Use of metformin before and during assisted reproductive technology in non-obese young infertile women with polycystic ovary syndrome: a prospective, randomized, double-blind, multi-centre study.

BACKGROUND: To study the effect of metformin before and during assisted reproductive technology (ART) on the clinical pregnancy rate (CPR) in non-obese women with polycystic ovary syndrome (PCOS). METHODS: A multi-centre, prospective, randomized, double-blind study was conducted in eight IVF clinics in four Nordic countries. We enrolled 150 PCOS women with a body mass index <28 kg/m(2), and treated them with 2000 mg/day metformin or identical placebo tablets for ≥ 12 weeks prior to and during long protocol IVF or ICSI and until the day of pregnancy testing. The primary outcome measure was CPR. Secondary outcome measures included spontaneous pregnancy rates during the pretreatment period, and the live birth rate (LBR). RESULTS: Among IVF treated women (n = 112), biochemical pregnancy rates were identical in both groups (42.9%), and there were no significant differences in the metformin versus the placebo group in CPR [39.3 versus 30.4%; 95% confidence interval (CI): -8.6 to 26.5]. The LBR was 37.5 versus 28.6% (95% CI: -8.4 to 26.3). However, prior to IVF there were 15 (20.3%) spontaneous pregnancies in the metformin group and eight (10.7%) in the placebo group (95% CI: -1.9 to 21.1; P = 0.1047). According to intention to treat analyses (n = 149); significantly higher overall CPR were observed in the metformin versus placebo group (50.0 versus 33.3%; 95% CI: -1.1 to 32.3; P = 0.0391). LBR was also significantly higher with use of metformin versus placebo (48.6 versus 32.0; 95% CI: 1.1 to 32.2; P = 0.0383). No major unexpected safety issues or multiple births were reported. More gastrointestinal side effects occurred in the metformin group (41 versus 12%; 95% CI: 0.15 to 0.42; P < 0.001). CONCLUSIONS: Metformin treatment for 12 weeks before and during IVF or ICSI in non-obese women with PCOS significantly increases pregnancy and LBRs compared with placebo. However, there was no effect on the outcome of ART per se. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT00159575.

Fertility and ageing.

The late 20th century trend to delay birth of the first child until the age at which female fecundity or reproductive capacity is lower has increased the incidence of age-related infertility. The trend and its consequences have also stimulated interest in the possible factors in the female and the male that may contribute to the decline in fecundity with age; in the means that exist to predict fecundity; and in the consequences for pregnancy and childbirth. In the female, the number of oocytes decreases with age until the menopause. Oocyte quality also diminishes, due in part to increased aneuploidy because of factors such as changes in spindle integrity. Although older male age affects the likelihood of conception, abnormalities in sperm chromosomes and in some components of the semen analysis are less important than the frequency of intercourse. Age is as accurate as any other predictor of conception with assisted reproductive technology. The decline in fecundity becomes clinically relevant when women reach their mid-30s, when even assisted reproduction treatment cannot compensate for the decline in fecundity associated with delaying attempts at conceiving. Pregnancies among women aged >40 years are associated with more non-severe complications, more premature births, more congenital malformations and more interventions at birth.

[Embryonic stem cells and therapeutic cloning]. / Embryonale stamceller og terapeutisk kloning.

Increased interest in the therapeutic use of human stem cells has emerged following significant progress in ongoing research. The cloning of a sheep, the isolation of human embryonic stem cells, and the discovery that adult stem cells may be reprogrammed taken together give substance to hopes that novel principles of treatment may be developed for a variety of serious conditions. Embryonic stem cells are derived from pre-embryos at the blastocyst stage and may give rise to all bodily tissues and cells. Animal models have demonstrated that embryonic stem cells when transplanted into adult hosts may differentiate and develop into cells and tissues applicable for treatment of a variety of conditions, including Parkinson's disease, multiple sclerosis, spinal injuries, cardiac stroke and cancer. Transplanted embryonic stem cells are exposed to immune reactions similar to those acting on organ transplants, hence immunosuppression of the recipient is generally required. It is, however, possible to obtain embryonic stem cells that are genetically identical to the patient's own cells by means of therapeutic cloning techniques. The nucleus from a somatic cell is transferred into an egg after removal of the egg's own genetic material. Under specific condition the egg will use genetic information from the somatic cell in organising the formation of a blastocyst which in turn generates embryonic stem cells. These cells have a genetic composition identical to that of the patient and are suitable for stem cell therapy.

