Are follicle stimulating hormone measurements predictive of ovarian response to hyperstimulation with human menopausal gonadotrophin?

Previous studies have suggested that elevated serum follicle stimulating hormone (FSH) concentrations are associated with a poor ovarian response to hyperstimulation with human menopausal gonadotrophin (HMG) in in vitro fertilisation (IVF) programmes. We have used the day 2 serum FSH concentration to determine the dose of HMG administered in women under 40 years. If the FSH concentration was below 9 IU/l, a constant dose of 150 IU HMG were administered; if above 9 IU/l a constant dose of 300 IU HMG was used. Women over the age of 40 years were given 300 IU HMG regardless of their serum FSH concentration. This retrospective study was undertaken to assess whether this approach was beneficial for the younger women and also whether the FSH concentration was predictive of outcome in older women. The study included all women < 40 years (n = 143) and > 40 years (n = 32) having their first IVF treatment cycle during 1994. In the younger women, there was no difference in the number of cancelled treatment cycles (9.7% vs. 7.5%); the number of follicles present (9.6 vs. 8.2); serum oestradiol concentration (6971 pmol/l vs. 6686 pmol/l); number of eggs collected (7.9 vs. 5.7); number of embryos created (3.7 vs. 3.6); and pregnancy rate (13.5% vs. 15%) between women with normal (n = 103) or elevated (n =40) FSH concentrations. By using the serum FSH concentration to select women in whom a poor response was expected, and administering a higher dose of HMG, a similar ovarian response was produced and the pregnancy rate was similar to those in women with normal FSH concentrations. Women over 40 years with elevated serum FSH concentrations (n = 17) had a significantly (P < 0.05) higher cancellation rate (17.6% vs. 0%) and fewer number of eggs collected (6.9 vs. 2.5) than the group with normal FSH concentrations (n = 15). One woman conceived in each group. These findings confirmed previous studies showing that the serum FSH is predictive of ovarian response. This study confirmed the value of measuring the day 2 serum FSH concentration as a predictor of response; and it provides a scientific approach to determine the dose of HMG administered for IVF stimulation. A satisfactory response to induction of ovulation will be achieved using 150 IU HMG in women with FSH < 9 IU/l but if the FSH is raised i.e. above 9 IU/l, 300 IU is required to achieve a similar response.

Use of buserelin and low-dose human menopausal gonadotropin for in vitro fertilization in women at risk of ovarian hyperstimulation syndrome.

PURPOSE: The purpose of the present study was (i) to assess the value of using a low dose of hMG (75 IU/day) to achieve ovarian stimulation in women who have previously shown an exaggerated response to a standard dose of 150 IU human menopausal gonadotropin/day in a desensitization (group I) or flare-up (group II) protocol and (ii) to determine whether the choice of GnRH-a regimen in a subsequent cycle, namely, a desensitization or flare-up protocol, influenced the effectiveness of the low dose of hMG. RESULTS: In group I, 75% (12/16) and 57% (8/14) of the subsequent desensitization and flare-up protocols, respectively, were cancelled because of inadequate ovarian response. Similarly, the cancellation rates in group II were 10 of 10 and 7 of 11 (64%), respectively. The total cancellation rate (groups I and II together) with the desensitization protocol was higher than that using the flare-up protocol (P < 0.05). CONCLUSION: The simple use of a reduced dose of hMG (75 IU/day) for subsequent in vitro fertilization in women to minimize the risk of the development of ovarian hyperstimulation is of limited benefit since a large proportion then shows an inadequate response. This is particularly pronounced with a subsequent desensitization protocol which does not utilize endogenous gonadotropins to initiate follicular development.

The routine assessment of sperm motility at room temperature and 37 degrees C.

The World Health Organization (WHO) laboratory manual (1992) states that assessment of sperm motility can be performed at either 37 degrees C or room temperature (20-24 degrees C). The motility of spermatozoa in 44 semen samples (22 fresh samples and 22 frozen-thawed samples) was assessed at both of these temperatures and a significant difference in the motility profiles was noted, specifically an increase at 37 degrees C in the percentage (expressed here as median and ranges) of spermatozoa with excellent progressive motility and an overall increase in the percentage with total progressive motility. With fresh samples the excellent progressive motility increased from 41 (19-53) to 54 (30-66) and the overall motility from 58.5 (39-74) to 65.0 (40-79). With the frozen-thawed samples the excellent motility increased from 14 (1-33) to 25 (6-45) and the overall motility from 30.5 (14-51) to 33.0 (16-52). As the WHO laboratory manual was published 'In response to a growing need for the standardisation of procedures for the examination of human spermatozoa' it is proposed that only one temperature for routine analysis should be used, namely 37 degrees C, which may have more physiological relevance and eliminate effects of fluctuations in ambient laboratory temperature.

