Decline of follicular oocyte maturation inhibitor coincident with maturation and achievement of fertilizability of oocytes recovered at midcycle of gonadotropin-treated women.

To examine whether a decline in follicular oocyte maturation inhibitor (OMI) is associated with attainment of oocyte maturation and fertilizability, OMI was measured in follicular fluid (FF) of 39 follicles of 20 normal women given human menopausal gonadotrophin and human chorionic gonadotrophin to induce follicular growth and maturation. Oocytes were aspirated per laparoscope, the fluid was saved, and the egg was observed, incubated, and inseminated with the husband's sperm. Concepti that developed to the 4- to 8-cell stage were transferred to the uterus and the women were followed for pregnancy. OMI activity in each FF was measured by using cultured cumulus-enclosed porcine oocytes (30-40 oocytes per FF sample). Estrogen, progesterone, oocytes (30-40 oocytes per FF sample). Estrogen, progesterone, and delta 4-androstenedione were measured in FF by radioimmunoassay. The FF of 13 preovulatory follicles yielding oocytes that were mature and fertilizable had significantly less OMI activity (mean +/- SEM) (0.58 +/- 0.10 unit/ml) compared to follicles yielding immature oocytes (2.8 +/- 0.56 units/ml; n = 9), atretic oocytes (5.5 +/- 2.5 units/ml; n = 7), or preovulatory oocytes with fractured zonae (1.9 +/- 0.63 units/ml; n = 7). The estrogen concentration (mean +/- SEM) of preovulatory follicles yielding mature fertilizable eggs or mature eggs with fractured zonae was greater (396 +/- 34 ng/ml; n = 20) compared to follicles yielding immature or atretic eggs (203 +/- 59 ng/ml; n = 9 and 97 +/- 47 ng/ml; n = 7, respectively; P less than 0.05). Progesterone concentration (mean +/- SEM; ng/ml) of FF was generally elevated in all preovulatory follicles (635 +/- 53) compared to immature or atretic follicles (230 +/- 64 and 76 +/- 17, respectively; P less than 0.05). It may be concluded that in normal follicle maturation there is a decline in OMI in the follicle containing an oocyte that becomes mature and fertilizable. There is also an increase in estrogen, progesterone, and follicle size. It is also possible to have an abnormal follicle maturation when there is an increase in size as well as FF, estrogen, and progesterone, but withut a decline in OMI--a situation which can lead to production of a nonfertilizable oocyte.     

Oocyte fertilization in vitro is associated with high follicular immunoreactive beta-endorphin levels

Follicular fluid (FF) levels of immuno reactive beta-Endorphin (i.r. beta-EP), i.r. gamma-Endorphin (i.r. gamma-EP), i.r. alpha-melanocyte-stimulating hormone (i.r. alpha-MSH), androgens and estrogens were measured in 76 preovulatory follicles obtained, after gonadotropin stimulation from 19 women undergoing in vitro fertilization (IVF). The aim of the study was to investigate the relationships existing between peptide contents of FF and both oocyte-cumulus-corona-complex (OCCC) maturity and the success of IVF. Peptides and steroids were measured by RIA after FF extraction with liquid chromatography and ethyl-ether, respectively. Out of the total of 76 oocytes, 52 were fertilized in vitro and 35 of them underwent normal cleavage and were transferred. Among the three peptides, only i.r. beta-EP levels were higher in FF from follicles which contained oocytes that were subsequently fertilized (127.6 +/- 16.2 pmol/L mean +/- SE) than in FF from follicles which contained oocytes that did not subsequently fertilized (62.9 +/- 8.4, p less than 0.04). Independent of subsequent fertilization, i.r. alpha-MSH values in FF were 5 times higher than those of i.r. beta-EP and i.r. gamma-EP. In the presence of a morphologically mature oocyte, FF i.r. gamma-EP levels (165.2 +/- 45.3 pmol/L) were higher than in FF from follicles yielding immature (63.6 +/- 13.5, p less than 0.01) or luteinized (32.7 +/- 9.2, p less than 0.01) oocytes. Steroid levels in FF did not change in relation to oocyte maturity or subsequent oocyte fertilization.(ABSTRACT TRUNCATED AT 250 WORDS)     

The effect of the follicular fluid adenosine 3',5'-monophosphate degradation rate on successful fertilization and cleavage of human oocytes

Follicular fluid (FF) and oocytes were obtained from 19 women for in vitro fertilization. Ovulation was induced with clomiphene citrate and human menopausal gonadotropin. Thirty-seven FF samples containing mature oocyte-corona-cumulus complexes were used to measure steroids, gonadotropins, and cAMP. The FF specimens were divided into three groups: follicles yielding ova that were fertilized and cleaved (group A), follicles containing fertilized ova without further cleavage (group B), and follicles containing nonfertilized oocytes (group C). The FF levels of progesterone, 17 beta-estradiol, LH/hCG, and FSH did not differ significantly among the groups. Mean FF cAMP concentrations declined from 17.5 +/- 2.5 pmol/mL 15 min after follicle aspiration to 3.6 +/- 0.9 pmol/mL after 3 h. The initial FF cAMP concentration did not differ significantly among three groups. However, the cAMP degradation rate in group A (119 +/- 13 X 10(-4) pmol/min) was significantly greater than that in group B (62 +/- 10 X 10(-4) pmol/min) or group C (75 +/- 8 X 10(-4) pmol/min). In conclusion, an increased intrafollicular cAMP degradation rate was associated with successful fertilization and cleavage of human oocytes in vitro. These data suggest that the degradation rate of FF cAMP may be a marker of optimal follicular development and oocyte maturation.     

