Steroid secretion by human oocyte-coroma-comulus complexes associated with conceptions following in vitro fertilization

Seventy-one oocyte-corona-cumulus complexes (OCCCs) were obtained from eight women who conceived and delivered following in vitro fertilization (IVF) and eight women who were matched for the number of embryos transferred into the uterus but failed to conceive following the procedure. The steroid secretion of these OCCCs was assessed during a 24-hr culture period. Intermediate (N=35) and mature (N=36) complexes did not differ in steroid secretion, and progesterone was the major steroid secreted. OCCCs associated with a fertilized oocyte (N=59) produced significantly (P<0.02) higher levels of progesterone than those associated with nonfertilized oocytes (N=12) (515±53 vs 231±70 ngl OCCC/24 hr) but comparable levels of estradiol (9.17±1.14 vs 6.55±2.0 ng/OCCC/24 hr) and testosterone (0.34 ±0.09 vs 0.22±0.05 ng/OCCC/24 hr). Steroid production by OCCCs of oocytes which continued to cleave was not different from that of oocytes which did not cleave following fertilization, but the estradiol-to-testosterone ratio was significantly higher (31.4±3.5 vs 19.1±3.3; P<0.02) in the former group. The degree of cleavage was not related to the levels of steroid secretion. Cleaved oocytes associated with viable pregnancies (N=28) were derived from OCCCs which secreted significantly less testosterone than those which did not result in a pregnancy following transfer (N=26) (0.27±0.04 vs 0.41±0.04 ng/OCCC/24 hr). It is suggested that the steroids secreted by the corona-cumulus cells may directly affect the oocyte quality and its capacity to continue developing in the uterus.     

Cholesterol and steroid levels in human follicular fluids of human menopausal gonadotropin-induced cycles for in vitro fertilization

Fifty-eight follicular fluids (FF) were obtained from 18 women undergoing in vitro fertilization (IVF). Follicular development was induced by human menopausal gonadotropin (hMG) and follicular aspiration was performed 36 hr after an ovulatory dose of human chorionic gonadotropin (hCG). Two stages of oocyte-corona-cumulus complexes (OCCCs) morphological maturation was identified in this population: intermediate and mature. FF from which intermediate and mature OCCCs were obtained did not differ in 17-estradiol (E₂, progesterone (P), and cholesterol levels. Fifty OCCCs were fertilized and eight were not fertilized. No difference was found in E₂, P, and cholesterol levels in those two populations of OCCCs. Forty hours after insemination 50% of the oocytes were at the two-cell stage and 50% were at the three-cell stage. Steroids and cholesterol levels did not differ in FF from which those two groups of embryos originated. A direct correlation was found among the levels of cholesterol, E₂, and P in the FF. An inverted ratio of high-density lipoprotein (HDL) to low-density lipoprotein (LDL) and very low-density lipoprotein (VLDL) was found in FF compared to serum in 10 women. It is concluded that FF cholesterol levels have no value in predicting follicular maturation.     

Results of in vitro fertilization cycles in women aged 43-45 years.

Our objective was to evaluate the results of in vitro fertilization (IVF) cycles in the elderly (43-45 years old) female population. All consecutive women aged 43-45 years admitted to our IVF unit from January 1996 to December 2001 were enrolled in the study. Ovarian stimulation characteristics, number of oocytes retrieved, number of embryos transferred and pregnancy rate were assessed. Seven hundred and eight consecutive IVF cycles in 276 patients were evaluated. Two hundred and seven cycles were cancelled (cancellation rate 29.2%). Forty-seven patients achieved a clinical pregnancy (pregnancy rate 6.6% per cycle and 9.4% embryo transfer) with a 30% live birth rate. In patients who underwent embryo transfer, there were no differences between conception and non-conception cycles in patient's age, number of gonadotropin ampules used, length of ovarian stimulation, number of oocytes retrieved, fertilization rate or cleavage rate. However, the conception cycles were associated with a significantly lower peak estradiol level (p < 0.04) and higher number of total (p < 0.03) and good-quality (p < 0.005) embryos transferred, in addition to a lower ratio of estradiol level/number of follicles > 14 mm on day of human chorionic gonadotropin administration and of estradiol level/number of oocytes retrieved. We conclude that, although older female age is a major contributor to IVF failure, successful IVF cycles can be expected in patients aged 43-45 years in the presence of low ratios of peak estradiol to either number of follicles > 14 mm on day of human chorionic gonadotropin administration or number of oocytes retrieved that reach the stage of embryo transfer with at least two good-quality embryos.     