Detection of cytokines (interleukin-1, interleukin-6, transforming growth factor-beta) and soluble tumour necrosis factor receptors in embryo culture fluids during in-vitro fertilization.

Several lines of evidence suggest that a number of immunoactive cytokines participate in early reproductive events such as implantation and placental development. Furthermore, cytokines may influence embryo growth and differentiation. In the present study, the production of tumour necrosis factor (TNF), interleukin-1 (IL1), interleukin-6 (IL6) and transforming growth factor-beta (TGF beta) during the first 48 h after oocyte retrieval during in-vitro fertilization was investigated. In addition, the question was raised whether soluble receptors may contribute to cytokine activity regulation in early reproduction, and concentrations of TNF and IL6 receptors in culture media were determined. Finally, an investigation of whether any association exists between cytokine concentrations and embryo morphology was performed. Media from 256 embryos were analysed. IL1, IL6 and TGF beta were produced during the 48 h culture period, whereas no TNF was detected. Levels of IL1 and IL6 were significantly higher in media from the first 24 h culture period than from the second period, whereas TGF beta concentrations in supernatants from the two observation periods did not differ. IL6 receptors were not detected, whereas TNF receptors (p75) appeared in media from the 24-48 h culture period. Granulosa, cumulus and sperm cells are potential sources of cytokine production, especially during the first 24 h period. The contribution of the embryo to cytokine/cytokine receptor production remains an open question. No significant correlation was observed between cytokine/cytokine receptor concentrations and embryo morphological score.

Androgen metabolism in the different lobes of the prostate gland of intact, gonadectomized or hypophysectomized rats with or without androgen substitution.

The effect of gonadectomy or hypophysectomy and the effect of substitution with testosterone, upon the reductive and oxidative metabolic transformations of testosterone, 4-androstene-3,17-dione, 17 beta-hydroxy-5 alpha-androstane-3-one, 5 alpha-androstane-3 alpha, 17 beta-diol and 5 alpha-androstane-3 beta, 17 beta-diol using NAD(H) and NADP(H) as added cofactors were examined in homogenates from the ventral (VP), lateral (LP) and dorsal prostate (DP), and coagulating gland (CG) of adult Wistar rats. The fall in serum testosterone induced by gonadectomy or hypophysectomy led to reduced activity of 5 alpha-reductase, NADP(H)-dependent 3 alpha-hydroxysteroid oxidoreductase (3 alpha-HSOR), NAD-dependent 3 alpha-HSOR, 3 beta-HSOR and 17 beta-HSOR, indicating that these enzymes are androgen dependent. The NADP dependent oxidation of 5 alpha-androstane-3 alpha, 17 beta-diol in LP increased upon hypophysectomy and gonadectomy. Testosterone substitution of gonadectomized or hypophysectomized rats generally maintained enzymatic activity at the level of the control group, except for the 5 alpha-reductase activity which fell in spite of this treatment. Hypophysectomy or gonadectomy had different effects on the activity of several androgen metabolizing enzymes; 5 alpha-reductase activity in DP and CG, NAD(H) dependent 3 alpha-HSOR in VP and NADPH dependent 3 beta-HSOR activity in LP, DP and CG. There were marked differences between the rat prostatic lobes, both concerning the activity of the androgen metabolizing enzymes and the responses to the different treatment modalities.

High concentrations of the soluble p55 tumour necrosis factor receptor in human seminal plasma.

