains; the cytoplasm of S cells was regularly labelled. If exogenous FITC-AFP uptake could be used to distinguish labelled F cells from S cells (with application for identification and selection of F cells), the immunocytochemically stained endogenous AFP could allow S cells to be distinguished from F cells (using the dilutions of antibodies still staining the S cells but which lead to the absence of F-cell labelling). The two procedures, which can be used independently or together, may constitute differential markers for S cell and F cultures in, i.e., nerve regeneration of neurofibroma studies using the model of mixed S and F culture also containing other types of cells.

Why lose an in vitro fertilization cycle when stimulation fails?

Different protocols using agonists of GnRH-a have been proposed for the ovarian stimulation in IVF cycles. In case of stimulation failure with the flare-up protocol, we have investigated whether an immediate switch to pituitary blockade by the long-acting analog may avoid the cycle to be canceled. This procedure allows a rescue of cycles among poor responders and does not have any deleterious effect on the recruitment of follicles, oocyte quality, fertilization, and pregnancy rates.