Systemic Progesterone Concentrations Research Articles

Synergistic Activation of the Human Type II 3β-Hydroxysteroid Dehydrogenase/Δ5-Δ4 Isomerase Promoter by the Transcription Factor Steroidogenic Factor-1/Adrenal 4-binding Protein and Phorbol Ester

Steroidogenic factor-1/adrenal 4-binding protein (SF-1/Ad4BP) is an orphan nuclear receptor/transcription factor known to regulate the P450 steroid hydroxylases; however, mechanisms that regulate the activity of SF-1/Ad4BP are not well defined. In addition, little is known about the mechanisms that regulate the human steroidogenic enzyme, type II 3beta-hydroxysteroid dehydrogenase (3beta-HSD II), the major gonadal and adrenal isoform. Regulation of the 3beta-HSD II promoter was examined using human adrenal cortical (H295R; steroidogenic) and cervical (HeLa; non-steroidogenic) carcinoma cells. H295R cells were transfected with a series of 5' deletions of 1251 base pairs (bp) of the 3beta-HSD II 5'-flanking region fused to a chloramphenicol acetyltransferase (CAT) reporter gene followed by treatment with or without phorbol ester (phorbol 12-myristate 13-acetate; PMA). CAT assay data indicated that the region from -101 to -52 bp of the promoter was required for PMA-induced expression. A putative SF-1/Ad4BP regulatory element, TCAAGGTAA, was identified by sequence homology at -64 to -56 bp of the promoter. Cotransfection of HeLa cells with the -101 3beta-HSD-CAT construct and an expression vector for SF-1/Ad4BP increased CAT activity 49-fold. Subsequent treatment with PMA induced an unexpected synergistic increase in transcriptional activity 540-fold over basal. Mutation of the putative response element (TCAAGGTAA to TCAATTTAA) abolished SF-1-induced CAT activity and the synergistic response to PMA. Gel mobility shift assays confirmed that SF-1/Ad4BP interacts with the putative element and transcripts for SF-1/Ad4BP were detected in H295R cells by Northern analysis. These data are the first to demonstrate 1) regulation of a non-cytochrome P450 steroidogenic enzyme promoter by SF-1/Ad4BP, 2) a powerful synergistic effect of PMA on SF-1/Ad4BP-induced transcription, and 3) the importance of the SF-1/Ad4BP regulatory element in the regulation of the 3beta-HSD II promoter.

Luteal function in unilaterally hysterectomized ewes treated with gonadotropin-releasing hormone.

An experiment was conducted to examine effects of GnRH administered to ewes during metestrus on subsequent luteal and uterine functional interrelationships. Treatments consisted of GnRH (0 or 100 micrograms/d) and uterine status (intact or unilaterally hysterectomized [UHYST]). On d 12 of an estrous cycle, all ewes were unilaterally ovariectomized and one-half of these ewes were subjected to contralateral UHYST. Corpora lutea in the remaining ovary were enucleated. One-half of the intact and UHYST ewes were injected i.v. with 2 mL of GnRH on d 2 and 3 after subsequent estrus, and the remaining ewes were injected similarly with 2 mL of saline. Jugular blood samples were collected at 15-min intervals after GnRH or saline injection and analyzed for serum LH. Caudal vena caval and(or) jugular blood were collected daily from d 5 to 10 and on d 12 and 14 of the cycle and analyzed for progesterone, oxytocin, and prostaglandin F2 alpha (PGF2 alpha). Injection of ewes with GnRH increased serum concentrations of LH within 60 min compared with those of saline-treated ewes (P = .01). Treatment with GnRH did not alter concentrations of oxytocin in intact or UHYST ewes (P > .10) but on d 12 and 14 mean jugular concentrations of oxytocin were greater (P = .01) in intact than in UHYST ewes. Vena cava plasma concentrations of PGF2 alpha did not differ (P > .10) among treatments. Treatment with GnRH did not affect (P > .10) serum concentrations of progesterone, but concentrations of this steroid over the sampling period tended to be greater (P = .09) in UHYST ewes than in intact ewes. In conclusion, treatment of intact and UHYST ewes with GnRH failed to alter systemic concentrations of oxytocin, PGF2 alpha, and progesterone; however, the concentrations of oxytocin were affected by unilateral hysterectomy.

