Supplementary growth hormone treatment of women with poor ovarian response to exogenous gonadotrophins: changes in serum and follicular fluid insulin-like growth factor-1 (IGF-1) and IGF binding protein-3 (IGFBP-3)

The effects of supplementary growth hormone (GH) treatment uponinsulin-like growth factor-1 (IGF-1), IGF binding protein-3(IGFBP-3) concentrations in serum and ovarian follicular fluidwere investigated in women undergoing buserelin human menopausalgonadotrophin (HMG) ovulation induction for in-vitro fertilization.Women (n = 40), aged 24–39 (mean 35 years), who showedpoor ovarian responses to HMG, were recruited and randomly dividedinto two groups. Each patient received two cycles of ovulationinduction, one with GH (12 IU/day x 12 days/HMG/buserelin) andanother with placebo/HMG. Serum IGF-1 increased substantiallyduring the GH treatment and remained significantly higher thanthe control 2 days after the last GH injection. Serum IGFBP-3fell steadily during the placebo/HMG treatment and to a nadiron the day of oocyte retrieval (P <0.05 compared to serumbefore any treatment). In contrast, IGFBP-3 was increased (P<0.01) during the GH administration and returned to the controllevel 2 days after GH injection. Serum oestradiol concentrationson the eighth day of HMG and the day of human chorionic gonadotrophin(HCG) were not significantly different between the two groups.Serum IGF-1 was highly correlated with IGFBP-3 before any treatment(r = 0.433, P < 0.001). This correlation disappeared afterbuserelin, placebo/HMG treatment in the control group, but itwas maintained during GH/HMG treatment (r = 0.343, P = 0.04).Follicular fluid concentrations of GH and IGF-1, not IGFBP-3or oestradiol, were significantly elevated in the GH-treatedwomen. Serum IGF-1 on the day of oocyte retrieval was highlycorrelated to the follicular fluid IGF-1 in both groups. Therelationships between the follicular fluid GH and IGF-1 werecompletely opposite in the two groups, being positive in thecontrol group and negative in the GH-treated group. In the controlgroup, significant correlations were found between follicularfluid concentrations of IGF-1 and IGFBP-3, and GH and IGFBP-3which were not found in the GH-treated group. There were nocorrelations found between follicular fluid concentrations ofGH or IGF-1 or IGFBP-3 and oestradiol. The results clearly demonstratethat the normal GH, IGF-1, IGFBP-3 relationships can be alteredby treatments which influence the ovarian—pituitary axis;the significance of such changes to ovulation remains to bediscovered.     

High levels of immunoreactive endothelin-1 in human follicular fluids

Follicular fluids were obtained from 180 follicles of 15 womenundergoing follicular aspiration for in-vitro fertilization.Follicular development was induced by a combination of buserelinacetate and human menopausal gonadotrophin. Endothelin-1 (ET-1)concentrations in human follicular fluids were determined byspecific radioimmunoassay. ET-1-like immunoreactivity (ET-1-LI)ranged from 338 to 928 pg/ml. ET-1-LI concentrations in follicularfluids obtained from immature (< 15 mm) follicles were significantlyhigher than those from mature (15–25 mm) and post-mature(25 mm) follicles. No correlation was found between the concentrationof ET-1-LI, on the one hand, and that of oestradiol, progesterone,testosterone, prolactin, luteinizing hormone, insulin-like growthfactor-I, prostaglandin E₂ or platelet activating factor onthe other, in follicular fluids. However, a significant positivecorrelation was observed between ET-1-LI concentration and folliclestimulating hormone and IGF-II concentrations, respectively.These data suggest that the high concentration of ET-1 foundin follicular fluids may play some physiological role in folliculardevelopment.     