Adenosine 3',5'-monophosphate levels in human follicular fluid: relationship to oocyte maturation and achievement of pregnancy after in vitro fertilization

cAMP, estradiol (E2), and progesterone levels were determined in 24 follicular fluid samples obtained from 8 women who conceived after in vitro fertilization and in 47 samples from 26 women who did not. Follicular development was induced by human menopausal gonadotropin, and maturation of retrieved oocytes was assessed by the degree of cumulus mucification and corona dispersal. The mean follicular fluid cAMP concentration was significantly (P less than 0.001) lower in women who became pregnant than in those who did not (106 vs. 241 pmol/ml), while the mean E2 level was significantly (P less than 0.01) higher (727 vs. 497 ng/ml), and the progesterone to E2 ratio was significantly (P less than 0.05) lower (9.5 vs. 18.0). Overall, follicles of immature, intermediate, and mature oocytes did not differ in cAMP content. However, intermediate and mature oocytes from women who became pregnant were derived from follicles containing significantly (P less than 0.01) lower cAMP levels than those of women who did not become pregnant (66 and 122 vs. 233 and 288 pmol/ml, respectively). Furthermore, fertilized oocytes leading to conception originated from follicles with significantly (P less than 0.001) lower cAMP concentrations than the follicles that yielded nonfertilized oocytes or fertilized oocytes not leading to conception (92 vs. 270 and 240 pmol/ml, respectively). Similarly, significantly (P less than 0.05) lower cAMP levels were found in the follicular fluid of cleaved oocytes resulting in a pregnancy compared to those that did not (86 vs. 236 pmol/ml). It is concluded that low levels of cAMP are associated with successful fertilization and cleavage of human oocytes in vitro resulting in viable pregnancies and may, therefore, be used as a marker of optimal follicular development in in vitro fertilization cycles.     

Gonadotropin and prolactin levels in follicular fluid of human ova successfully fertilized in vitro

Follicular fluid (FF) and oocytes were obtained from 94 follicles of 36 women for fertilization in vitro. Ovulation was induced with human menopausal gonadotropin, and follicular aspiration was performed 36 h after an ovulatory injection of hCG. The concentrations of immunoreactive hCG, FSH, and PRL were correlated with the degree of maturation of the oocyte-corona-cumulus complex mass (OCCC), fertilization, rate of cleavage, and the incidence of pregnancy after embryo transfer. Immature OCCC were derived from follicles that contained significantly lower levels of FSH than those from which intermediate and mature OCCC were derived (5.2 +/- 0.6 vs. 11.1 +/- 1.2 mlU/ml; P less than 0.05). FF from oocytes that were successfully fertilized contained higher levels of both hCG and FSH than FF surrounding oocytes that did not fertilize (136.7 +/- 8.7 vs. 108.5 +/- 10.3 mlU/ml hCG; 10.55 +/- 0.6 vs. 5.3 +/- 0.8 mIU FSH, respectively). There was no correlation between early embryonic growth rate and FF concentrations of FSH, hCG, and PRL. Ova reaching the two-cell stage 40 h after fertilization in vitro were associated with the same FF concentrations of FSH, hCG, and PRL as those that cleaved to the four-cell stage. The PRL concentration in FF was significantly higher in mature fertilized ova and in fertilized ova that were associated with a successful pregnancy. It is suggested that the intrafollicular concentration of FSH is associated with the degree of mucification of the OCCC, but FF levels of both FSH and hCG are associated with successful fertilization. High levels of PRL in FF were associated with successful pregnancy and may imply a role of this hormone in oocyte maturation.     

Flow cytometric analysis of deoxyribonucleic acid in human granulosa cells from in vitro fertilization cycles: relationships to oocyte maturity and fertilizability and to follicular fluid steroids

We measured the mitotic activity of granulosa cells, sex steroid concentrations in follicular fluids, and the maturity and fertilizability of oocytes from 49 follicles. Flow cytometric measurements of DNA were used to determine the percentage of cells in G0/G1, S, and G2/M phases of the cell cycle. Mitotic index was designated as the percentage of granulosa cells in S + G2/M. The progesterone concentration and the progesterone to estradiol ratio in follicular fluids were inversely correlated to mitotic index (r = -0.506; P less than 0.001, and r = -0.320; P less than 0.02, respectively). Estradiol and androstenedione levels did not correlate with the mitotic index. The mitotic index was higher in follicles with immature oocytes [25.6 +/- 2.0% (+/- SE); n = 7] than in follicles with mature oocytes (15.6 +/- 1.2%; n = 41; P less than 0.001). The mitotic index of granulosa cells was lowest in follicles with oocytes that fertilized (15.5 +/- 1.8%), higher in follicles with oocytes that remained unfertilized (18.5 +/- 1.3%; P less than 0.03), and highest in follicles with oocytes that fertilized abnormally (24.0 +/- 2.1%; P less than 0.02). Differences in maturity or fertilizability of oocytes were not associated with variations in follicular fluid progesterone concentrations. The study supports the concept that mitotic activity is decreased when granulosa cells become luteinized. During early follicular growth it is assumed that estradiol and perhaps androstenedione may be important regulators of cell division. Our findings suggest that progesterone, perhaps acting as an antiestradiol, is more important in controlling granulosa cell division of preovulatory follicles during the late follicular phase.     

