The effect of deglycosylated human chorionic gonadotropin on corpora luteal function in healthy women.

Fourteen healthy young women were studied through a control and a treatment menstrual cycle in two series of experiments. In the first series, they were given one of four doses of deglycosylated human chorionic gonadotropin (hCG) as a 24-hour infusion during the mid-luteal phase of the cycle. In these studies, there were no significant alterations of the length of the luteal phase of the treatment cycle, and there was no decrease in serum progesterone (P) during the infusion. In fact, serum P increased during the infusion. In the second series of studies, five subjects were given a 48-hour infusion of normal saline during the control cycle, and a 48-hour infusion of deglycosylated alpha-intact beta-hCG during the treatment cycle, both being administered during the mid-luteal phase. Treatment did not alter luteal phase duration and, again, increased serum P. It is concluded that deglycosylated preparations of hCG are not clinically useful as luteinizing hormone antagonists, probably because of residual agonist activity.

Adenylate cyclase activity of the corpus luteum during the oestrous cycle of the pig.

Basal adenylate cyclase values for corpora lutea (CL) removed from cyclic gilts on Days 3, 8, 13 and 18 were 178 +/- 61, 450 +/- 46, 220 +/- 25 and 208 +/- 18 pmol cAMP formed/min/mg protein, respectively. Basal activity was significantly elevated on Day 8 (P less than 0.001). LH-stimulatable adenylate cyclase values for CL from Days 3, 8, 13 and 18 were 242 +/- 83, 598 +/- 84, 261 +/- 27 and 205 +/- 17 pmol cAMP formed/min/mg protein respectively. Serum progesterone concentrations of 12 gilts bled every 2 days through one complete oestrous cycle ranged from 1.1 to 26.9 ng/ml with highest values between Days 8 and 12. The decline in serum progesterone concentrations was coincident with the decrease in basal adenylate cyclase activity. There was no LH-stimulatable adenylate cyclase activity present in the CL at the specific times of the oestrous cycle examined. We conclude that progesterone secretion by the pig CL is apparently dependent on basal activity of adenylate cyclase.

Deglycosylated human follitropin: characterization and effects on adenosine cyclic 3',5'-phosphate production in porcine granulosa cells.

Chemical deglycosylation of gonadotropins with hydrogen fluoride (HF) has facilitated the investigation of the structure-function relationship of the individual peptide and oligosaccharide moieties in the mechanism of hormone action. These studies have dealt almost exclusively with lutropin or human choriogonadotropin. We report here the chemical characterization and biological properties of deglycosylated human follitropin (degly hFSH). Results indicate that deglycosylation of hFSH by HF removes 89% of the total carbohydrate without disruption of the peptide chain or significant loss of amino acid residues. However, a change in the conformation of the molecule was observed by measurement of the far-ultraviolet circular dichroic spectrum. The degly hFSH showed a 44% reduction in binding when tested in a FSH radioimmunoassay utilizing a polyclonal antibody. Binding of the degly hFSH to FSH-responsive tissues showed that the altered hormone bound with equal or better avidity than the intact hormone while the association constants were approximately the same for both preparations. The degly hFSH alone did not stimulate the FSH-stimulatable adenylyl cyclase (AC) activity of cellular homogenates of small follicle porcine granulosa cells. Furthermore, degly hFSH was a potent antagonist of hFSH-stimulatable AC activity when coincubated with intact hFSH. In intact granulosa cells, both the hFSH and the degly hFSH stimulated cAMP production and release by these cells. However, the degly hFSH was one-tenth as effective as the intact hormone.(ABSTRACT TRUNCATED AT 250 WORDS)

Inhibition of adenylyl cyclase activity in rat corpora luteal tissue by glycopeptides of human chorionic gonadotropin and the alpha-subunit of human chorionic gonadotropin.