Endocrinology: Growth hormone, oestradiol, progesterone and testosterone concentrations in follicular fluid after ovarian stimulation with various regimes for assisted reproduction

Follicular fluid samples and oocytes were obtained from 75 women(87 cycles), who participated in an assisted conception programme.Determinations of the concentration of oestradiol, progesterone,testosterone and growth hormone were performed in all follicularfluid samples. Patients were stimulated with the following regimes:group A (24 cycles, 94 samples), human menopausal gonadotrophin(HMG) (three ampoules/day) and human chorionic gonadotrophin(HCG); group B (23 cycles, 53 samples), HMG/HCG with prednisolone(7.5 mg/day) after cycle programming with oral contraceptives;group C (40 cycles, 60 samples), buserelin with HMG/HCG. Oestradiolconcentrations (mean ± SEM) were significantly higher(P < 0.05) in group A (320.1 ± 27.3 ng/ ml) and thoseof growth hormone in both groups A and C (3.8 ± 0.2 and3.2 ± 0.15 ng/ml, respectively), as compared to the othergroups, whereas progesterone and testosterone concentrationswere similar in all groups. The mean concentrations of oestradiol,progesterone, testosterone and growth hormone were significantlyhigher (P < 0.01) in follicular fluid with oocytes of intermediatematurity than with mature oocytes (382.5 ng/ml, 7847.5 ng/ml,1704.5 ng/dl and 3.7 ng/ml versus 217.8 ng/ml, 5488.4 ng/ml,1313.6 ng/dl and 2.7 ng/ml, respectively). On the other hand,only oestradiol concentrations were significantly higher infollicular fluid of fertilized compared to non-fertilized oocytes.Concentrations of the other hormones analysed, except growthhormone, were similar in follicular fluid from pregnant andnon-pregnant women after assisted reproduction. Growth hormone,on the other hand, was significantly lower (P < 0.05) infollicular fluid from pregnant compared to non-pregnant women(2.8 versus 3.5 ng/ml). It is concluded that intermediate maturityoocytes and oocytes which will be subsequently fertilized arefound in follicles with higher follicular fluid concentrationsof growth hormone and steroids. Moreover, oocytes leading topregnancy after in-vitro fertilization and embryo transfer arederived from follicles with lower growth hormone concentrationsin follicular fluid.     

Gonadotrophin surge-attenuating factor bioactivity is present in follicular fluid from naturally cycling women

Rat pituitary monolayer bioassays were used to compare gonadotrophinsurge-attenuating factor (GnSAF) bioactivity in follicular fluidfrom 12 follicles in 10 spontaneously cycling women with thatin pooled follicular fluid from women undergoing ovulation induction.Expressed as ED₅₀s (µl follicular fluid/well producing50% of maximal effect), GnSAF bioactivity was detectable inall spontaneous follicular fluid samples (1.4–33.3 µl/well)and in follicular fluid from women undergoing ovulation induction(6.8 µl/well). This GnSAF bioactivity was unaffected bypre-incubation with an inhibin antibody. When the data weregrouped according to whether the recovered oocytes fertilizedin vitro or not, the fertilized group contained significantlygreater GnSAF bioactivity than the unfertilized group (5.3 ±1.1 and 14.1 ± 2.6 µl/well respectively, P <0.05). While both inhibin bioactivity (9.7 ± 1.4 and28.9 ± 12.1 µl/well) and immunoreactivity (36.8± 2.2 and 21.0 ± 3.0 and ng/ml) were also greater(P < 0.01) in the fertilized compared with the unfertilizedgroups respectively, there were no other significant differencesbetween the two groups. We conclude that GnSAF is found in follicularfluid from spontaneously cycling women, supporting in-vivo evidencefor the involvement of GnSAF in feedback control of the ovary-pituitaryaxis.     