Periovulatory follicular fluid hormone levels in spontaneous human cycles

We measured follicular fluid hormone levels in 48 normally cycling infertile women who underwent follicle puncture and oocyte retrieval during diagnostic laparoscopy at time-bracketed intervals after an endogenous LH surge. Follicular fluid LH, FSH, PRL, estrone (E1), estradiol (E2), progesterone (P), androstenedione (A), and testosterone (T) concentrations and P/E2 and A/E2 ratios were determined. Oocytes were classified as germinal vesicle (gv), metaphase I (mI), metaphase II (mII), or degenerating (dg). Follicular fluid (ff) hormone levels then were correlated with the stage of oocyte maturation. There were no differences in ff E1 or E2 levels at any stage of oocyte maturation, except that the mean ff E2 concentration was significantly (P less than 0.05) lower in ff containing dg oocytes [2,474 +/- 1,435 (+/- SE) nmol/L] than in those containing the other oocyte stages. The mean P levels were significantly (P less than 0.0001) higher in ff containing mI (48,781 +/- 10,240 nmol/L) and mII (41,801 +/- 11,098 nmol/L) oocytes than in ff containing gv oocytes (1371 +/- 696 nmol/L). The mean A level was highest (P less than 0.01) in dg-associated ff. Similarly, T was highest (P less than 0.05) in ff containing dg (52 +/- 14 nmol/L) oocytes than in ff containing mI (10.7 +/- 10.1 nmol/L) or mII (10.1 +/- 4 nmol/L) oocytes, and it was also elevated (P less than 0.05) in gv ff (72 +/- 33 nmol/L) compared to mII ff. The above differences also were reflected in the P/E2 ratio, which was significantly higher (P less than 0.05) in mI and mII ff, as well as in the A/E2 ratio, which was higher (P less than 0.05) in ff containing mI and mII oocytes compared to ff containing gv or dg oocytes. These data define the evolving changes in the microenvironment of the follicular fluid of preovulatory follicles of normally cycling women. They also provide reference points for analysis of ff obtained from women during stimulated cycles intended for in vitro fertilization.     

Human follicular fluid insulin concentrations

In mammals, insulin stimulates granulosa cell aromatase activity and steroid production and is a regulating factor of oocyte maturation. To assess the role of insulin in human follicular and oocyte maturation, human follicular fluid was obtained 32-36 h after hCG administration at the time of oocyte recovery for in vitro fertilization. Follicular fluid insulin levels, measured by RIA, ranged from undetectable (less than 2 microU/ml) to 65.4 microU/ml. In women treated with human menopausal gonadotropin (n = 21), clomiphene citrate (n = 4), and human menopausal gonadotropin/clomiphene citrate (n = 14), follicular fluid insulin concentrations were 18.0 +/- 4.3 (+/- SE), 10.2 +/- 4.2, and 12.0 +/- 3.8 microU/ml, respectively (P = NS). Similarly, there was no significant difference in follicular fluid insulin concentrations in follicles with mature (n = 33) or immature (n = 6) oocytes (13.3 +/- 2.7 vs. 24.7 +/- 9.5 microU/ml) or in oocytes which eventually did (n = 35) or did not (n = 4) fertilize (16.4 +/- 3.0 vs. 3.2 +/- 0.8 microU/ml). Follicular fluid insulin levels (n = 30) correlated positively with follicular fluid progesterone levels (P less than 0.05), but not with follicular fluid estradiol or androstenedione levels or the estradiol to androstenedione ratio. The relationship of follicular fluid insulin and progesterone levels suggests that, as in other mammals, follicular fluid insulin may have a physiological role in follicular maturation.     

Plasma levels of plasminogen activator inhibitor type 1, beta- thromboglobulin, and fibrinopeptide A before, during, and after treatment of acute myocardial infarction with alteplase

Plasma levels of plasminogen activator inhibitor type-1 (PAI-1), beta- thromboglobulin (beta TG), and fibrinopeptide A (FPA) were followed over 24 hours in 30 patients treated with alteplase for acute myocardial infarction. Samples were taken at baseline (T Oh), after 90 minutes (under alteplase, no heparin, T 1.5h), after 120 minutes (under alteplase and heparin, T 2h), 30 minutes after thrombolytic therapy (T 3.5h), as well as 12 hours (T 12h) and 24 hours (T 24h) after baseline. PAI-1 antigen levels (55 +/- 9 ng/mL at T Oh, mean +/- SEM) decreased to 35 +/- 5 (T 1.5h) and 40 +/- 6 (T 2h) ng/mL under alteplase, before increasing to 84 +/- 22 (T 3.5h), 130 +/- 30 (T 12h), and 64 +/- 7 (T 24h) ng/mL after therapy, P less than .001. A high baseline PAI-1 activity (18 +/- 3 ng/mL) decreased to 2.0 +/- 0.4 (T 1.5h) and 1.7 +/- 0.2 (T 2h) under alteplase and increased to 32 +/- 5 (T 12h) and 19 +/- 3 (T 24h) ng/mL after therapy (P less than .0001). beta TG levels (339 +/- 105 ng/mL at T Oh) decreased to 203 +/- 48 (T 2h), 154 +/- 51 (T 3.5h), 187 +/- 40 (T 12h), and 142 +/- 32 (T 24h) ng/mL under heparin (P less than .01). FPA levels (34 +/- 9 ng/mL at T Oh) increased to 85 +/- 15 ng/mL under alteplase alone (T 1.5h) and normalized under heparin (11 +/- 4, 6 +/- 2, 4 +/- 2, and 3 +/- 1 ng/mL at T 2h, T 3.5h, T 12h, and T 24h, respectively). A high level of FPA at T 3.5h correlated with reocclusion (33 +/- 12 ng/mL, n = 4 v 2.9 +/- 0.5 ng/mL, n = 21, P less than .005). We conclude that plasma levels of PAI- 1 antigen as well as activity markedly increase after alteplase therapy of acute myocardial infarction. The high activity of PAI-1 and decreasing beta TG levels suggest that platelets do not contribute significantly to this phenomenon. The marked increase of FPA levels under recombinant tissue-type plasminogen activator alone and its normalization under heparin emphasize the important role of concomitant anticoagulation in controlling further intravasal fibrin generation under alteplase.     