Fetal development in cattle with multiple ovulations.

Treatment of lactating and nonlactating parous cows (n = 379) with 12 mg of FSH-P to evaluate development of multiple bovine fetuses resulted in ovulation rates ranging from 1 to 27 corpora lutea (CL). Fertilization rate (i.e., ova fertilized at 6 to 8 d postmating, 80.0%) was not affected by ovulation rate. The percentage of fetuses developing normally at 51 to 53 d postmating decreased (P less than .01) as ovulation rate increased; 1 CL, 100.0%; 2 CL, 100.0%; 3 CL, 66.7%; 4 CL, 45.8%; 5 CL, 33.3%; 6 to 10 CL, 13.6%; and greater than 10 CL, 8.9%. Of the 86 cows permitted to calve, 47 produced singles, 22 twins, 9 triplets, 7 quadruplets, and 1 quintuplets. Calf birth weight and gestational length decreased (P less than .01) as the number of calves born increased from one to two to three. Smaller decreases (P less than .05) in birth weight occurred among triplets, quadruplets, and quintuplets, whereas gestational length did not differ (P greater than .1) among these groups. Systemic progesterone concentrations in the dam were proportional (P less than .01) to the number of fetuses in utero between d 126 and 266 for dams gestating one, two, or three or more fetuses; estrone sulfate was lower (P less than .01) in dams with one than in those with two or more fetuses. Placental weight (i.e., cotyledons plus intercotyledonary membranes) per fetus at 52 +/- 1 d of gestation and at term decreased as the number of fetuses increased. The chorioallantoic membranes were often fused among multiple fetuses and contained either all viable or all dead fetuses, but not both, within the same anastomosed placental unit. These results suggest that ovulation rate is the first limiting factor to increasing cow productivity for beef cattle because some bovine females had the capacity to gestate up to three fetuses per uterine horn, or a total of five fetuses, above which pregnancy was terminated.

The relationship between ovulation response and systemic progesterone concentration in steroid immunised heifers

The relationship between ovulation response and systemic progesterone concentration in steroid immunised heifers

Systemic progesterone concentration following human chorionic gonadotropin administration at various times during the estrous cycle in beef heifers.

In Trial 1, 26 heifers were allotted randomly to a control group or one of four groups to receive a single injection of 3,000 IU hCG on d 1, 4, 7 or 10 of an estrous cycle. Heifers next completed a nontreated cycle, and at their third estrus were reallotted to one of the five groups described previously. Estrous cycle length was extended in cycle 1 but was not altered during the nontreated cycle or in cycle 3. Administration of hCG on d 4 or 7 increased (P less than .05) mean serum progesterone (P4) over the first 16 d of the estrous cycle by .9 and .8 ng/ml, respectively. In Trial 2, 23 heifers were allotted randomly to one of two groups to receive a placebo or a single injection of 3,000 IU hCG on d 4 of an estrous cycle. Heifers were inseminated artificially at subsequent estrus. On d 4 postbreeding, treatments were repeated. Administration of hCG on d 4 of the prebreeding estrous cycle increased (P less than .05) mean P4 over the first 16 d by .9 ng/ml, whereas mean P4 over the first 16 d postbreeding was not affected by a second injection of hCG on d 4 postbreeding. Administration of hCG increased (P less than .05) first-service pregnancy rate (92 vs 55%). In conclusion, progesterone concentrations were enhanced by hCG given on d 4 or 7 of the estrous cycle, and pregnancy rate was increased when hCG was administered both on d 4 of the prebreeding cycle and d 4 postmating.