Endocrinology: Presence and possible function of activin-like substance in human follicular fluid

We assessed the presence of an activin-like substance in humanfollicular fluid that was obtained from women undergoing in-vitrofertilization using a bioassay for activin A. Activin activitywas not detected in crude follicular fluids; the bioactivityof standard activin A was inhibited by the addition of follicularfluid. After the follistatin (binding protein of activin A)was removed from follicular fluid using a purification procedure,activin activity was detected in the follicular fluids (meanconcentration: 131 ± 40 ng/ml). Activin activity wasinhibited by the addition of follistatin to fluid. The concentrationof activin activity was substantially higher (100-fold) thanthat reported in serum. The concentration negatively and significantlycorrelated with the number of developed follicles in the ovary(r = 0.501, P < 0.01). These results suggest that activinA and its binding protein are present in follicular fluid inlarge amounts and that they may have a role in local ovarianregulation.     

Fertilization and early embryology: Human follicular fluid inhibits the binding of human spermatozoa to zona pellucidain vitro

The effect of human follicular fluid on human zona pellucidabinding of spermatozoa was investigated using the hemizona bindingassay (HZA). This effect was compared to that of progesterone,a known component of human follicular fluid. Exposure of spermatozoato 25% pooled human follicular fluid for 1 h significantly reducedthe number of spermatozoa bound to zona pellucida when comparedto those without human follicular fluid treatment (149.1 ±30.7 versus 177.1 ± 33.8, P 0.01). The same phenomenonwas observed after 3 h of treatment The corresponding numbersof bound spermatozoa were 140.4 ± 19.1 and 200.2 ±23.4 (P 0.0001). Progesterone (1.0µg/ml) stimulated thezona pellucida-binding capacity of spermatozoa significantlyunder the same conditions (P 0.01). The numbers of bound spermatozoaafter 1 and 3 h progesterone treatment were 235.5 ± 44.7(control, 168.1 ± 32.9) and 204.3 ± 27.4 (control,162.3 ± 20.1) respectively. HZA comparing the effectsof human follicular fluid and progesterone at concentrationsequivalent to those found in human follicular fluid using matchinghemizonae confirmed the inhibitory effect of human follicularfluid on sperm binding to zona pellucida (80.4 ± 28.4versus 149.8 ± 35.2, P 0.05). This inhibitory effectwas also found in another eight individual human follicularfluid samples. Both human follicular fluid and progesteronedid not affect the motility and viability of the treated spermatozoawhen compared to the controls with the same incubation period.Although more spermatozoa underwent the acrosome reaction after1 and 3 h of human follicular fluid treatment than in the control,the extent was comparable to those after progesterone treatmentThese results suggested that human follicular fluid inhibitedthe zona pellucida-binding capacity of spermatozoa in vitro.This inhibitory effect of human follicular fluid was not mediatedby progesterone, and did not result from the effects of humanfollicular fluid on sperm motility, viability and acrosome reaction.     

Follicular fluid pH changes following intraperitoneal exposure of Graafian follicles to carbon dioxide: a comparative study with follicles exposed to ultrasound

Follicular aspiration to obtain oocytes is generally performedvia laparoscopy after creating a pneumoperitoneum with carbondioxide. Such a procedure has been shown to reduce the rateof in-vitro fertilization of human oocytes and affect the rateof cleavage of rabbit embryos. These adverse effects may becaused by a reduction in follicular fluid pH due to diffusionof carbon dioxide into the folilde. In laparoscopic oocyte retrievals,a negative correlation was observed between duration of CO₂exposure and follicular fluid pH, whereas In ultrasound-guidedretrievals, the pH remaIned unchanged. The mean pH In 78 folliclesaspirated at laparoscopy was 7.22 ± 0.03 compared with7.62 ± 0.01 in 35 follicles aspirated under ultrasoundguidance (P = 0.0003). The results also indicate that oocytesin preovulatory follicles are surrounded by fluid that is morealkaline than plasma. Hence, the acidic environment treatedby CO₂ may be deleterious to sub sequent reproductive functionof the oocyte.     