The acrosome reaction to ionophore challenge test: assay reproducibility, effect of sexual abstinence and results for fertile men.

Since the human acrosome reaction is considered a prerequisite for normal fertilization and the spontaneous acrosome reaction rate is low, laboratory tests using calcium ionophores to induce the acrosome reaction have been devised and applied to the investigation of patients. The introduction of any new laboratory test into routine clinical practice is usually accompanied by the determination of intra- and inter-subject variability within the normal population, and the derivation of reference values to distinguish between affected and unaffected populations. The acrosome reaction to ionophore challenge (ARIC) test was evaluated and found to have (i) intra- and inter-assay coefficients of variation of 10.8 and 18.8% respectively, (ii) a high degree of intra-subject variability for three subjects studied over a 10 week period, (iii) a high degree of inter-subject variability when aliquots of 20 ejaculates of donor semen of proven fertility were tested, and (iv) no effect of length of sexual abstinence on ARIC values. The results of this study suggest that the use of fresh semen samples from subjects of proven fertility for quality control purposes in the ARIC test may be inadequate due to the high degree of intra-subject variability, and that this problem may be overcome by utilizing a frozen quality control sample. The results also suggest that an isolated negative ARIC test is not necessarily indicative of functionally incompetent spermatozoa, and highlight the importance of examination of the normal population prior to the clinical application of such a test.

Assisted conception using buserelin and human menopausal gonadotrophins in women with polycystic ovary syndrome.

OBJECTIVE: To compare the outcome of in vitro fertilisation (IVF) and gamete intrafallopian transfer (GIFT) cycles in women with or without ultrasound features of polycystic ovary syndrome (PCOS). DESIGN: A consecutive series from January to December 1989. SUBJECTS: Twenty-five women with PCOS scheduled for assisted conception. The controls were 139 women with normal ovaries. SETTING: A single centre specialist fertility unit, Manchester, UK. INTERVENTIONS: Pituitary desensitisation was with buserelin. In the PCOS group ovarian stimulation was with 1 ampoule (75 iu FSH) of hMG/day in 12 women (Group I) and two ampoules/day in 13 (Group II). The controls (Group III) were given two ampoules of hMG daily. Human chorionic gonadotrophin (hCG; 10,000 iu) was given when three follicles measured > or = 20 mm diameter. MAIN OUTCOME MEASURES: Serum oestradiol (E2) concentrations, number of follicles, clinical pregnancies, features of the ovarian hyperstimulation syndrome (OHS). RESULTS: Women with PCOS (Groups I or II) had more follicles > or = 14 mm diameter on the day of the hCG injection (P < 0.005), higher serum E2 concentrations on the day after the hCG (P < 0.05) and more oocytes retrieved (P < 0.05) than the controls. The OHS was more prevalent in those with PCOS (32% versus 6.5%; P < 0.05). The clinical pregnancy rate per embryo transfer (27% versus 22%) or gamete transfer (25% versus 39%) and the rate of spontaneous miscarriage (33% versus 12%) were not statistically different. CONCLUSIONS: The pregnancy rate and outcome of pregnancy following IVF or GIFT in women with or without PCOS are similar. Women with PCOS are at a higher risk of developing OHS.

Does elective cryopreservation of all embryos from women at risk of ovarian hyperstimulation syndrome reduce the incidence of the condition?

OBJECTIVES: To analyse the incidence and factors associated with the ovarian hyperstimulation syndrome (OHS) in our IVF/GIFT programme before and after the introduction of a strategy to cryopreserve all embryos from women judged to be at risk. DESIGN: Two hundred forty-one consecutive IVF/GIFT cycles from January to December 1989. SETTING: Specialist fertility unit, Manchester, UK. INTERVENTIONS: Pituitary suppression was effected by a daily subcutaneous injection of buserelin (500 micrograms) beginning 7 days before the expected menses. The ovarian stimulation was with variable amounts of human menopausal gonadotrophin. Ovulation was induced with 10,000 i.u. human chorionic gonadotrophin (hCG). From January to May (period A), gametes/embryos were replaced and 2000 i.u. hCG given, irrespective of the serum oestradiol (E2) concentration. From June to December (period B), all the embryos from women with an E2 > 3500 pg/ml on the day of ovulatory trigger were electively cryopreserved. MAIN OUTCOME MEASURES: Serum E2, features of moderate or severe OHS, clinical pregnancies. RESULTS: The OHS occurred in 10/105 (9.5%) and 12/136 (8.8%) cycles in periods A and B, respectively. Fewer women (6% versus 60%, P < 0.05) who had their embryos cryopreserved developed severe OHS compared with women with an E2 > 3500 pg/ml who became pregnant after gamete/embryo transfer in period A. The main factors associated with the development of OHS were serum E2 concentrations > 3500 pg/ml, whether gamete/embryos were replaced and the additional hCG given, the occurrence of a pregnancy and the presence of polycystic ovary disease. CONCLUSION: The elective cryopreservation of all embryos from women with high E2 levels reduced the severity, but not the incidence of symptomatic OHS.