Influence of SLA haplotype on ovulation rate and litter size in miniature pigs

Systemic blood was collected from and surgery performed on sows of 3 strains of miniature swine bred for specific SLA (swine MHC) haplotypes (a, c and d) from Day 2 to Day 6 after mating (first day of mating = Day 0). Ovulation rate was determined by counting corpora lutea and embryos were flushed from the uterus. Progesterone, oestradiol-17 beta and oestrone were quantitated in blood plasma and uterine flushings by RIA. SLAd/d females had a higher ovulation rate than SLAa/a or SLAc/c females (11.50 +/- 0.87 vs 9.11 +/- 0.68 and 8.17 +/- 0.83, respectively; P less than 0.01). Oestrone was higher than oestradiol-17 beta in systemic plasma (56.5 +/- 6.4 vs 33.0 +/- 4.7 pg/ml, P less than 0.01) while oestradiol-17 beta was higher than oestrone in uterine flushings (19.8 +/- 1.4 vs 14.9 +/- 1.5 pg/horn, P less than 0.10). Systemic progesterone concentration was correlated with day after mating (r = 0.93, P less than 0.01). There was no effect of haplotype on any of the hormone concentrations measured. Litter size was analysed from 99 matings amongst SLAa/a, SLAa/c, SLAa/d, SLAd/c and SLAd/d sires and dams. Litter size from -/d and d/d sows or from d/d boars were larger (P less than 0.05) than for all other matings. Although ovulation rate was higher in SLAd/d sows, the significant effect of sire SLA genotype on litter size suggests an additional effect of the d haplotype on embryonic survival.

Reproduction in the vervet monkey (Cercopithecus aethiops): endometrial oestrogen and progestin receptor dynamics during normal and prolonged menstrual cycles.

Sixteen individually caged adult female vervet monkeys (Cercopithecus aethiops), whose reproductive parameters had been studied for 5 years, were hysterectomized/ovariectomized during three reproductive states; i.e. the late follicular (15.4 +/- 4.7 (S.D.) days) and luteal (27.8 +/- 4.7 days) phases of the normal cycle (20-50 days), and during prolonged intermenstrual intervals (PII; 99.0 +/- 2.5 days since the previous menses). These latter animals showed characteristics of both follicular and luteal phases; i.e. their ovaries contained both corpora lutea and large antral follicles and systemic oestradiol and progesterone concentrations were raised. Analysis of cytoplasmic oestrogen and progestin receptors (CER and CPR) revealed that endometrium during PII had CER levels of 0.58 +/- 0.07 pmol/mg protein, similar to those of the follicular phase (0.60 +/- 0.12); CPR levels (1.20 +/- 0.87) were not different from those of the luteal phase (1.05 +/- 0.45). The ratio of CPR to CER during the luteal phase was about tenfold higher than that of the follicular phase. Levels during PII were intermediate between the two phases. Under receptor-activating conditions, the DNA-binding components of the PII cytoplasmic fraction underwent over 40% loss while those present during both phases of the normal cycle doubled. The hormone-binding sites at all times remained intact indicating that the DNA and hormone-binding sites are distinct on both CER and CPR. Less than 50% interaction of CER/CPR with DNA-cellulose was obtained, indicating the presence of only limited quantities of cytoplasmic activating factors which may be a prerequisite for receptor binding to the genome. During PII, factors which deactivate DNA-binding sites may also have been induced. Extensive accumulation of nuclear oestrogen receptor was evident in PII endometrium with 80% being salt-resistant. This level is higher than that in the follicular and luteal phases (37 and 52% respectively). These data, suggesting a possible aberration of receptor activation in vitro and receptor processing in vivo, may be indicative of endometrial dysfunction during PII. This could lead to a delay in menstruation.

Evidence for uterine metabolism of progesterone during early pregnancy in the pig

Two experiments were conducted to examine whether the 40 or 50% decrease in systemic progesterone (P 4) concentrations between Days 13 and 21 postmating in the pig results from decreased ovarian P 4 secretion or increased uptake of P 4 by the uterus. In Experiment I, five nonpregnant (NP) and four pregnant (P) gilts were sham-operated, and five NP gilts were hysterectomized (HYST) on Days 7 to 9 postestrus or postmating (first day of estrus or mating = Day 0). Femoral arterial blood was obtained once daily from Day 10 until the subsequent estrus (NP gilts) or Day 21 (P and HYST gilts). In Experiment II, blood was collected daily from both utero-ovarian veins of two NP and three P gilts from Days 11 to 18. Femoral arterial P 4 concentrations were similar for all gilts in Experiment I from Days 10 to 14. For NP gilts, femoral arterial P 4 declined (P < 0.01) after Day 14 to reach basal levels by Day 17. Progesterone in femoral arterial blood of P gilts declined (P < 0.01) from Days 13 to 16 and then remained constant through Day 21. Concentrations of P 4 in femoral arterial blood of HYST gilts remained constant from Days 13 to 21 and were greater (P < 0.01) than for P gilts from Days 15 to 21. In Experiment II, P 4 concentrations in utero-ovarian venous blood were similar until Day 14 between NP and P gilts. Utero-ovarian P 4 of NP gilts then declined (P < 0.01) to reach basal levels by Day 16. P 4 concentrations in utero-ovarian venous blood of P gilts increased (P < 0.05) for Days 14 to 18. These results demonstrate that ovarian P 4 secretion increases during early pregnancy in the pig. Further, the absence of a decline in P 4 concentrations in femoral arterial blood of HYST gilts suggests that the declining systemic P 4 levels observed during early pregnancy are a result of uterine uptake and(or) metabolism.