Long-lasting coronary vasoconstrictor effects and myocardial uptake of endothelin-1 in humans

The effect of intravenous administration of the endothelium-derived vasoconstrictor peptide endothelin-1 (ET-1 0.2, 1 and 8 pmol kg^?1 min^?1) on coronary blood flow in relation to plasma ET-1 as well as blood lactate and glucose levels were investigated in six healthy volunteers. Coronary sinus blood flow was measured by thermodilution. Administration of ET-1 elevated arterial plasma ET 35-fold, dose-dependently increased mean arterial blood pressure from 95±5 mmHg to 110±6 mmHg (P<0.01) and reduced heart rate from 64±4 beats min^?1 to 58±4 beats min^?1 (P<0.05) at 8 pmol kg^?1 min^?1. Coronary sinus blood flow was reduced maximally by 23±4% (P<0.01) and coronary vascular resistance increased by 48±11% (P<0.01). Coronary sinus oxygen saturation decreased from 35±1% to 22±2% at 2 min after the infusion (P<0.01). A coronary constrictor response was observed at a 4-fold elevation in plasma ET. The reduction in coronary sinus blood flow lasted 20 min and coronary sinus oxygen saturation was still reduced 60 min after the infusion. Myocardial oxygen uptake or arterial oxygen saturation were not affected by ET-1. Myocardial lactate net uptake decreased by 40% whereas glucose uptake was unaffected. At the highest infusion rate there was a net removal of plasma ET by 24±3% over the myocardium (P<0.05). The results show that ET-1 induces long-lasting reduction in coronary sinus blood flow via a direct coronary vasoconstrictor effect in healthy humans observable at a 4-fold elevation in plasma ET-1. Furthermore, there is a net removal of circulating ET-1 by the myocardium.     

Immunology: Assessment of the relevance of zona pellucida antibodies in follicular fluid of in-vitro fertilization (IVF) patients

The presence of anti-zona pellucida antibodies in the follicularfluid of 11 women who underwent in-vitro fertilization (IVF)and embryo transfer was analysed. Only infertile couples withtubal or unexplained pathologies were included in our study,which was aimed at investigating the relationship between anti-zonapellucida antibodies in follicular fluid and failed fertilization.Whether or not these antibodies were present in some or allfollicles in the same patient was also investigated. Out of55 follicular fluids analysed, 36.3% were positive to the testand no fertilization was observed in oocytes from these follicles,while 63.6% were negative, and the oocyte fertilization rateassociated with these was 51.4%. The presence of anti-zona pellucidaantibodies was positively correlated with the degree of fertilizationfailure (P <0.001 x² test).     

The effects of the somatostatin analogue octreotide on ovulatory performance in women with polycystic ovaries

The elevated luteinizing hormone (LH) and androgen concentrationscharacteristic of women with polycystic ovaries (PCO) are consideredcrucial factors in their infertility. The somatostatin analogueoctreotide lowers LH and androgen concentrations in women withPCO. The effects of octreotide given concurrently with humanmenopausal gonadotrophin (HMG) were therefore compared withthat of HMG alone in 28 infertile women with PCO resistant toclomiphene. In 56 cycles of combined HMG and octreotide therapythere was more orderly follicular growth compared with the multiplefollicular development observed in 29 cycles in which HMG wasgiven alone (mean number of follicles > 15 mm diameter onthe day of human chorionic gonadotrophin (HCG) administration:2.5 ± 0.2 and 3.6 ± 0.4 respectively; P = 0.026).There was a significantly reduced number of cycles abandoned(>4 follicles > 15 mm diameter on day of HCG) in patientstreated with octreotide + HMG, so that HCG had to be withheldin only 5.4% of cycles compared to 24.1% with HMG alone (P <0.05). The incidence of hyperstimulation was also lower on combinedtreatment. Octreotide therapy resulted in a more ‘appropriate’hormonal milieu at the time of HCG injection, with lower LH,oestradiol, androstenedione and insulin concentrations. Althoughgrowth hormone concentration was similar on both regimens, significantlyhigher insulin growth factor-I concentrations were observedon the day of HCG in women on combined therapy than on HMG alone.     