Follicular fluid steroid levels in dysmature and mature follicles from spontaneous and hyperstimulated cycles in normal and anovulatory women

Follicular maturity and atresia have been defined previously, both hormonally and microscopically, in normal ovulatory women. Ovarian hyperstimulation with clomiphene citrate and human menopausal gonadotropins in normal women is carried out for the purpose of aspirating oocytes from several large follicles for in vitro fertilization. Alterations in follicular fluid (FF) hormone levels occur with hyperstimulation regimens, and some of these large follicles (greater than 18 mm) appear morphologically atretic. We have used the term dysmature to describe those large follicles that have an abnormal oocyte morphological appearance and cannot be fertilized in vitro. Mature follicles have been defined by their size, their oocyte morphological appearance, and their ability to be fertilized in vitro. FF from small (2-3 mm) and large (greater than 18 mm) mature and dysmature follicles were obtained from 10 untreated ovulatory women. Mature and dysmature follicles also were obtained from clomiphene and human menopausal gonadotropin-treated normal (n = 11) and anovulatory (n = 5) women. In untreated cycles, the FF steroid content of the small follicles characterized these follicles to be atretic. FF from dysmature follicles from spontaneous untreated cycles had higher concentrations of dihydrotestosterone, 5 alpha-androstane-3 alpha,17 beta-diol, and 17 beta-estradiol (E2) and lower progesterone (Prog) and Prog to E2 ratios (P less than 0.05). Compared to mature follicles from untreated patients, hyperstimulated mature follicles from ovulatory women had higher FF E2 concentrations and lower Prog to E2 ratios (P less than 0.05). In ovulatory patients, the FF concentrations of testosterone were higher and FF Prog and Prog to E2 ratios were lower (P less than 0.05) in the dysmature than in the mature follicles. Mature follicles from hyperstimulated ovulatory patients and those from hyperstimulated anovulatory patients were similar except for lower FF Prog, higher FF E2, and lower Prog to E2 ratios in the anovulatory group. Dysmature follicles from hyperstimulated anovulatory patients had lower FF androstenedione and dihydrotestosterone, but were generally similar to mature follicles. The percentages of dysmature follicles occurring among all large (greater than 18 mm) follicles that were aspirated were similar in ovulatory (34%) and anovulatory (45%) patients. FF steroid concentrations did not correlate with serum levels of testosterone and E2 at the time of follicle aspiration in any patient group. In conclusion, FF androgen concentrations in hyperstimulated follicles were unrelated to morphological maturity.(ABSTRACT TRUNCATED AT 400 WORDS)     

Oocyte maturity and human follicular fluid prostanoids, gonadotropins, and prolactin after administration of clomiphene and pergonal

Follicular fluid (FF) and oocytes were obtained from 130 follicles of 52 women in whom ovulation was induced with human menopausal gonadotropin (Pergonal) and clomiphene citrate. Follicular aspiration was performed 36 h after an ovulatory injection of hCG. The concentrations of LH, FSH, PRL, and the prostanoids prostaglandin E2 (PGE2), PGF2 alpha, PGI2 (as 6-oxo-PGF2 alpha), and thromboxane A2 (as TXB2) in the FF were measured by RIA and related to the degree of maturation of the oocyte-corona-cumulus complex mass (OCCC). FF obtained from follicles with immature OCCC contained significantly lower concentrations of all four prostanoids (median concentrations, picograms per mL: PGE2, 88; PGF2 alpha, 85; 6-oxo-PGF1 alpha, 40; TXB2, 50) than those with intermediate OCCC (PGE2, 175; PGF2 alpha, 325; 6-oxo-PGF1 alpha, 130; TXB2, 65) and mature OCCC (PGE2, 425; PGF2 alpha, 860; 6-oxo-PGF1 alpha, 235; TXB2, 78; all P less than 0.01). There were no significant differences between the maturity of the complexes and the concentrations of LH, FSH, or PRL. There were significant correlations between the FF concentrations of LH and FSH and those of all of the prostanoids, but not with PRL, concentrations. These results indicate that the synthesis of prostanoids in the human Graafian follicles may be modulated by gonadotropins and consolidates the view that prostanoids may play a role in human oocyte maturation and ovulation.     

Markers of sodium and volume homeostasis in pregnancy-induced hypertension.