Effects of prostaglandin E 1 or E 2 (PGE 1; PGE 2) on luteal function and binding of luteinizing hormone in nonpregnant ewes

Two studies were conducted to determine the effects of PGE 1 or PGE 2 on luteal function and binding of luteinizing hormone (LH) to luteal cell membranes in nonpregnant ewes. In Study I, ewes (n=5 per group) received an injection of vehicle (VEH) or 333 μg of PGE 1 or PGE 2 into the tissue surrounding the ovarian vascular pedicle (intrapedicle) on day 7 postestrus. Systemic progesterone concentrations of PGE 1-treated ewes were greater (P<0.01) than those of VEH-treated ewes at 24 and 48 hr after injection. For PGE 2-treated ewes, progesterone concentrations were greater (P<0.01) than for VEH-treated ewes only at 24 hr. Neither PGE 1 nor PGE 2 affected luteal weights or LH binding capacity at 48 hr. Treatment with PGE 1, however, increased (P<0.10) endogenously bound LH at this time. In Study II, ewes (n=5 per group) received an intrapedicle injection of VEH, or 10 mg of PGE 1 or PGE 2 on day 8 postestrus. Systemic progestrone concentrations in PGE 1-treated ewes were less (P<0.01) than for VEH-treated ewes at 24 hr, but by 72 hr were not different from those of VEH-treated ewes. For PGE 2-treated ewes, systemic progesterone declined steadily to reach low values by 72 hr. Prostaglandin E 2 had no effect on luteal binding of LH at 72 hr, whereas PGE 1 increased (P<0.05) LH binding capacity and endogenously bound LH. Although PGE 2 had no apparent effect on luteal binding of LH in these studies, PGE 1 may enhance the function of ovine corpora lutea by stimulating an increase in their binding of LH and capacity to bind LH when the CL receives a luteolytic signal.

Evidence for an oscillator other than luteinizing hormone controlling the secretion of progesterone in cattle.

The objective of this study was to quantify and compare the frequencies of pulses in ovarian and systemic concentrations of progesterone, systemic concentrations of luteinizing hormone (LH) and rate of ovarian blood flow. Blood was collected simultaneously from previously implanted catheters in the ovarian venous circulation and jugular vein on Day 12 or 13 of estrous cycles from 4 nonlactating dairy cows. Blood was collected at a rate of 2.5 ml/min for 5 min out of every 10 min over an 8-h period. The mean rate of blood flow in the ovarian artery during the 5-min collection period was estimated by an electromagnetic blood flow transducer. Pulses were observed over time in both ovarian and systemic concentrations of progesterone at frequencies that ranged between 0.625 and 0.875 cycles/h (1.1 to 1.5 h/cycle) among the animals. Only one or two episodes of release of LH were observed during the 8-h period, and transient increases in blood flow to the ovaries were associated temporally with each episode of LH release. The estimated frequencies for release of LH and increased blood flow were the same for each animal and ranged between 0.250 and 0.375 cycles/h. A second cycle with a frequency similar to that for LH was evident in the spectral density functions for ovarian and systemic concentrations of progesterone. This cycle was eliminated when the cycle for LH was removed from the data for progesterone, but the magnitude and frequency of the pulses in progesterone were not affected.(ABSTRACT TRUNCATED AT 250 WORDS)

Conception, embryonic development and corpus luteum function in beef cattle open for two consecutive breeding seasons