Androstenedione and progesterone concentrations in preovulatory follicular fluid correlate with successful fertilization and cleavage of human oocytes in vitro

OBJECTIVE: To determine if androstenedione (A) and progesterone (P) concentrations in preovulatory follicular fluid (FF) correlate with successful fertilization and cleavage of human oocytes in vitro. DESIGN: A retrospective randomized trial. SETTING: Hospital department of obstetrics and gynecology. PATIENTS: Fifty-five patients, ages 24 to 39 years, with normal menstrual cycles undergoing in vitro fertilization (IVF) for tubal infertility. INTERVENTIONS: Multiple follicular development was induced with clomiphene citrate and human menopausal gonadotropin. MAIN OUTCOME MEASURES: Relationships among FF steroid hormone, morphological maturity of oocyte-corona-cumulus complexes, and fertilization and cleavage of oocytes. RESULTS: Follicles with mature oocyte-corona-cumulus complexes and unfertilized oocytes contained significantly greater amounts of A (P less than 0.05) than those with mature oocyte-corona-cumulus complexes and fertilized ova, indicating the occurrence of atretic changes. Follicles yielding successfully fertilized and cleaved ova had significantly greater amounts of P (P less than 0.05) and A (P less than 0.01) but similar levels of 17 beta-estradiol compared with follicles yielding fertilized ova that failed to cleave. CONCLUSIONS: Follicles yielding oocytes that cleaved as a result of IVF have both a shift in steroidogenesis from estrogen to progestin accumulation and declining aromatase activity, thus reflecting progressive luteinization of the follicles.     

The effect of cocaine on oocyte development and the follicular microenvironment in the rabbit

We examined the effects of cocaine exposure in the rabbit on in vitro oocyte development and on steroidal content of follicular fluid (FF) and serum progesterone (P). Cocaine hydrochloride (0, 10, 20, 40, or 80 mg/kg) was administered daily subcutaneously for 5 days to New Zealand White female rabbits before superovulation. On the last day of cocaine administration, animals were given human chorionic gonadotropin intravenously, and laparotomy was performed 6 to 8 hours later. During laparotomy, ovaries were removed, the number of follicles recorded, oocytes retrieved, and FF was obtained. In vitro fertilization (IVF) was then performed on the oocytes and the rate of cleavage observed. For all cocaine dosage groups, no differences were observed in the number of follicles present, number of oocytes retrieved, or IVF and cleavage rates. Cocaine did, however, decrease periovulatory serum P, and FF P, whereas FF estradiol concentrations increased. This suggests that short-term cocaine exposure affects the follicular steroid milieu, possibly by delaying granulosa cell luteinization.     

The Follicular Endocrine Environment in Stimulated Cycles of Women with Endometriosis: Steroid Levels and Embryo Quality

Objective: To assess the endocrine milieu in follicles of stimulated cycles comparing women with and without endometriosis. Steroids were measured in follicular fluid (FF) and in in vitro culture of granulosa-luteal cells, and this status was related to the quality of the embryos obtained after IVF.

Design: Case-control study.

Setting: IVF program at the Instituto Valenciano de Infertilidad.

Patient(s): Twenty-four women with laparoscopically documented endometriosis and 26 controls undergoing IVF.

Intervention(s): Individual follicular aspiration, oocyte isolation, FF storage, and preparation of luteinized granulosa cells for culture; oocyte insemination and embryo cleavage in standard IVF.

Main Outcome Measure(s): Serum (day of ovum pickup) and FF measurements of estradiol, progesterone, testosterone, and androstenedione. Secretion of progesterone was measured in the cell-conditioned medium. Results were compared between patients with endometriosis and controls, as well as between oocytes that yielded embryos of different quality.

Result(s): Levels of progesterone in the FF increased with the severity of the disease, whereas testosterone accumulation in the FF decreased with the severity of the disease. An increase in progesterone accumulation in vitro was observed in basal and hCG-induced granulosa cell cultures. No difference was observed in terms of embryo quality, and no steroid marker was able to identify follicles with oocytes that displayed embryos of good or bad quality under the inverted microscope.