Somatostatin in human follicular fluid

To demonstrate the presence of somatostatin in human pre-ovulatoryfollicular fluid, and to assess the role of this peptide infollicular maturation, a total of 66 follicular fluid sampleswere obtained from 26 patients at the time of oocyte recoveryfor in-vitro fertilization. Follicular fluid concentrationsof somatostatin, oestradiol, progesterone and androstenedionewere measured by immunoassays. Somatostatin concentrations inconcomitantly obtained plasma samples were also analysed. Follicularfluid somatostatin concentrations ranged from undetectable (<1.5pmol/1) to 109.4 pmol/1. The mean ±SE somatostatin concentrationsin follicular fluid (12.8± 1.8 pmol/1) were significantly(P< 0.0001) increased compared to corresponding plasma concentrationsof somatostatin (6.5 ± 0.2 pmol/1). A significant andpositive correlation existed between follicular fluid and plasmasomatostatin concentrations (r = 0.27; P < 0.03). No differencesin either follicular fluid or plasma somatostatin concentrationswere found between different stimulation protocols or diagnosticgroups. Neither did follicular fluid somatostatin concentrationvary with follicular size. Similarly, no differences in somatostatinconcentrations were found between follicular fluids associatedwith fertilized (13.2 ± 2.1 pmol/1) or non-fertilizedoocytes (10.5± 1.6 pmol/1). Follicular fluid concentrationsof somatostatin correlated positively with those of progesterone(r % 0.30; P = < 0.04), but not with those of oestradiolor androstenedione or with the androstenedione/oestradiol ratio.The relationship between follicular fluid somatostatin and progesteroneconcentrations suggests that follicular fluid somatostatin mayhave a physiological role in follicular maturation and the luteinizationprocess.     

Ovarian stimulation with buserelin/HMG/HCG: prospective randomized study of short versus long protocol

The combined administration of the gonadotrophin-releasing hormone(GnRH) agonist buserelin and human menopausal gonadotrophin(HMG) was evaluated in 527 cycles (428 patients) of an assistedreproduction programme. All women were randomly allocated accordingto the ovulation induction protocol into two groups: group I(short protocol; 318 cycles) was given buserelin (1 mg/day)intranasally from cycle day 1 and HMG (2 ampoules/day) fromday 3 until human chorionic gonadotrophin (HCG) administration:group H (long protocol; 209 cycles) was given buserelin (1 mg/day)intranasally from cycle day 1 for at least 14 days and then2 ampoules HMG/day were added, increasing progressively accordingto the ovarian response. The number (mean ± SEM) of folliclesdeveloped was higher in group II than in group I (9.1 ±0.4 versus 7.7 ± 0.3, respectively; P < 0.05). Moreoocytes were retrieved in group II (8.4 ± 0.5) than ingroup I (6.5 ± 0.3) (P < 0.001), as well as more embryos(6.3 ± 0.5 and 4.0 ± 0.3, respectively; P <0.001). Moreover, in group II there was a better correlationbetween oestradiol and the total follicular volume (r = 0.5391)on cycle day 0 compared with group I (r = 0.458), while oestradiolvalues were similar between the two groups. No differences wereobserved in the cancellation rate, fertilization rate and maturityof the oocytes between the two groups. The pregnancy rate pertransfer was slightly better in group II (25.8%) than in groupI (19.4%), but this difference was not significant. More stimulationdays were needed in group II than in group I (11.8 ±0.2 and 10 ± 0.2, respectively) (P < 0.001) and moreHMG ampoules (37.7 ± 1.4 and 27.9 ± 0.1, respectively)(P < 0.001). In conclusion, the administration of the longprotocol is associated with a higher number of follicles developed,oocytes retrieved and embryos obtained, while it seems morepromising concerning the pregnancy rates. Nevertheless, treatmentwith this protocol increases the stimulation days and the numberof HMG ampoules administered and hence the cost.     