Normal pregnancy is associated with increased levels of digitalis-like factor (DLF) and erythrocyte sodium-lithium countertransport (RBC CTT), which return to normal levels postpartum. Patients with pregnancy-induced hypertension (PIH) have greater increases in both factors than women with normotensive pregnancies. This study was designed to determine if both abnormalities are observed concomitantly in PIH, if they correlate with blood pressure, if they correlate negatively with a hormonal index of volume status (PRA), and if they differ in women with and without proteinuria. Twenty-six normotensive women and 26 women with PIH were studied in the third trimester. Thirteen of these patients were also studied 6 months postpartum. Women with PIH, compared to those who were normotensive, had higher RBC CTT (0.49 +/- 0.04 vs. 0.36 +/- 0.03 mmol Li/L cells.h; P = 0.004) and DLF (0.30 +/- 0.3 vs. 0.20 +/- 0.03 microgram digoxin equiv./L; P = 0.01) and lower PRA [4.58 +/- 0.76 vs. 7.34 +/- 0.86 ng/mL.h (1.27 +/- 0.21 vs. 2.04 +/- 0.24 ng/L.s); P = 0.001]. All three parameters correlated significantly with diastolic blood pressures (RBC CTT and DLF positively (P less than or equal to 0.02) and PRA negatively (P = 0.03). Comparisons of DLF, RBC CTT, and PRA demonstrated a significant correlation of RBC CTT and DLF for normotensive pregnant women only (r = 0.38; P = 0.05). Patients with PIH were further analyzed according to whether proteinuria (24-h urinary protein, greater than 0.30 g; urine dipstick, greater than or equal to 2+) was present or absent. There was no significant difference in diastolic blood pressure or PRA between the hypertensive subpopulations, although there was a tendency for those without proteinuria to have lower PRAs [3.85 +/- 0.80 ng/mL.h (1.07 +/- 0.02 ng/L.s)] than those with proteinuria [5.31 +/- 1.30 ng/mL.h (1.48 +/- 0.36 ng/L.s)]. RBC CTT was significantly higher (P less than 0.05) in women with PIH without proteinuria, whereas serum DLF was significantly higher in women with PIH with proteinuria (P less than 0.05). In 13 women studied 6 months postpartum, there was a significant reduction in serum DLF, RBC CTT, and PRA for all women and in blood pressure for women who had had PIH (P less than 0.01). Thus, women with PIH, compared to normotensive pregnant women, had abnormalities in a variety of factors known to be volume sensitive or indicative of salt- and volume-sensitive forms of hypertension.(ABSTRACT TRUNCATED AT 400 WORDS)     

Correlation between follicular fluid content and the results of in vitro fertilization and embryo transfer. II. Inhibin and aromatase inhibitor activity

The inhibin content and aromatase inhibitor activity (AIA) of 72 follicular fluids (FF) obtained from 42 women undergoing in vitro fertilization (IVF) and embryo transfer (ET) were studied as a function of IVF ET outcome. Inhibin levels were determined by bioassay (BA) and RIA; AIA was measured by BA. The inhibin content of follicles characterized as immature by their estradiol (E2) levels and E2/progesterone (P) ratios was significantly lower (P less than 0.05) than that of mature follicles (i.e. leading to pregnancy). The mean AIA for mature follicles were significantly lower than AIA in groups where pregnancy was not obtained. AIA for follicles from which a pregnancy was obtained for each ET was also significantly lower than that in FF characterized as immature of hypermature. The highest E2/AIA and inhibin BA/AIA ratios were associated with the highest incidence of successful IVF ET outcome. No correlation was found between AIA and inhibin, on the one hand, and E2, delta 4-androstenedione, E2/P, and PRL, on the other. However, a positive correlation was found between inhibin (RIA and BA) and P, reflecting the production of inhibin by granulosa cells during luteinization. These studies allowed us to conclude that FF inhibin levels do not differ according to IVF ET outcome, but are an index of follicular maturation. AIA not only constitutes an index of follicular maturation and granulosa cell luteinization, but is of predictive value for IVF ET outcome as E2/AIA and inhibin BA/AIA ratios.     

Effects of incremental infusions of arginine vasopressin on adrenocorticotropin and cortisol secretion in man

Arginine vasopressin (AVP) regulates ACTH release under certain conditions, and exogenously administered AVP is used clinically to stimulate ACTH secretion. We attempted to determine at what plasma concentration AVP can stimulate ACTH release. Six normal men were given infusions of AVP (Ferring) or vehicle between 1600 and 1700 h on five occasions: 1) saline (30 mL/h); 2) 10 ng AVP/min; 3) 30 ng AVP/min; 4) 100 ng AVP/min; and 5) 300 ng AVP/min. Plasma AVP, ACTH, and cortisol concentrations were measured every 10 min during the infusions. Basal plasma AVP levels were less than 1 ng/L (less than 0.92 pmol/L). The lowest AVP dose raised plasma AVP into the range found in fluid-deprived subjects (7-8 ng/L;6.5-7.3 pmol/L), but had no effect on plasma ACTH concentrations. AVP in a dose of 30 ng/min also had no effect. The 100 ng AVP/min dose raised plasma AVP concentrations to 51.4-65.5 ng/L (46-60 pmol/L). This increase led to a transient insignificant increase in plasma ACTH from 13.9 +/- 1.2 (+/- SEM) ng/L (3.1 +/- 0.3 pmol/L) to 20.0 +/- 1.4 ng/L (4.4 +/- 0.3 pmol/L), while plasma cortisol rose significantly from 146 +/- 10 to 209 +/- 19 nmol/L (P less than 0.01) after 60 min of infusion. The 300 ng AVP/min dose raised plasma AVP levels to about 260 ng/L (239 pmol/L); the maximal plasma ACTH and cortisol levels were 39.5 +/- 5.0 ng/L (8.7 +/- 1.1 pmol/L; P less than 0.01) and 348 nmol/L (P less than 0.01), respectively. Thus, peripheral plasma AVP levels have to be raised high above the physiological range before ACTH release is stimulated. We conclude that any AVP reaching the adenohypophysis through the peripheral circulation is of much less importance for the regulation of ACTH secretion than is AVP derived from the pituitary portal circulation.     