Conclusion(s): The data show differences in the steroidogenesis of follicles from stimulated women with and without endometriosis. These changes indicate good endocrine health but are not predictive of embryo quality.     

Correlation between follicular fluid content and the results of in vitro fertilization and embryo transfer. I. Sex steroids

The steroid content of 72 follicular fluids (FF) obtained from 42 women subjected to ovulation induction with the luteinizing hormone-releasing hormone analogue D-Trp6 and human menopausal gonadotropin was studied in terms of the evolution of in vitro fertilization (IVF) and embryo transfer (ET) results. The FF were classified into several categories based on oocyte evolution. Individual values of FF estrone and estradiol (E2), as well as androstenedione and testosterone could not be correlated with ET outcome. However, FF progesterone (P) levels for follicles leading to pregnancy were significantly lower when compared with those in the other categories. The correlation between the E2/P ratio and E2 permitted the definition of a band wherein IVF-ET outcome was successful and enabled the characterization of different functional follicular maturational states.     

Immunoreactive epidermal growth factor concentrations in follicular fluid obtained from in vitro fertilization

Immunoreactive epidermal growth factor (EGF) was measured in follicular fluid (FF) obtained at the time of oocyte retrieval for in vitro fertilization from cycles with (91 follicles) and without (128 follicles) the gonadotropin-releasing hormone agonist leuprolide acetate (LA). Follicular fluid immunoreactive EGF levels in the non-LA cycles correlated with androstenedione but not estradiol or progesterone levels from follicles with prophase I oocytes or from all follicles taken together, but not from metaphase I or metaphase II oocyte containing follicles alone. In non-LA cycles, FF immunoreactive EGF levels were lower in follicles that contained metaphase I or II oocytes than prophase I oocytes. Additionally, FF immunoreactive EGF levels were lower in follicles containing metaphase I or II oocytes from non-LA than LA cycles. We conclude that immunoreactive EGF is present in FF. Leuprolide acetate may affect FF immunoreactive EGF levels.     

The impact of high progesterone levels in the follicular phase of in vitro fertilization (IVF) cycles: A comparative study

Estrogen (E₂) and plasma progesterone (P₄) levels are valuable parameters for follicular development in in vitro fertilization (IVF) cycles. Furthermore, the progesterone concentration prior to, during, and following human chorionic gonadotropin (hCG) administration is an important marker for the detection of early luteinization and premature ovulation. The pattern of hormonal profile in relation to the number of oocytes retrieved, fertilized, and cleaved and the fate of the pregnancies achieved were compared in three groups of patients treated by the same protocol. Group I included 22 women who conceived with high progesterone levels on day hCG+1 (P_4'>2.5 ng/ml). Group II included 43 women who conceived with low P₄ values (P_4'<2.5 ng/ml), while group III included 46 patients in whom no pregnancies occurred. A significant decrease in fertilization, cleavage, and pregnancy rates was observed in patients with high progesterone levels on day hCG+1, compared to those with normal levels. Nevertheless, it is suggested that cycles with high P₄ levels in the preovulatory phase should not be canceled, as a fair chance for pregnancy still exists.     

Relationship between the steroid and prolactin concentration in follicular fluid and the maturation and fertilizaton of human oocytes

Seventyeight follicles and their follicular fluid were aspirated from 46 women undergoing in vitro fertilization (IVF) procedures after stimulation of the ovaries with a low-dose human menopausal gonadotropin/human chorionic gonadotropin stimulation regimen. The concentrations of estradiol (E₂), progesterone (P), testosterone (T), and prolactin (PRL) were measured in follicular fluid and related to the maturation of the oocyte-corona-cumulus complex (OCCC) and the fertilization of oocytes. Follicles containing mature oocytes had significantly higher follicular fluid E₂ and P levels than follicles with intermediate and immature oocytes. A constant decrease in PRL and T values with advancing follicular maturation was observed. Similar results were obtained when the fertilizing ability of the oocytes was examined. The gradual decline in follicular fluid PRL and T levels during follicular development was connected with increasing E₂ and P biosynthesis and therefore seems to be an important precondition for normal follicular and oocyte maturation.     