Soluble TNFRs (tumour necrosis factor receptors) inhibit in-vivo and in-vitro bioactivities of TNF, and thus the secretion of soluble TNFRs could be a physiological principle to attenuate the bioactivities of TNF. Two types of TNFR have been identified and both forms can be released from cells. In this study, soluble TNFRs in seminal plasma from three groups of men were analysed: from men with normal semen quality (n = 32), with reduced semen quality (n = 7) and vasectomized men (n = 3). Sensitive and specific enzyme-linked immunosorbent assays were utilized to detect soluble TNFRs in seminal plasma, based on capture antibodies directed against non-TNF-binding sites of the TNFRs and digoxigenin (DIG)-labelled TNF. The mean +/- standard deviation levels of p55 TNFR were 56.4 +/- 20, 64.4 +/- 17 and 45.4 +/- 5 ng/ml in the three groups, respectively. The concentration of p75 TNFR was < 1 ng/ml in all groups. The results suggest an exclusive existence of high amounts of the soluble p55 TNFR in seminal plasma. Seminal plasma from vasectomized men contained p55 TNFR at approximately the same levels as the specimen from the two other groups, indicating that the source of p55 TNFR is not the testis but rather some tissue more distal in the male genital tract, such as the prostate or the seminal vesicles. The soluble p55 TNFR was purified from human seminal plasma, using affinity and gel filtration chromatography. Further characterization of the purified p55 revealed a protein with a molecular weight of approximately 22 kDa, both under reducing and non-reducing conditions.(ABSTRACT TRUNCATED AT 250 WORDS)

Formation of antisperm antibodies in women treated with fallopian tube sperm perfusion.

Fallopian tube sperm perfusion (FSP) is a combination of ovarian stimulation and intra-uterine insemination using a large volume (4 ml) of inseminate containing 10(7)-10(8) spermatozoa. The inseminate will flush the Fallopian tubes and some of it will end up in the pouch of Douglas. In the present study, we have investigated whether the FSP method will result in the formation of serum antisperm antibodies in the female. A total of 184 treatment cycles were given to 128 women. The indications for treatment were: unexplained infertility (n = 35), various infertility diagnoses (n = 28) and donor insemination (n = 65). Prior to treatment, 11 (8.6%) women had a positive tray-agglutination test (Friberg) and/or a positive immunobead test. After completing one to four treatment cycles, another six (4.7%) women had developed serum antisperm antibodies. The antibodies induced by the treatment were of isotype IgM and directed against the tail-tip of the spermatozoa. Two of the women, who prior to the treatment had antisperm antibodies, showed an increase in antibody titre during treatment. There was no statistically significant difference in the pregnancy rate between the women with antisperm antibodies and the women without. In our opinion, the small risk of developing antisperm antibodies is no contra-indication for treating infertile couples with FSP.

Fallopian tube sperm perfusion (FSP) versus intra-uterine insemination (IUI) in the treatment of unexplained infertility: a prospective randomized study.

Prospective randomization of 60 couples with unexplained infertility was performed for treatment either with intrauterine insemination (IUI), using a volume of 0.5 ml of the inseminate, or Fallopian tube sperm perfusion (FSP), using a volume of 4 ml of inseminate. The protocols for ovarian stimulation and induction of ovulation were the same in the two groups. The two groups were similar concerning age of the female at the start of treatment and the number of follicles > 15 mm diameter, the serum oestradiol concentrations and the endometrial thickness on the day of human chorionic gonadotrophin (HCG) administration. The mean (+/- SD) number of motile spermatozoa inseminated was significantly higher in the FSP group than in the IUI group (52 +/- 5 x 10(6) and 28 +/- 3 x 10(6) respectively). In the FSP group, 30 women were given a total of 52 treatment cycles; 14 clinical pregnancies occurred in this group, giving a pregnancy rate of 26.9% per cycle and 46.7% per woman. In the IUI group, 28 women were given a total of 51 treatment cycles; five clinical pregnancies occurred, giving a pregnancy rate of 9.8% per cycle and 17.9% per woman. The pregnancy rates per cycle and per woman in the FSP group were significantly higher than in the IUI group (P < 0.05, chi-square test). This study indicates that in the treatment of couples with unexplained infertility, Fallopian tube sperm perfusion (FSP) is more successful than intra-uterine insemination (IUI).

Treatment of unexplained infertility. Fallopian tube sperm perfusion (FSP).