High-fertility (control cows) and low-fertility (cows and heifers not pregnant after two consecutive breeding seasons — twice-open) cyclic bovine females were treated with a single injection of 1000 IU of human chrionic gonadotropin (HCG) or 100 μg of gonadotropin releasing hormone (GnRH) to enhance and/or hasten corpus luteum formation and progesterone secretion, and improve conception rate in the low-fertility females. Hormone treatments were administered to 38 parous control cows, 34 twice-open parous cows, and 27 twice-open nonparous heifers immediately after natural mating by a fertile bull. Blood samples were collected on Days 3, 6, 9, 12, and 18 after mating for determination of systemic progesterone concentrations. Pregnancy rate at necrospy approximately 33 days after mating (range 31–37) was higher in control cows (73.0%) than in twice-open cows (48.4%; P<0.05) or twice-open heifers (34.6%; P<0.01). Pregnancy rate was not affected by the HCG or GnRH treatment. The HCG treatment increased plasma progesterone concentrations in twice-open heifers but not in control or twice-open cows. Progesterone was unaffected by the GnRH treatment. Systemic progesterone concentrations were higher in control than in twice-open females but did not differ between pregnant and nonpregnant females of Days 3, 6, 9 and 12 after mating. Enhanced gonadotropin stimulation at estrus by injection of either HCG or GnRH did not increase pregnancy rate or systemic progesterone concentrations (except in HCG-treated twice-open heifers) in low- or high-fertility females. Lower pregnancy rates in twice-open females were not associated directly with the lower systemic progesterone concentrations.

Doses of prostaglandin F 2α effective for induction of parturition in goats

Twelve mixed breed does were injected with different doses of prostaglandin F 2α (PGF 2α) or saline on day 144 of gestation. Four each received single intramuscular injections of 5.0 or 2.5 mg PGF 2α, or 1.0 ml saline (controls). Systemic progesterone (P 4) concentrations were determined daily from day 144 until the day of kidding. Does receiving 5.0 mg PGF 2α, 2.5 mg PGF 2α, or saline kidded within mean (± SD) hours and range (hours) of 35 ± 8.6 and 28–48, 43 ± 11.8 and 29–57, and 111 ± 79.1 and 41–200, respectively. Mean (± SD) concentrations of P 4 (ng/ml) on the day of injection and on day 1 postinjection were 5.2 ± 2.6 and 0.7 ± 0.9, 5.3 ± 2.2 and 1.1 ± 1.0, and 6.4 ± 3.9 and 4.1 ± 2.6 for does receiving 5.0 mg PGF 2α, 2.5 mg PGF 2α, or saline, respectively. It was concluded that 5.0 mg and 2.5 mg PGF 2α effectively shortened the interval from injection to parturition, but that this interval was not as predictable as that previously reported with 20 mg PGF 2α.

A luteolytic interaction between estradiol benzoate and prostaglandin F2 alpha in cattle.

A possible luteolytic interaction between prostaglandin F2 alpha (PGF2 alpha) and estradiol benzoate (E2B) in cattle was investigated by randomly assigning 20 heifers to one of four groups of a 2 x 2 factorially designed experiment. The treatments for the groups consisted of im administration of: 1) 200 micrograms of E2B, given twice daily on d 10, 11 and 12 of the estrous cycle plus 7 mg PGF2 alpha, given at a separate site, but concurrently with the last injection of E2B; 2) the vehicles for E2B (sesame oil) and PGF2 alpha (saline); 3) E2B and saline and 4) sesame oil and PGF2 alpha. Administration of both E2B and 7 mg PGF2 alpha resulted in luteolysis as evidenced by a shorter mean length of the estrous cycle (P less than .05) when compared with the vehicle-treated control group, and a decline in systemic concentrations of progesterone to less than 1 ng/ml in four of five animals. Administration of PGF2 alpha with sesame oil was luteolytic in only one of five animals and E2B plus saline had no effect on the mean length of the estrous cycle or systemic concentrations of plasma progesterone. These observations suggested a luteolytic interaction between E2B and PGF2 alpha. This luteolytic interaction was examined further in a second experiment. Corpora lutea was removed 12 h after treatment with 200 micrograms E2B or .5 ml sesame oil, administered twice on d 10, 11 and 12 of the cycle, and incubated with luteinizing hormone (LH) and PGF2 alpha in vitro. The effect of both LH and PGF2 alpha was to increase progesterone synthesis (P less than .01), and this effect was observed irrespective of E2B pretreatment. These findings were not consistent with the hypothesis that the luteolytic site of action for either E2B or PGF2 alpha was inhibition of the steroidogenic effects of LH as assessed under in vitro conditions.