Perifollicular vascularity monitoring and scoring: a clinical tool for selecting the best oocyte

The aim of our study was to assess relationship between perifollicular vascularity and outcome of in vitro fertilization (IVF) cycles. In our study, women who received embryos originating from oocytes developed in well vascularized follicles had a statistically higher pregnancy rate than women who received embryos chosen without considering follicles of origin.     

A preliminary study on in vitro fertilization and embryo transfer

Forty-two women with infertility were enrolled in an in vitro fertilization and embryo transfer (IVF/ET) program, from July, 1986 to February, 1988, at the First Affiliated Hospital, Human Medical University. CC/hMG/hCG and hMG/hCG regimens for ovulation induction were used for 35 and 7 patients. Monitoring methods consisted of daily follicular ultrasonography and serum estradiol measurements. Human chorionic gonadotropin 10,000IU was administered when the leading follicle reached 18mm in diameter and serum E2 level was equal to or more than 1,480 pmol/L (400 pg/ml). Twenty-one laparoscopies and nineteen laparotomies for oocyte retrieval were performed. The IVF/ET results using CC/hMG/hCG for ovarian stimulation were as follows: an average of 6.1 follicles were aspirated and 3.5 oocytes recovered for one case. The recovery rate and fertilization rate was 57.0% and 60.1% respectively. In twenty-five women one to five embryos were transferred and a clinical pregnancy occurred in one who received three embryos. Laparotomy for oocyte retrieval showed that more oocytes could be obtained than laparoscopy. The purity of CO2 used for laparoscopy was considered to affect the IVF results.     

Bromocriptine inhibition of anesthesia-induced hyperprolactinemia: effect on serum and follicular fluid hormones, oocyte fertilization, and embryo cleavage rates during in vitro fertilization

Thirty-two patients undergoing in vitro fertilization (IVF) were given bromocriptine either 1 or 12 hours before anesthesia or received no drug to determine what effect suppression of transient, anesthesia-induced hyperprolactinemia would have on peripheral and follicular fluid hormones, fertilization and cleavage rates, and pregnancy. Thirty minutes after anesthesia, there was a 120-ng/mL rise in serum prolactin (PRL) in control patients versus an insignificant change in women given bromocriptine. Levels of PRL in follicular fluid were significantly less, and estradiol (E2) levels were higher (P less than 0.05) in all bromocriptine-treated patients compared with controls, whereas follicular fluid levels of progesterone (P), inhibin activity, and midluteal serum P were unaffected. Although fertilization and pregnancy rates were similar, a greater proportion of fertilized oocytes from bromocriptine-treated patients advanced to cleaving embryos compared with controls (95% versus 63%, respectively; P less than 0.001). We conclude that bromocriptine, given before anesthesia, can suppress transient, anesthesia-induced hyperprolactinemia and dramatically alter follicular fluid concentrations of PRL and E2. Although these changes in hormonal milieu affected neither oocyte fertilization nor pregnancy rate in our IVF patients, they seemed to have a positive influence on embryonic development after IVF.     

Effects of leuprolide acetate on follicular fluid hormone composition at oocyte retrieval for in vitro fertilization

The follicular fluid (FF) in 91 follicles from 17 women treated with leuprolide acetate (LA) before stimulation with gonadotropins for in vitro fertilization were analyzed for estradiol (E2), progesterone (P), androstenedione, prolactin, and human chorionic gonadotropin (hCG) and compared with the concentrations in 128 follicles from 31 women treated with gonadotropins alone. The FF E2 concentration in LA-treated patients was significantly lower than in non-LA patients for all oocyte maturational stages. Follicles containing metaphase II oocytes had significantly lower concentrations of P and hCG in LA-treated patients. These differences persisted when analysis was limited to follicles whose oocytes fertilized normally. These data indicate that in the presence of LA, normal oocyte maturation can occur despite lower intrafollicular concentrations of E2 and P.     