PATIENTS AND METHODS: Fifty-one couples with unexplained infertility were enrolled in the fallopian tube sperm perfusion (FSP) program. FSP is in short a combination of ovarian hyperstimulation, ovulation induction and intrauterine and intrafallopian tube insemination using a sperm suspension of 4 ml volume. RESULTS: One hundred cycles were started; 93 of these were completed, resulting in 27 pregnancies (29.0% per cycle). The pregnancy rate in the first treatment cycle (41.2%) was significantly higher than in the subsequent treatment cycles (14.3%, p < 0.01). Total pregnancy loss was 22.2% (2 ectopic pregnancies and 4 spontaneous abortions). The multiple birth rate was 14.2% (two sets of twins and one set of triplets). Twenty-two couples not conceiving following a maximum of three FSP treatment, were offered IVF treatment. Forty-three IVF cycles were started resulting in 27 embryo replacements in 15 patients. Seven pregnancies occurred in six couples. In 11 of the couples treated, IVF treatment revealed a possible explanation for the infertility; development of multiple small follicles following ovarian hyperstimulation, empty follicle syndrome and partial or total fertilization failure. Only in five of the couples who did not conceive, no cause for the infertility could be found. CONCLUSIONS: In the treatment of couples with unexplained infertility, the FSP procedure seems to give a birth rate of about 40% in less than two treatment cycles. FSP is less invasive and less expensive than alternative procedures such as GIFT, ZIFT and IVF and does not require the expertise and facilities for culturing of human oocytes and embryos.

[Fertilization in vitro--how large a proportion of couples will have a child?]. / In vitro-fertilisering--hvor stor andel av parene får barn?

In Norway, the efficacy and the efficiency of IVF treatment for infertile couples have been questioned. Information about this topic is important for the general public, the infertile population, the gynaecologists and the health authorities. We have followed a cohort of 485 infertile couples accepted for in vitro fertilization treatment. During a 14 month period starting 1 January 1989 a total of 214 women have either delivered or are pregnant in the second or third trimester. This number represents 44% of the 485 women accepted for IVF treatment, irrespective of whether they were treated or not, and 50% of the couples who completed at least one IVF cycle. There was no difference in cycle efficiency (pregnancy rate) between the first to third cycle, but this was significantly lower in cycle 4-6. There was no difference in delivery rate between groups of patients with different diagnoses of female infertility, but an additional male factor reduced the delivery rate significantly. Replacement of cryopreserved/thawed embryos gave a 5% increase in the delivery rate per couple included in the study, and an increase of 13% in the number of deliveries.

The efficacy and efficiency of an in-vitro fertilization programme including embryo cryopreservation: a cohort study.

A cohort of 485 couples starting their first in-vitro fertilization (IVF) attempt between January, 1989 and February, 1991 inclusive, were followed until June 1, 1992. A total of 1086 treatment cycles were initiated (mean 2.2, range 1-6). Of these, 235 (21.8%) cycles were cancelled, giving a total of 851 embryo replacements (mean 1.7, range 1-5). After IVF treatment, 189 women have either delivered or have an ongoing pregnancy in the second or third trimester. This gives a baby take-home rate of 17.4% per started cycle and 22.2% per embryo replacement. For 91 (18.6%) of the couples, the treatment was abandoned prior to completion of the three scheduled IVF attempts and 57 (11.7%) of these had no completed IVF cycles. In the group of couples with reduced sperm quality, the delivery rate was significantly lower than that of the other groups. A total of 193 women had embryos cryopreserved in at least one IVF cycle; 124 of these women started a frozen embryo replacement cycle and 88 had at least one cycle with replacement of frozen/thawed embryos, resulting in 25 deliveries/ongoing pregnancies. Due to the Norwegian law on assisted procreation 65 (33.7%) of the women have had their frozen embryos thawed and discarded after 12 months of storage. The cryopreservation programme, with the limitations of the Norwegian law, gives a 5.2% increase in the baby take-home rate for women entering the IVF programme, an increase of 13.2% in the number of ongoing pregnancies/deliveries and an 11.6% increase in number of children/viable fetuses.(ABSTRACT TRUNCATED AT 250 WORDS)

Fallopian tube sperm perfusion used in a donor insemination programme.