Outcome of treatment subsequent to the elective cryopreservation of all embryos from women at risk of the ovarian hyperstimulation syndrome.

From 1st June 1989 to 31st May 1991, 78 women with a serum oestradiol level greater than 3500 pg/ml on the day of the ovulatory trigger, following pituitary suppression with buserelin and ovarian stimulation with human menopausal gonadotrophins (HMG), had all their embryos electively cryopreserved at the pronucleate stage to minimize the risk of developing ovarian hyperstimulation syndrome (OHS). Treatment with buserelin was continued in the luteal phase. A median of 19 oocytes (range 7-43) was obtained and 12 embryos (range 1-37) frozen per cycle. Twenty-one (27%) women developed OHS (six severe). Women developing OHS had higher (P less than 0.05) serum oestradiol concentrations on the 7th day after oocyte retrieval, compared to those who did not. No differences were found for any of the following criteria: aetiology of infertility, age, total dose of HMG, number of oocytes, fertilization rate or freeze-thaw survival of embryos. Subsequently, 125 frozen-thawed embryo replacements have been undertaken, using buserelin and hormone replacement therapy (HRT) (n = 93) or natural cycles (n = 32). The overall freeze-thaw survival and implantation rates per embryo were 71.8 and 11.7%, respectively. The pregnancy rates in natural cycles (19%) and buserelin/HRT cycles (29%) were not significantly different.

Cryopreservation of human embryos at the pronucleate, early cleavage, or expanded blastocyst stages.

Supernumerary embryos following treatment by IVF or GIFT were cryopreserved at the pronucleate, early cleavage or expanded blastocyst stages. The success of embryo cryopreservation at these stages was evaluated in terms of (i) the proportion of embryos surviving the freeze/thaw procedure; (ii) the proportion of patients reaching embryo replacement; and (iii) the incidence of pregnancy per replacement. Significantly more embryos survived when frozen/thawed at the pronucleate (44/61; 72%) or early cleavage stages (48/80; 60%), than at the expanded blastocyst stage (13/34; 38%). A significantly higher proportion of patients had embryo replacements when embryos were frozen/thawed at the pronucleate (17/19; 89%) or early cleavage stages (21/24; 88%), than at the expanded blastocyst stage (9/17; 53%). Following replacement of frozen/thawed pronucleate and early cleavage stage embryos, clinical pregnancy rates of 8/17 (47%) and 3/21 (14%) clinical pregnancies were achieved, respectively. No pregnancies were achieved following replacement of frozen/thawed expanded blastocysts.

Fertilization in vitro of supernumerary oocytes following gamete intra-fallopian transfer (GIFT).

Gamete intra-Fallopian transfer (GIFT) was performed in 130 treatment cycles over a 17-month period. In 91% (118/130) of the cycles one or more oocytes were available for insemination in vitro and only GIFT cycles with supernumerary oocytes were included in the present study. Pituitary and ovarian suppression was achieved with buserelin followed by stimulation of multifollicular development by human menopausal gonadotrophin (HMG). Failure of supernumerary oocytes to fertilize was associated with a significantly reduced pregnancy rate (3/23; 13%) compared to cycles where fertilization occurred in vitro (35/95; 37%). These findings demonstrate that the outcome of IVF of supernumerary oocytes may be of particular diagnostic value in couples where the female partner has not conceived following treatment by GIFT after pituitary down-regulation with buserelin and ovarian stimulation with HMG.

The cryopreservation of donor semen by a simplified method: use in an IVF and GIFT programme.