Relationship between midcycle luteinizing hormone surge quality and oocyte fertilization

OBJECTIVE: To determine whether alterations in preovulatory follicular fluid (FF) levels of LH, FSH, and steroids are associated with the probability of fertilization. DESIGN: Retrospective analysis of prospective study results. SETTING: Reproductive medicine clinic of a university teaching hospital. PATIENT(S): Infertile women, with unstimulated, apparently regular cycles in an IVF research program. INTERVENTION(S): Measurement of preovulatory FF levels of LH, FSH, E2, and P and serum LH levels by fluoroimmunometry. MAIN OUTCOME MEASURE: Oocyte fertilization. RESULT(S): There were 84 transferable embryos (rate of normal fertilization and cleavage, 67%), and 41 oocytes (33%) failed to fertilize. Analysis of the matched FF showed that the median concentration of FF LH was significantly higher for cleaving embryos than for unfertilized oocytes (14.6 vs. 10.4 IU/L). Serum LH concentrations were similarly higher in cycles with cleaving embryos. There were no statistically significant differences in FF concentrations of FSH, E2, or P in the two groups. CONCLUSION: Reduced preovulatory FF LH levels are associated with impaired fertilization of oocytes in vitro, despite normal FF FSH and steroid levels.     

Does gravidity influence the success of in vitro fertilization-embryo transfer cycles?

OBJECTIVE: To evaluate the influence of gravidity on the results of in vitro fertilization (IVF)-embryo transfer (ET) cycles. PATIENTS AND METHODS: All consecutive women aged <35 years admitted to our IVF unit from January 2002 to December 2004 were enrolled in the study. Only patients undergoing one of their first three IVF cycle attempts were included. Gravidity, ovarian stimulation characteristics, number of oocytes retrieved, number of embryo transferred and clinical pregnancy rate were assessed. RESULTS: Three hundred and forty-two consecutive IVF cycles were evaluated. One hundred and sixty-one cycles were from nulligravidas and 181 from women with a history of at least one previous clinical pregnancy. Forty-eight (29.8%) clinical pregnancies were observed in the nulligravida group and 56 (30.9%) in the gravida group. There were no differences between nulligravidas and gravidas in causes of infertility, length of ovarian stimulation, peak estradiol and progesterone levels, number of oocytes retrieved, fertilization rate and number of embryos transferred. Gravidas were significantly older (30.4 vs. 27.6 years, p < 0.001) and used more gonadotropin ampoules (36.1 vs. 31.8, p < 0.004) compared with the nulligravidas. CONCLUSIONS: Patient gravidity has no influence on the likelihood of achieving pregnancy in IVF-ET cycles.     

Transvaginal ultrasound-guided embryo transfer improves outcome in patients with previous failed in vitro fertilization cycles

OBJECTIVE: To determine the effect of transvaginal ultrasound-guided ET in IVF cycles performed on patients who had previously failed to conceive from IVF and compare the results to previous cycles where ultrasound guidance was not used. DESIGN: Retrospective clinical study.Setting: Private practice IVF program. PATIENT(S): One hundred twenty-nine women undergoing consecutive cycles of IVF where fresh embryos were transferred. INTERVENTION(S): Transvaginal ultrasound guidance was used during transfer of embryos. MAIN OUTCOME MEASURE(s): Patient age, number of ampules of gonadotropin used, maximum E(2) level, number of oocytes retrieved, number of two pronuclei embryos obtained, number of embryos transferred, mean embryo score, implantation and pregnancy rate. RESULT(S): There was no difference in any of the clinical parameters measured in IVF cycles resulting in pregnancy when transvaginal ultrasound-guided ET was used compared to the failed cycles when there was no ultrasound guidance. Of the patients who previously had failed IVF cycles and subsequently had IVF cycles with ultrasound guidance, those who became pregnant had higher mean embryo scores than those who did not become pregnant. Overall implantation and pregnancy rates were higher during the study period when transvaginal ultrasound guidance was used than in the previous 3 years when it was not used. CONCLUSION(S): Transvaginal ultrasound-guided ET may be responsible for successful IVF cycles in patients who had previously failed to conceive when embryos were transferred by the clinical touch method. Transvaginal ultrasound guidance may also be responsible for an overall increase in embryo implantation and pregnancy compared to the use of the clinical touch method.     