A controlled study comparing patients with and without polycystic ovaries undergoing in-vitro fertilization

The outcome of in-vitro fertilization and embryo transfer (IVF—ET)was compared in 76 patients with polycystic ovaries (PCO) diagnosedon pre-treatment ultrasound scan, and 76 control patients whohad normal ovaries and were matched for age, cause of infertilityand stimulation regimen. Despite receiving significantly lesshuman menopausal gonadotrophin (HMG), patients with PCO, ascompared with controls, had significantly higher serum oestradiollevels on the day of human chronic gonadotrophin administration(5940 ± 255 versus 4370 ± 240 pmol/1, P < 0.001),developed more follicles (14.9 ± 0.7 versus 9.8 ±0.6, P < 0.001) and produced more oocytes (9.3 ± 0.6versus 6.8 ± 0.5, P = 0.003). However, fertilizationrates were reduced in the PCO patients (52.8 ± 3.4% versus66.1 ± 3.4%, P = 0.007). There was no significant differencein cleavage rates. The pregnancy rate/embryo transfer was 25.4%in the PCO group and 23.0% in the group with normal ovaries.There were three high order multiple pregnancies in the PCOgroup compared with none in the group with normal ovaries. Ofthe PCO patients, 10.5% developed moderate/severe ovarian hyperstimulationsyndrome (OHSS) compared with none of the controls (P = 0.006).Patients with and without PCO undergoing IVF have comparablepregnancy and livebirth rates. However, it is important to diagnosePCO before ovarian stimulation is initiated as these patientsare more likely to develop moderate or severe OHSS following1VF—ET.     

The double heterozygote of two endothelin-1 gene polymorphisms (G8002A and -3A/-4A) is related to big endothelin levels in chronic heart failure

The aim of this study was to focus on the relationship among the associated genotypes of G (8002) A and -3A/-4A endothelin-1 (ET-1) gene polymorphisms and some clinical and/or biochemical parameters in Czech (Caucasian) patients with chronic heart failure. Included in the study were 103 patients with chronic heart failure (functional classes NYHA II-IV, ejection fraction < 40%). The ET-1 gene polymorphisms were detected by polymerase chain reaction (PCR) and restriction fragment length polymorphism methods. A significant decrease in the ET-1-associated genotype AG3A4A number (double heterozygote) was observed in CHF patients with plasma big endothelin levels above 0.7 pmol/L compared to those with levels below 0.7 pmol/L (OR = 0.19; 95% confidence interval = 0.06-0.57; P = 0.005; Pcorr = 0.03). We found a significant decrease in the AG3A4A genotype number in the other groups compared to the group of patients with both big endothelin and endothelin-1 levels under 0.7 pmol/L (OR = 0.22; 95% confidence interval = 0.07-0.79; P = 0.02). The double heterozygote variants of two ET-1 gene polymorphisms were associated with significantly less risk for chronic heart failure with higher levels of big endothelin.     

Pregnancy: Oxygen promotes contraction by endothelin-1 in human umbilical artery

The influence of oxygen on the contractile response to endothelin-1in the human umbilical artery was investigated in vitro. Segmentsof human umbilical artery were suspended in organ baths to recordthe circular motor activity induced by endothelin-1 at a pO₂of 12 kPascal (kPa) or 45 kPa. Endothelin-1 induced a concentration-dependentcontraction which was significantly larger at 45 kPa O₂ comparedwith the contractile response at 12 kPa O₂.     