Differential regulation of inhibin A and inhibin B by luteinizing hormone, follicle-stimulating hormone, and stage of follicle development

Inhibin B and inhibin A exhibit unique patterns of secretion across the follicular phase of the menstrual cycle. To test the hypothesis that the distinct patterns of inhibin B and inhibin A secretion result from differential regulation by LH and FSH, a series of controlled experiments was designed to dissect the specific effects of LH and FSH at distinct stages of follicle development. After GnRH agonist desensitization, women with small antral follicles were treated with recombinant human LH (rhLH), rhFSH, or rhFSH and estradiol (E(2)). rhLH or rhFSH was also administered when follicles reached the preovulatory stage in gonadotropin-stimulated or spontaneous cycles. At the small antral stage of development, rhFSH, but not rhLH, administration increased inhibin B (17.4 +/- 4.6 to 321.0 +/- 97.0 pg/mL; P < 0.05), inhibin A (0.6 +/- 0.1 to 2.6 +/- 0.6 IU/mL; P < 0.05), and E(2) [15.8 +/- 3.6 to 95.3 +/- 26.9 pg/mL (58.0 +/- 13.2 to 349.8 +/- 98.7 pmol/L); P < 0.05]. The inhibin B increase preceded inhibin A by 48 h. Addition of E(2) to FSH resulted in a greater increase in inhibin B (23.2 +/- 6.4 to 865.2 +/- 294.5 pg/mL; P < 0.05) than FSH alone (P < 0.05). At the preovulatory stage, rhLH administration increased inhibin A (15.9 +/- 10.3 to 21.5 +/- 13.7 IU/mL; P < 0.05) and E(2) [669.4 +/- 285.5 to 943.6 +/- 388.1 pg/mL (2457.4 +/- 1048.1 to 3464.0 +/- 1424.7 pmol/L); P < 0.05], but not inhibin B, as did rhFSH administration in spontaneous cycles [E(2): 226.4 +/- 102.7 to 264.7 +/- 121.0 pg/mL (831.1 +/- 377.0 to 971.7 +/- 444.2 pmol/L); P < 0.05; inhibin A: 2.6 +/- 1.3 to 3.7 +/- 1.9 IU/mL; P < 0.05; and inhibin B: 76.3 +/- 32.2 to 77.6 +/- 32.8 pg/mL; P = NS]. These findings suggest that increases in both FSH and E(2) in the early follicular phase result in increased inhibin B secretion at early stages of follicle development, whereas the selective LH rise in the late follicular phase favors inhibin A secretion from more mature follicles. Thus, both differential secretion of LH and FSH and the stage of follicle development determine the patterns of inhibin A and inhibin B secretion in the normal menstrual cycle.     

Correlation of human follicular fluid inhibin activity with spontaneous and induced follicle maturation

We sought to correlate the inhibin activity of individual ovarian follicles (greater than 16 mm in diameter) from untreated (7 patients; 7 follicles), clomiphene-stimulated (150 mg/day; menstrual cycle days 5-9; 9 patients, 14 follicles), and human menopausal gonadotropin (hMG)-stimulated (150 IU/day; menstrual cycle days 3-11; 8 patients; 23 follicles) ovarian cycles and to correlate these results with the follicular fluid (FF) steroid concentration. Follicular aspirates were obtained via laparoscopy from 24 regularly menstruating patients when the diameter of the largest follicle reached 20 mm, as determined by serial ultrasonography. FF concentrations of estradiol, progesterone, testosterone, 17-hydroxyprogesterone, and androstenedione were determined by RIA. Inhibin activity was determined using the inhibition of basal 24-h FSH secretion by dispersed rat anterior pituitary cells. Inhibin values were highest among the follicles aspirated from those patients who received hMG [277 +/- 31 (+/- SE) U/ml] compared to untreated subjects (51 +/- 13 U/ml) or those who received clomiphene (96 +/- 14 U/ml). Estradiol was highest in FF from untreated patients (2295 +/- 1155 ng/ml) compared to levels in patients who received hMG (368 +/- 1.76 micrograms/ml) or clomiphene (1049 +/- 174 ng/ml). FF progesterone values were highest in untreated patients (9.4 +/- 2.59 micrograms/ml) compared to those in hMG-treated (5.04 +/- 1.76 micrograms/ml) and clomiphene-treated patients (7.82 +/- 1.24 ng/ml). FF 17-hydroxyprogesterone values (7.82 +/- 1.24 ng/ml). FF 17-hydroxyprogesterone values were similarly higher in the untreated (1.55 +/- 0.21 micrograms/ml) and clomiphene-treated (2.54 +/- 0.27 micrograms/ml) patients than in the hMG-treated group (0.73 +/- 0.09 micrograms/ml). FF androstenedione (untreated, 50.7 +/- 30 ng/ml; clomiphene-treated, 73.4 +/- 23.4 ng/ml; hMG-treated, 60.2 +/- 19.8 ng/ml) and testosterone (6.66 +/- 2.45, 5.98 +/- 1.46, and 6.39 +/- 2.16 ng/ml, respectively) concentrations in all three patient groups were similar. In untreated patients, there was a highly significant positive correlation between intrafollicular inhibin activity and FF estradiol, testosterone, and androstenedione concentrations and a statistically significant negative correlation between intrafollicular inhibin activity and FF progesterone concentrations. Patients receiving clomiphene therapy demonstrated at least two different response patterns, one with a positive and one a negative correlation between intrafollicular inhibin activity and FF steroid concentrations. The patients receiving hMG therapy had no statistically significant correlation between intrafollicular inhibin     