Progesterone-induced hyperventilation in the guinea pig

Progesterone-induced hyperventialtion has thus far not been reported in animals other than humans. Accordingly we investigated the effects of chronic progesterone treatment on ventilation in guinea pigs. Virgin female guinea pigs were divided into those treated with cholesterol as controls (C), progesterone (P), or estrogen plus progesterone (E&P). All hormones were administered in silastic capsules placed subcutaneously for 28 days. On day 28 arterial blood gases and respiratory rates were determined. Arterial pH was elevated in P- and E&P-treated animals compared with controls (7.43 ± 0.04 and 7.43 ± 0.03 vs 7.39 ± 0.05, respectively, P < 0.05). Arterial P CO 2 was reduced in both progesterone-treated groups compared to controls ( 33.9 ± 5.8 ( P), 33.1 ± 2.8 ( E&P), 38.0 ± 4.3 ( C) ), and was correlated with systemic progesterone concentrations in animals receiving E&P (r = −0.85, P < 0.01) but not in animals receiving P alone (r = −0.07, P = N.S.). Arterial [HCO 3 −], P o 2 and respiratory rates were not different between groups. We conclude that chronic progesterone administration produces hyperventilation in guinea pigs, and that estrogen facilitates this action of progesterone.

Doses of prostaglandin F 2α effective for induction of estrus in goats

A total of 11 cycling does weighing between 24 and 50 kg were injected with varying dosages of prostaglandin F 2α (PGF 2α) between 7 and 10 days into each estrous cycle. Five injections each of 1.25, 2.5, 5.0, or 7.5 mg PGF 2α were alternated with five injections of 1.0 ml saline. Saline treated does served as controls. All does were teased twice daily with a buck and observed for signs of estrus for 5 days post-injection. Daily systemic concentrations of progesterone (P 4) were determined by radioimmunoassay. The mean (± S.E.) hours from injection to estrus was 47 ± 3.3, 42 ± 4.3, 44 ± 8.5, and 43 ± 5.5 for does receiving 1.25, 2.5, 5.0, and 7.5 mg PGF 2α, respectively. None of the does receiving saline exhibited estrus in the 5-day post-injection observation period. Mean (± S.E.) concentrations of systemic P 4 in all does on the day of injection was 4.22 ± 0.45 ng/ml. Concentrations 24 hours post-injection were 0.21 ± 0.02, 0.15 ± 0.05, 0.17 ± 0.04, 0.16 ± 0.04, and 4.5 ± 1.36 ng/ml for does receiving 1.25, 2.5, 5.0, and 7.5 mg PGF 2α, and 1.0 ml saline, respectively. The results suggested that 1.25 mg PGF 2α was effective for induction of estrus in the cycling goat.

Blood flow to the corpus luteum-bearing ovary and ipsilateral uterine horn of cows during the oestrous cycle and early pregnancy.

Total ovarian blood flow of cows (sum of flows through the ovarian artery and uterine branch of the ovarian artery), measured by electromagnetic flow probes, followed a consistent cyclic pattern, being highest from Day 10 of the oestrous cycle (oestrus = Day 0) until 4 days before the subsequent oestrus. During this period of high ovarian blood flow, blood of uterine origin flowed to the ovary through the uterine branch of the ovarian artery and was found to comprise 20-40% of the ovarian blood flow. Between Days -4 and -3, ovarian blood flow dropped (P less than 0.01) from 3.2 +/- 0.5 to 1.0 +/- 0.4 ml/min, coincident with a reversal of flow through this anastomosis. Uterine arterial blood flow was highest (P less than 0.05) from 1 day before oestrous to the day of oestrus when compared to the remaining days of the oestrous cycle. Throughout the oestrous cycle, ovarian blood flow was positively correlated with systemic concentrations of progesterone (r = 0.55, P less than 0.01) and negatively correlated with systemic concentrations of oestradiol-17 beta (r = -0.42, P less than 0.01). In contrast, uterine arterial blood flow was negatively correlated with systemic progesterone (r = -0.25, P less than 0.05) and positively correlated with systemic oestradiol (r = 0.45, P less than 0.05). These data provide direct evidence in the cow that a portion of the blood flowing to the ovary containing a fully functional corpus luteum is contributed by the ipsilateral uterine artery.