Adjuvant growth hormone therapy in poor responders to in-vitro fertilization: a prospective randomized placebo-controlled double-blind study

The objective of the study was to assess the effect of growthhormone (GH) supplementation to a combined gonado-trophin-releasinghormone agonist/human menopausal gonadotrophin (GnRHa/HMG) treatmentprotocol on ovarian response in ‘poor responders’undergoing in-vitro fertilization (IVF). GH or a placebo wereadministered in a prospective randomized double-blind manner.A total of 14 poor-responder patients (oestradiol < 500 pg/ml,less than three oocytes retrieved in two previous IVF cycles)were randomly allocated to a combined treatment of either GnRHa/HMG/GH (18 IU on alternate days, total dose 72 IU) or GnRHa/ HMGplacebo. No difference was found between the study and controlgroups in the number of HMG ampoules used, the number of follicles(>14 mm) and serum oestradiol concentrations on the day ofadministration of human chorionic gonadotrophin (HCG), the numberof oocytes retrieved and fertilized, and the number of embryostransferred. The GH group (n = 7) did not show a better ovulatoryresponse in the study cycles; mean ± SD serum oestradiolon day of HCG 411 ± 124 versus 493 ± 291 pg/ml,aspirated oocytes 2.2 ± 1.5 versus 1.9 ± 2.0.Interestingly, when the above results for the placebo groupwere compared with their previous cycles (serum oestradiol 403± 231 pg/ml; 0.4 ± 0.5 aspirated oocytes), a non-specificeffect was found. Follicular recruitment, oestradiol secretionby mature follicles and the number of oocytes retrieved in poorresponders were not improved by GH supplementation.     

Ovary and ovulation: Nitric oxide concentrations in the follicular fluid and apoptosis of granulosa cells in human follicles

To study the relationship between follicular atresia, apoptosis,and nitric oxide (NO) generation in follicular development,steroidogenesis, NO levels in follicular fluid and apoptosiswere analysed in the various sized follicles of women receivingovarian stimulation with human meno-pausal gonadotrophin (HMG)—humanchorionic gonado-trophin (HCG) treatments for in-vitro fertilization(IVF)-embryo transfer. The follicles were divided into threegroups by diameter: large follicle, 18 mm; medium follicle,12 and 15 mm; small follicle, 10 mm. Follicular fluid was obtainedfrom 20 women 34 h after HCG administration, and the concentrationsof oestradiol, progesterone and testosterone, and nitrite, nitrate,arginine and citrulline were measured. Granulosa cells obtainedfrom each group of follicular fluid were stained with Hoechstdye, and nuclear morphology was examined by a fluorescence microscopy.Oestradiol and progesterone concentrations in large follicleswere significantly (P < 0.01) higher than those in mediumor small follicles, and testosterone concentrations in smallfollicles were significantly (P < 0.01) higher than thosein large follicles. There were no significant differences inthe concentrations of nitrite, nitrate, arginine and citrullineamong three groups. The percentage of apoptotic cells with nuclearfragmentation was significantly (P < 0.01) higher in smallfollicles than in large follicles. The present results suggestedthat small follicles with poor response to HMG may undergo atresiathrough apoptosis. No significant difference in the follicularNO level between large and small follicles led us to speculateon a different responsiveness to NO in these two types of follicles.     

Hormonal characteristics of follicular fluid from women receiving either GnRH agonist or hCG for ovulation induction

BACKGROUND: A recent prospective randomized study from our groupcompared GnRH agonist (0.5 mg buserelin) and hCG (10 000 IU)for triggering of ovulation following a flexible antagonistprotocol. The agonist group showed a poor reproductive outcomedespite luteal phase support with progesterone and estradiol(E₂). In the present prospective observational study, the healthstatus of follicles from the above study was monitored by analysingthe hormonal content of frozen/thawed follicular fluid samples.The aim was to test whether the poor reproductive outcome couldbe related to a defective pre-ovulatory follicular maturationresulting in oocytes with a compromised developmental competence.METHODS: Hormone concentrations were measured in two individualfollicular fluid samples from each of 32 women receiving buserelinand 37 receiving hCG, thus representing a subset of the folliclesretrieved. RESULTS: Follicular fluid levels of LH in the agonistgroup as compared with the hCG group was 11.1 ± 0.5 versus3.6 ± 0.3 IU/l (mean ± SEM; P < 0.001); FSH,6.3 ± 0.6 versus 3.3 ± 0.2 IU/l (P < 0.001);hCG, not determined versus 139±8 IU/l; E₂, 1.9 ±0.2 versus 1.8 ± 0.2 µmol/l (P > 0.10); progesterone,70 ± 4 versus 93 ± 6 µmol/l (P < 0.001);inhibin-A, 36.9 ± 3.1 versus 37.1 ± 2.5 ng/ml(P > 0.10) and inhibin-B, 35.6 ± 2.8 versus 40.1 ±3.1 ng/ml (P > 0.10). Thus, pronounced hormonal differencesexist in follicular fluid, and the collective concentrationof all three gonadotropins and the follicular fluid concentrationof progesterone were much higher in the group of women receivinghCG for ovulation induction. CONCLUSION: The study suggeststhat GnRH agonist results in proper pre-ovulatory follicularmaturation, but the ovulatory signal – probably in synergywith the resulting pituitary down-regulation – is toolow to support appropriate corpus luteum (CL) function.     