The cryopreservation of semen used in assisted reproduction procedures was carried out exclusively by a simplified method in which a mixture of semen and cryoprotectant was contained in 1-ml tuberculin syringes and plunged directly into liquid nitrogen. Donor semen samples halved and frozen in syringes and in straws in a controlled-rate freezer showed no significant difference in post-thaw motility (P = 0.217) or survival (P = 0.217) after 30 min. However, after 180 min the survival rate showed a significant reduction in syringes (P = 0.045). A significant difference (P less than 0.00008) in the rate of fertilization of oocytes was seen in IVF cycles using frozen-thawed donor sperm (58/142, 42%) when compared to fresh sperm from husbands (2315/3926, 59%). A significant reduction (P less than 0.00005) in fertilization rate was also observed in the case of supernumerary oocytes in GIFT cycles with the cryopreserved donor sperm (29/132, 22%) compared to the husbands' sperm (239/514, 46%). However, the pregnancy rate following IVF and embryo replacement was the same after fertilization with fresh sperm (75/351, 21%) as opposed to frozen sperm (3/14, 21%). Furthermore, a higher pregnancy rate was observed in GIFT with frozen donor sperm (9/19, 47%) than with fresh sperm from husbands (28/103, 27%), though this was not statistically significant (P = 0.079). These results show this simplified methods of semen cryopreservation to be effective when used in an IVF and GIFT programme, giving pregnancy rates comparable to fresh normospermic semen samples. The method is simple, quick and inexpensive.

Measurement of human chorionic gonadotropin during early pregnancy: a comparison of two immunoradiometric assays.

Evidence of implantation following either in vitro fertilization and embryo transfer (IVF-ET) or gamete intrafallopian transfer (GIFT) was obtained by collecting blood on days 10, 12, 14, 16, and 18 after oocyte recovery (day 0), and retrospectively measuring human chorionic gonadotropin (hCG). This was done using immunoradiometric assays for hCG, manufactured either by Serono Diagnostics Ltd. (MAIAclone) or Diagnostics Products Corporation (IRMA-count). The analysis of 63 serum samples by both kits showed a good correlation (r = 0.99) but the Serono (y) method gave values which were consistently greater (y = 1.58x + 4.89) than those of the DPC (x) method. A comparison of the hCG profile of singleton pregnancies measured with either the Serono (n = 33) or DPC (n = 22) kits gave a similar relationship. These results suggest that great care must be taken when comparing results from different laboratories using different kits. Also, consideration must be given to the possible loss of a continuous database should a laboratory change kit.

Quality control in an in-vitro fertilization laboratory: use of human sperm survival studies.

A bioassay procedure is described for quality control testing of various disposable items used in routine IVF procedures. This bioassay is performed over 4 days and uses the survival of human sperm in vitro at room temperature to assess which products are suitable for use. New products were tested for cytotoxicity using a general screening method and subsequent batches of every suitable item tested to detect interbatch variation. Products were considered suitable or unsuitable for use depending upon a calculated sperm survival index. Two main types of product were found to be cytotoxic, namely certain brands of syringe and surgical gloves, the common feature of both being the presence of rubber components. The bioassay was also used to investigate further the cytotoxic effect of the powdered and starch-free surgical gloves. The cytotoxic substances from both types of surgical glove were readily transferred to an embryo replacement catheter by touch, and washing of the gloves reduced this effect only moderately. The bioassay has proved inexpensive and convenient but more importantly it has been invaluable for detecting potential sources of cytotoxicity before they are introduced into a standard IVF protocol.

Fertilization of human oocytes in vitro by spermatozoa from oligozoospermic and normospermic men.

Sperm were isolated from the semen of oligozoospermic (less than 20 x 10(6) sperm/ml) and normospermic (greater than or equal to 20 x 10(6) sperm/ml) men in an in-vitro fertilization (IVF) programme. Oocytes from the female partners were inseminated with either 75 or 100 x 10(3) motile sperm and checked for fertilization after 16-20 h. A significant reduction in the overall fertilization rate of oocytes was seen for the oligozoospermic group compared to the normospermic group, at both insemination concentrations. In the oligozoospermic group, a fertilization rate of 31% (19/61) was achieved when oocytes were inseminated with 75 x 10(3) sperm, and 38% (9/24) when inseminated with 100 x 10(3) sperm. This compared with rates of 57% (397/696) and 64% (650/1018), respectively, for normospermic cases at both insemination concentrations. No evidence of fertilization was seen in 36% (4/11) and 67% (4/6) of oligozoospermic cases when 75 or 100 x 10(3) sperm were used, compared with values of 13% (17/133) and 9% (20/212), respectively, in normospermic cases. After excluding zero cases, the fertilization rate of oocytes for the oligozoospermic group (75%; 9/12) was similar to the normospermic group (70% 650/935) when 100 x 10(3) sperm were used. However, when 75 x 10(3) sperm were used, the fertilization rate for the oligozoospermic group (41%; 19/46) was significantly lower than that of the normospermic group (62%; 397/645). Following the transfer of embryos into the female partner, clinical pregnancies were diagnosed in 2/7 (29%) oligozoospermic cases and 27/267 (10%) normospermic cases.(ABSTRACT TRUNCATED AT 250 WORDS)