Ninety-six couples were enrolled in a programme of Fallopian tube sperm perfusion (FSP). FSP is a method combining ovarian stimulation, ovulation induction and intra-uterine insemination with a 4 ml volume of sperm suspension. Frozen/thawed donor semen was used in all cycles. A total of 196 treatment cycles were started and of these 172 were completed. Twenty cycles were cancelled because of maturation of too many follicles, a low ovarian response, or formation of ovarian cysts. Five of the women enrolled in the programme did not have any inseminations. Forty-eight pregnancies occurred (27.9% per cycle) among 45 women (49.5%). There were five spontaneous abortions and one ectopic pregnancy. Sixteen pregnancies are on-going (greater than 12 weeks) and 26 women have delivered (including two sets of triplets and one set of twins). The pregnancy rate declined from the first attempt (34.1%) to the fourth attempt (14.3%). The pregnancy rate was significantly higher in women with three to five mature follicles compared to women with two or fewer mature follicles (P less than 0.05). Cycles in which human chorionic gonadotrophin (HCG) was administered on day 11 or later had a higher pregnancy rate than cycles where HCG was administered on day 10 (P less than 0.01). There was no difference in pregnancy rates between insemination with 8-10 x 10(6) motile spermatozoa and insemination with greater than 30 x 10(6) motile spermatozoa. FSP used in a programme of artificial insemination by donor using frozen/thawed donor semen is relatively simple to perform and gives a high pregnancy rate per cycle.

Fallopian tube sperm perfusion: first clinical experience.

We have developed an insemination method using a large (4 ml) volume of the inseminate. This method incorporates ovarian stimulation, isolation of an optimal number of motile spermatozoa and, finally, Fallopian tube sperm perfusion (FSP). In a clinical study, 139 couples with various causes of infertility, had a total of 239 treatment cycles. The pregnancy rate per treatment in groups with endometriosis, ovulation disorders, tubal impairment, combined male and female factors and subnormal sperm quality were low, ranging from 2.7% to 7.7%. In patients with unexplained infertility, the pregnancy rate per treatment was 26.9% and for this group, the pregnancy rate in the first treatment cycle was 37.3%. In the cervical hostility group, two pregnancies occurred after five treatments. FSP seems to be a favourable treatment for couples with cervical hostility or unexplained infertility. About half of the women in these groups conceived after three treatment cycles. FSP is easier to perform and is less expensive than other methods of assisted procreation, such as in vitro fertilization and gamete intra-Fallopian transfer.

Dose-dependent induction of ornithine decarboxylase and S-adenosyl-methionine decarboxylase activity by testosterone in the accessory sex organs of male rats.

The dose-dependent induction of ornithine decarboxylase (ODC) and adenosylmethionine decarboxylase (AMDC) activity in the different lobes of the prostate and the seminal vesicles (SV), 24 hours after administration of testosterone to castrated Wistar rats, has been studied. ODC and AMDC activities were low in all lobes 10 days after castration. A dose of approximately 300 micrograms testosterone/100 g body weight (B.W.) gave an ODC activity of 50 percent of maximum response, and at 600 micrograms/100 g B.W. maximum activity was reached in all the prostatic lobes and the SV. In the lateral and dorsal prostate, and the coagulating gland, the dose of testosterone giving 50% of maximum AMDC activity was reached after administration of between 450 and 600 micrograms/100 g B.W. In the ventral prostate and SV, the dose giving a 50% response was approximately 700 micrograms/100 g B.W. In conclusion, all prostatic lobes showed a clear dose-response relationship concerning the activity of ODC and AMDC following administration of different doses of testosterone. We have found minor differences in androgen responsiveness between the lobes when looking at the dose requirements for induction of AMDC activity. The dose-response curves could possibly be useful as a rapid in vivo bioassay for compounds with anti-androgenic properties in the prostate.

IVF in the Nordic countries 1981-1987: a collaborative survey.

This report concerns the development of in-vitro fertilization (IVF) in the Nordic countries from the start of 1981 to January 1, 1988. All the 24 clinics in the Nordic countries which treat infertile couples with IVF participated in the study. The number of IVF clinics in the Nordic countries has increased from four in 1981 to 24 in 1987. In the same period, the number of people who spent some of their professional time as a part of an IVF team increased from 17 to 150. The majority of the clinics (19) based their activity mainly on governmental funding, while five of the clinics were private. During the period from 1981 to 1987, the Nordic clinic changed their methods of oocyte retrieval. In 1981, all oocyte retrievals were done using laparoscopy. Transvesical, ultrasound-guided follicular punctures domainated in 1985 and transvaginal ultrasound-guided punctures were used in 81.5% of the retrievals in 1987. A total of 852 pregnancies were reported. Of these, 313 (36.8%) were miscarriages, including 66 (7.7%) ectopic pregnancies. At January 1, 1988, 433 babies had been born, including 48 sets of twins, 14 sets of triplets and two sets of quadruplets. This gave a multiple birth rate of 18.2%. Premature births (less than 36 weeks of gestation) were reported in 14.6% of the births. Five babies (1.1%) died in the neonatal period and five (1.1%) were born with various types of malformation. One hundred and eight-eight pregnancies were in progress at January 1, 1988.