Indirect evidence has indicated that the carbohydrate moieties of the glycoprotein hormones are involved in the activation of the receptor-adenylyl cyclase system of reproductive tissues. In the present study, we have isolated the glycopeptides (GP) from human chorionic gonadotropin (hCG), the alpha-subunit of hCG, fetuin, and bovine gamma-globulin (b gamma G). These along with a number of synthetic oligosaccharides were tested for their ability to inhibit adenylyl cyclase (AC). There was less than 0.001% cross-reactivity of the GP from hCG, hCG alpha, fetuin, and b gamma G when tested in a double-antibody hCG radioimmunoassay or rat corpora lutea radioreceptor assay. The GP of fetuin, b gamma G, and the synthetic oligosaccharides did not inhibit AC activity of 2000 g corpora lutea membranes when coincubated with 100 ng of hCG/mL (ED50). However, when the GP of hCG and hCG alpha were included with intact hCG, there was a dose-related inhibition. Inhibition of cyclase activity was enhanced when the hCG GP were desialylated. This occurred without a change in the lag time of hCG activation which was calculated to be 1-1.5 min. Changing the concentration of ATP and Mg2+ did not affect the inhibitory effects of the hCG alpha GP on hCG-stimulated AC activity. Inhibition by hCG GP followed uncompetitive kinetics. The inhibition by the GP of hCG seems to be restricted to the LH/hCG-stimulatable AC system because the same dosage of hCG GP which inhibited the rat luteal AC system did not have any effect on the rat hepatocyte AC system when coincubated with glucagon or on NaF-stimulated activity in luteal membranes.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenylyl cyclase system of the small preovulatory follicles of the domestic hen: responsiveness to follicle-stimulating hormone and luteinizing hormone.

The purpose of this study was to determine if the granulosa cells of the small preovulatory follicles of the domestic hen are a target tissue for follicle-stimulating hormone (FSH). The third largest (F3), fourth largest (F4), and fifth largest (F5) follicles were removed from hens at 20, 12, 6 and 2 h before ovulation of the F1 follicle. Basal, FSH- and luteinizing hormone (LH)-stimulable adenylyl cyclase (AC) activities were measured in the granulosa cells. Isolated granulosa cells of the F5 follicle, obtained 20 h before ovulation of the F1 follicle, were incubated with ovine (o) or turkey (t) FSH and progesterone (P4) was assayed in the medium. Basal AC activity was similar for F5, F4 and F3 granulosa cells except for an increase (P less than 0.01) in F3 follicles removed 2 h before ovulation of the F1 follicle. The FSH-stimulable AC activity of F5, F4 and F3 granulosa cells was elevated over basal (P less than 0.01). The greatest responsiveness was seen in the F5 follicle and the least in the F3 follicle. LH-stimulable AC activity was absent in the F5 follicle but present in the F4 and F3 follicles with the greater responsiveness in the F3 follicle. Isolated F5 granulosa cells secreted significant amounts of P4 in response to oFSH and tFSH. The data indicate that: 1) FSH stimulates the AC system of granulosa cells of the smaller preovulatory follicles (F5 greater than F4 greater than F3) while LH stimulates the AC system of granulosa cells of the larger follicles (F3 greater than F4), and 2) FSH promotes P4 production by granulosa cells of F5 follicles.(ABSTRACT TRUNCATED AT 250 WORDS)

Steroid concentrations in isolated theca and granulosa layers of preovulatory follicles during the ovulatory cycle of the domestic hen.

The purpose of the present investigation was to measure the concentrations of progesterone (P4), testosterone (T) and estradiol-17 beta (E2) in isolated theca and granulosa layers of the five preovulatory follicles of the domestic hen. The largest follicle (F1), the second largest (F2), third largest (F3), fourth largest (F4) and fifth largest (F5) follicles were removed at 24, 18, 12, 6 and 2 h before the expected ovulation. Theca and granulosa layers were isolated and P4, T, E2 and protein concentrations determined. Protein concentrations of the granulosa and theca layers increased 5- and 15-fold, respectively, during the five ovulatory cycles prior to ovulation. As the follicle approached ovulation, there was a linear decrease in E2 concentration of the theca layer with the most significant decrease (P less than 0.001) occurring between 24 and 18 h of the ovulatory cycle. In the F3 and F4 theca layers, there was a significant increase (P less than 0.005) in E2 at 6 h of the ovulatory cycle. Fluctuations in T concentrations in theca and granulosa layers were similar. There was a significant increase (P less than 0.05) in T in both layers of the F2, F3 and F4 follicles at 6 h followed by a decrease (P less than 0.005) in the theca layers at 2 h of the ovulatory cycle. The P4 concentration of the granulosa layer increased gradually during follicular maturation, with the greatest increase occurring in the F2 and F1 granulosa layers between 18 and 12 h of the ovulatory cycle.(ABSTRACT TRUNCATED AT 250 WORDS)