Endocirnology: Comparison between prolactin, gonadotrophins and steroid hormones in serum and follicular fluid after stimulation with gonadotrophin-releasing hormone agonists and human menopausal gonadotrophin for an in-vitro fertilization programme

The inter-relationship between serum and follicular fluid prolactin,oestradiol, progesterone, follicle stimulating hormone (FSH),and luteinizing hormone (LH) in two groups of women was investigated.In group 1, 32 women were treated with gonadotrophin-releasinghormone agonist (GnRH-a) in a long term protocol and subsequentlystimulated with human menopausal gonadotrophin (HMG). In group2, 25 women were simultaneously stimulated with GnRH-a in ashort protocol with HMG. Follicular fluid was collected from54 follicles in group 1 and 47 follicles in group 2. Serum wasobtained on the day of human chorionic gonadotrophin (HCG) administration.Serum prolactin and oestradiol concentrations were significantlyhigher (P < 0.025 and P< 0.01, respectively) in group1 than in group 2. Serum LH (P < 0.005), FSH (P< 0.01)and progesterone (P < 0.025) were significantly lower ingroup 1 than in group 2. Follicular fluid prolactin was significantlyhigher (P < 0.005) in group 1. No differences were foundin follicular fluid progesterone and oestradiol. Follicularfluid LH was significantly lower (P < 0.005) in group 1.Serum prolactin correlated positively with oestradiol in bothgroups (P < 0.005 group 1; P < 0.02 group 2). No significantcorrelation was found between serum prolactin and LH in group1. We conclude that prolactin secretion is independent fromLH secretion. Hyperprolactinaemia, which is observed in womenstimulated with GnRH-a and HMG, is positively associated withincreased oestradiol.     

Localization and role of endothelin-1 and endothelin receptors in the human Fallopian tube.

The present experiments were designed to investigate the localization and role of endothelin-1 (ET-1) and endothelin receptors (ET(A) and ET(B)) in human Fallopian tubes obtained from patients in the follicular phase. Immunohistochemical studies revealed the predominant localization of ET-1 and of ET(B) receptors in the tubal epithelium and also within the muscle layer to a lesser degree. ET(A) receptors were dominant within the muscle layer. Scatchard plot analysis of the [(125)I]ET-1 binding also revealed the localization of ET(A) and ET(B) receptors on the Fallopian tubal membrane. A dissociation equilibrium constant of 34.6 +/- 3.3 pmol/l and a maximum binding site concentration of 1137.0 +/- 239.1 fmol/mg protein were obtained from the Scatchard plot analysis. Treatment of Fallopian tubal strips with ET-1 produced a tonic contraction which was inhibited by an ET(A) antagonist but not by an ET(B) antagonist. However, the increase in frequency and decrease in amplitude of rhythmic contractions caused by ET-1 were modulated by the ET(B) antagonist but remained unaffected by the ET(A) antagonist. These results suggest that ET-1 modulates the motility of the Fallopian tube through excitation of ET(A) and/or ET(B) receptors and possibly plays some role in oocyte capture.