Androgen regulation of polyamine synthesis in seminal vesicle and in different lobes of the rat prostate.

Endogenous levels of the polyamines putrescine, spermidine, and spermine have been examined in the 10(6) m/s2 supernatant of different lobes of the rat prostate and in the seminal vesicles of castrates, androgen-stimulated castrates, and intact controls. Content of the polyamines varied between the lobes, with spermidine highest in intact animals. After castration, the content of polyamines fell significantly in all lobes but the coagulating gland (CG). Spermidine levels were highest, except in the lateral prostate (LP) and CG, where the content of putrescine was highest. In castrated animals treated with testosterone propionate for 72 h, the amount of the three polyamines examined increased dramatically in the ventral prostate (VP) and moderately in the CG and seminal vesicle (SV). Concerning individual polyamines, spermidine increased significantly in all lobes, while putrescine increased significantly only in the two saccular parts of rat prostate, i.e., CG and SV. Spermidine content decreased significantly in the DP. Major differences in the content of the three polyamines--putrescine, spermidine, and spermine--in the various tissues studied have been found. Moreover, distinct differences among intact, castrated, and testosterone-treated castrated animals have been revealed.

[Freezing, thawing and transfer of human embryos]. / Nedfrysing, tining og tilbakesetting av humane embryoer.

We present our experience of cryopreservation of human embryos from 1 July 1987 to 1 July 1989. During this period, 667 in vitro fertilizing cycles were completed. In 173 (26%) of these cycles, a total of 805 surplus embryos were cryopreserved. 516 embryos were thawed and 247 of these were transferred to the uterine cavity in a total of 110 replacement cycles. This resulted in 22 (20%) pregnancies in which a total of 28 gestational sacs could be verified by ultrasound. Seven of the pregnancies aborted spontaneously in the first trimester and one pregnancy was ectopic. By 1 July 1989, five singleton pregnancies were in progress and 13 babies had been born from nine deliveries (one set of triplets and two sets of twins). 25% of the patients whose embryos were cryopreserved became pregnant in the in vitro fertilizing cycle after replacement of fresh embryos.

Separation of urinary metabolites of radiolabelled estrogens in man by HPLC.

A method to separate radiolabelled urinary estrogens by high performance liquid chromatography (HPLC) is described. Estrogen glucuronides were isolated from the urine of women receiving bolus injections of [4-14C]estrone or [4-14C]estradiol by adsorption on Sep-Pak C18 cartridges and subsequent DEAE Sephadex A25 column chromatography. Following enzyme hydrolysis, free estrogens were extracted and concentrated in methanol-water containing ascorbic acid. HPLC was performed either by C18 reversed phase chromatography using different concentrations of acetonitrile with or without tetrahydrofurane in phosphate buffer or methanol-water as mobile phases, or on a Diol column using chloroform-isooctane-n-hexane or isopropanol-isooctane-n-hexane as mobile phases. 3H-labelled estrogens were added as internal standards, and urinary [14C]estriol, [14C]estradiol and [14C]estrone concentrations could be measured with an interassay coefficient of variation less than 5%. Interassay coefficients of variation for [14C]2-hydroxyestriol, [14C]16 alpha-hydroxyestrone, [14C]2-hydroxyestradiol, [14C]2-hydroxyestrone and [14C]2-methoxyestrone were between 5 and 10%, while interassay coefficients of variation for [14C]4-hydroxyestrone was 14.6%. Recovery of the unstable catechol estrogen 2-hydroxyestrone was comparable to the recovery of the other estrogen metabolites, due to the addition of ascorbic acid throughout the different pre-chromatographic steps. Our method is suitable for the separation of the major labelled estrogen metabolites found in human urine following administration of radiolabelled estrone or estradiol.

A spleen pregnancy.

Non-tubal ectopic pregnancy is very rare. We report a case of spleen pregnancy, difficult to recognize clinically. The tests and investigations which led to the diagnosis and treatment are described briefly.