Pulsatile growth hormone secretion in patients with acromegaly and normal men: the effects of growth hormone-releasing hormone infusion

Twenty-four GH secretory patterns were studied before and during continuous infusions of GHRH in six patients with active acromegaly and in six normal adult men. GH release was episodic in both groups. Control subjects showed a normal diurnal variation in GH release, with the majority of GH released at night (2200-0800 h); mean levels were 1.5 +/- 0.4 (SE) ng/mL (day) and 4.2 +/- 0.8 ng/mL (night). Acromegalics had no diurnal variation in GH; levels were 45.3 +/- 13.7 ng/mL (day) and 39.8 +/- 12.2 ng/mL (night). Acromegalics demonstrated an increased frequency of GH pulses compared to normals (11.8 +/- 0.8 vs. 2.2 +/- 0.3/24 h). During continuous 24-h infusions of GHRH, the normal subjects continued to show a diurnal variation in GH release, but GH pulse frequency increased to a rate (11.7 +/- 1.4 pulses/24 h) very similar to that of the patients with acromegaly. In contrast, GHRH infusion did not alter the GH pulse frequency in the acromegalics. GHRH increased the mean levels of GH in both groups (patients 80.2 +/- 20.3 vs. 41.0 +/- 12.1 ng/mL, x +/- SE. P less than 0.05; controls 10.2 +/- 2.0 vs. 3.33 +/- 0.5 ng/mL, P less than 0.01). Some of the patients with acromegaly showed a progressive decline in GH levels during the infusion period, suggesting desensitization or exhaustion of releaseable stores; however, GH levels remained above basal values in all patients. After the 24-h GHRH infusions, the GH response to a bolus of GHRH was diminished in the normal subjects (2.1 +/- 0.9 vs. 16.8 +/- 5 ng/mL, x +/- SE; P less than 0.01) but not in the acromegalic patients (30.2 +/- 8.9 vs. 35.5 +/- 12.5 ng/mL; NS). These results indicate that GH release is episodic under basal conditions and during continuous GHRH infusion in both acromegalic and normal subjects, indicating the importance of other modulators of GH release, such as somatostatin, which may remain pulsatile even in acromegaly.     

Titrating luteinizing hormone surge requirements for ovulatory changes in primate follicles. II. Progesterone receptor expression in luteinizing granulosa cells

The events in granulosa cells that are initiated by the midcycle LH surge during luteinization of the primate follicle are poorly defined. This study was designed 1) to determine whether an ovulatory dose of hCG can induce progesterone receptors (PR) in macaque granulosa cells, and if so, 2) to begin titrating gonadotropin requirements for PR expression and progesterone production by luteinizing granulosa cells. Rhesus monkeys were treated with human FSH and LH for up to 9 days to stimulate the growth of multiple follicles. The next day, animals (n = 4-5/group) received: 1) no ovulatory stimulus; 2) 1000 IU hCG, im; 3) one injection of 100 micrograms GnRH, sc (GnRH-1); 4) three injections of GnRH (GnRH-3) at 3-h intervals (0800, 1100, and 1400 h); or 5) two injections of 50 micrograms GnRH agonist (GnRHa), sc, 8 h apart (0800 and 1700 h). Granulosa cells obtained by follicle aspiration 27 h after the hCG or initial GnRH/GnRHa injection or on days 8 or 10 from animals receiving no ovulatory stimulus were processed for indirect immunocytochemistry using a monoclonal antibody to human PR (JZB39). Specific staining for PR, determined by comparing cells incubated with PR antibody vs. a nonspecific antibody, was undetectable in granulosa cells from monkeys without an ovulatory stimulus. In contrast, the majority (64 +/- 5%) of cells from hCG-treated animals stained intensely for PR. In the GnRH/GnRHa groups, granulosa cells from only one animal (i.e. one GnRH-3 monkey) showed positive staining for PR. During 24-h culture in Ham's F-10 medium containing 10% monkey serum, basal progesterone production by cells from the hCG-treated group (2163 nmol/L.8 x 10(4) cells) was higher than that by cells from the no ovulatory stimulus/GnRH-1/GnRH-3/GnRHa groups (60, 111, 194, and 332 nmol/L, respectively). However, granulosa cells from the hCG-treated group were less responsive to hCG in vitro in terms of enhanced progesterone production (2 times control levels) than cells from the other four groups (up to 30 times control levels). This study provides direct evidence that an ovulatory dose of hCG induces PR expression in granulosa cells of luteinizing follicles during stimulated cycles in rhesus monkeys. However, repeated injections of GnRH/GnRHa that produced surge levels (greater than 100 ng/mL) of endogenous LH for up to 14 h failed to induce PR expression or progesterone production by granulosa cells. Thus, an extended LH surge more typical of that in the normal menstrual cycle (48-50 h) may be necessary for PR expression and luteinization of granulosa cells in primate follicles.     