Pregnancy rates in varying age groups after in vitro fertilization: a comparison of follitropin alfa (Gonal F) and follitropin beta (Follistim)

OBJECTIVE: Our purpose was to assess the efficacy of two recombinant follicle-stimulating hormones, follitropin beta (Follistim, Organon, West Orange, NJ) and follitropin alfa (Gonal F, Serono, Norwell, Mass) on pregnancy rates in varying age groups of women undergoing in vitro fertilization (IVF). STUDY DESIGN: Three hundred sixty-five IVF cycles were retrospectively compared, 233 by use of follitropin beta and 132 by use of follitropin alfa, both after gonadotropin-releasing hormone agonist down-regulation. Assignment to each medication was indiscriminate. The primary outcome measured was pregnancy evidenced by fetal heartbeat on ultrasonography. Secondary outcomes included days of stimulation, ampules per patient cycle, estradiol level on the day of human chorionic gonadotropin administration, total follicles present on the day of human chorionic gonadotropin administration, follicles greater than 14 mm, oocytes retrieved, mature eggs, fertilization rate, and embryos transferred. Outcomes were stratified by age, including women less than 36 years old, 36 to 39 years old, and more than 39 years old. RESULTS: There was no significant difference between follitropin beta and follitropin alfa in either the primary or secondary outcomes, although the pregnancy rate was significantly decreased with advancing age. CONCLUSION: Success rates are similar, when stratified by age, in women undergoing IVF with either follitropin beta or follitropin alfa.     

High progesterone/estradiol ratio in follicular fluid at oocyte aspiration for in vitro fertilization as a predictor of possible pregnancy

Eighteen women undergoing in vitro fertilization (IVF) procedures were studied. All had optimal (900 to 1600 pg/ml) peak serum estradiol (E2) response to the same stimulation regimen with clomiphene citrate and menotropins; fertilization rate was above 64%; and two to four embryos in two to eight cell stages were replaced in each patient. All were considered to have optimal chances for conception. The authors compared progesterone (P), E2, and P/E2 ratio in serum and follicular fluid (FF) at the time of oocyte aspiration in eight patients who conceived (group I) and ten who did not (group II). Mean serum P and E2 levels and serum P/E2 ratio were not significantly different between the groups. In contrast, mean FF P concentrations (ng/ml) were significantly (P less than 0.05) higher in group I (9721 versus 5385), as was FF P/E2 ratio (19.0 versus 11.8; P less than 0.02). There was no significant difference in mean FF E2 concentrations between the groups. These data indicate that in IVF cycles with optimal serum E2 response to the stimulation protocol, FF P and P/E2 ratio at the time of oocyte aspiration may be predictive of subsequent implantation and pregnancy.     

Follicular fluid concentrations of vascular endothelial growth factor, inhibin A and inhibin B in IVF cycles: are they markers for ovarian response and pregnancy outcome?

OBJECTIVE(S): The aim of this study was to measure concentrations of vascular endothelial growth factor (VEGF), inhibin A and inhibin B in follicular fluid (FF) of women undergoing to in vitro fertilization (IVF) cycles and to determine their relationship with ovarian response and pregnancy. STUDY DESIGN: Follicular fluid was collected from 58 patients undergoing oocyte retrieval for IVF. Ovulation was induced with GnRH analogues and gonadotropins. Follicular fluids of mature follicles (>17 mm) were aspirated and pooled for each patient. Follicular fluid steroid hormone levels (E2, P) and VEGF, inhibin A, inhibin B concentrations were studied. The serum levels of E2, P and VEGF were also assessed on the day of the oocyte retrieval. These parameters and characteristics of the cycles were compared between the pregnant (group 1) and non pregnant (group 2) patients. RESULTS: The serum and FF VEGF levels were found to be significantly lower in the group in whom the pregnancy was achieved (P < 0.001). The FF inhibin A and FF inhibin B were found to be significantly higher in pregnant group (P < 0.001). However, age, day 3 FSH, dosage of gonadotropin administered, fertilization rate, sperm count, motile and morphologically normal sperm percentage were not significantly different in the two groups. There was an negative correlation between VEGF and number of follicles, number of oocytes, FF inhibin A, FF inhibin B. The number of oocytes retrieved, the fertilization rate were positively correlated with FF inhibin B and FF inhibin A. CONCLUSION: This study demonstrated that decreased FF VEGF, serum VEGF and elevated FF inhibin A and B are associated with better ovarian response and high pregnancy rate.