Levels of steroid-binding proteins and steroids in human preovulatory follicle fluid and serum as predictors of success in in vitro fertilization-embryo transfer treatment

In an attempt to identify the embryos and cycles that have the best chances of resulting in establishment of pregnancies, after in vitro fertilization-embryo transfer (IVF-ET) treatment, the concentrations of sex hormone-binding globulin (SHBG) and cortisol-binding protein (CBP) were measured, using two new enzyme-linked immunosorbent assays in serum and follicle fluid (FF) from 30 women (125 FF) undergoing IVF-ET. The concentrations were compared to those of total estradiol and total progesterone, and correlated to oocyte cleavage and the establishment of pregnancies. Serum concentrations of CBP were significantly higher in women who became pregnant (1469 +/- 108) nM (+/- SEM] than in those who did not (CBP, 1200 +/- 58 nM; P less than or equal to 0.05). The concentrations of SHBG were not significant different in these two groups of women (72.4 +/- 9.3 and 60.8 +/- 4.2 nM, respectively; P greater than or equal to 0.10). By contrast, in FF significantly higher concentrations of both SHBG and CBP were found in women achieving pregnancy (SHBG, 56.1 +/- 2.8 nM; CBP, 1198 +/- 37 nM) than in those who did not (SHBG, 45.5 +/- 1.4 nM; P less than or equal to 0.001; CBP, 1079 +/- 29 nM; P less than or equal to 0.01). A positive correlation was found between serum and FF levels of both SHBG (r = 0.85; P less than or equal to 0.001) and CBP (r = 0.70; P less than or equal to 0.001). FF levels of estradiol and progesterone did not differ regardless of whether the oocyte cleaved. However, a significant reduction of estradiol was found in fluid from follicles in which the oocyte cleaved and resulted in pregnancy (3046 +/- 180 nM) than in fluid from follicles in which the oocyte cleaved but without establishment of pregnancy (4162 +/- 282 nM; P less than or equal to 0.001). There was no correlation between estradiol and SHBG and between progesterone and CBP. However, levels of FF progesterone above 15,000 nM combined with CBP concentrations above the mean concentration found in FF (1,127 nM) were related with oocyte cleavage in 87% of the cases. The overall cleavage rate is 56%. The higher levels of SHBG and CBP in serum compared to those in FF, and the positive relationship between serum and FF levels suggest that both proteins arise from the circulation. The similar levels in serum and FF indicate that neither SHBG nor CBP is responsible for maintaining the concentration gradient of estradiol and progesterone from follicle to plasma.(ABSTRACT TRUNCATED AT 400 WORDS)     

Serum bioactive follicle-stimulating hormone-like activity increases during pregnancy

It is presently accepted that high circulating steroid levels during pregnancy suppress pituitary immunoreactive FSH (I-FSH) secretion from early pregnancy to term. In this study we tested the hypothesis that bioactive FSH (B-FSH) in the sera of pregnant women would likewise be suppressed. Serum samples were obtained from one woman daily for 21 days after the I-LH surge, and in a second, every 10 min for 10 h on days 10, 20, and 27 after the I-LH surge. Single serum samples were also obtained from women at 2-41 weeks of gestation. In all samples, I-LH/hCG, I-FSH, B-FSH, progesterone (P), and estradiol (E2) were measured. In the first woman, serum I-FSH levels were suppressed from days 9-21 after the LH surge (0.2 +/- 0.04 ng/mL; 1.9 +/- 0.3 IU/L), whereas serum B-FSH increased linearly from 1.7 ng/mL (5.3 IU/L) on day 9 post-I-LH surge to 6.0 ng/mL (18.6 IU/L) on day 17 (r = 0.91; P less than 0.01) and remained elevated through day 21. In the second woman, the mean +/- SE of the serum I-FSH concentrations were 0.7 +/- 0.04, 0.1 +/- 0.01, and 0.1 +/- 0.01 ng/mL (5.5 +/- 0.3, 0.9 +/- 0.1, 1.1 +/- 0.1 IU/L) on days 10, 20, and 27 post-I-LH surge, respectively. The mean serum B-FSH concentrations increased from 3.1 +/- 0.2 ng/mL (9.6 +/- 0.7 IU/L) on day 10 to 7.5 +/- 0.6 ng/mL (23.2 +/- 1.7 IU/L) on day 27. Using the DETECT pulse analysis method at the 0.01 confidence level, the serum I-FSH pattern was relatively nonpulsate, while serum B-FSH averaged 8, 7, and 10 pulses/10 h on post-LH surge days 10, 20, and 27. Incremental peak amplitudes were 3.5 +/- 0.7, 4.6 +/- 0.4, and 8.0 +/- 1.3 ng/mL (10.9 +/- 2.2, 14.3 +/- 1.2, and 24.8 +/- 4.0 IU/L), respectively. In the cross-sectional study, serum I-FSH levels remained low, while serum B-FSH concentrations increased from 4.2 +/- 0.8 ng/mL (13.2 +/- 2.5 IU/L) during the early first trimester to 59.3 +/- 2.9 ng/mL (183.8 +/- 9.0 IU/L) by the late third trimester. There was no correlation between serum B-FSH and hCG levels. Incubates of term placentae secreted B-FSH in preference to I-FSH (i.e. B-FSH, 33.0 +/- 9.6 ng/h/g placental tissue; I-FSH, 0.7 +/- 0.1 ng/h/g).(ABSTRACT TRUNCATED AT 400 WORDS)