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1.
Opioids were found as factors affecting porcine ovarian steroidogenesis. The mechanism of opioid action, however, on porcine theca interna cells is completely unknown. Therefore, the present study was designed to investigate the possible involvement of two intracellular pathways, phospholipase C/protein kinase C and adenylyl cyclase/protein kinase A, in opioid signal transduction in porcine theca cells treated with mu opioid receptor agonist, FK 33-824. Incubation of the cells for 4 h with FK 33-824 at the dose 1 nM resulted in decreases in inositol phosphate accumulation as well as androstenedione (A(4)), testosterone (T), and estradiol (E(2)) secretions. Protein kinase C (PKC) inhibitors, staurosporine (1-100 nM), D-sphingosine (10-500 nM), and PKCi (100-2000 nM), both added alone and together with the opioid agonist, depressed release of the steroid hormones. PKC activator, phorbol ester (PMA, 1-100 nM), used alone was without effect on theca cell steroidogenesis, but added in combination with FK 33-824 abolished inhibitory influence of the opioid on A(4), T, and E(2) output. The steroid hormone secretion by PKC-deficient theca cells was inhibited by the opioid agonist. FK 33-824 also suppressed PKC activity reducing [(3)H]PDBu specific binding to theca cells, whereas ionomycin (a positive control) increased labeled phorbol ester binding to the cells. In the next experiment, cAMP release from theca cells during 2 and 4 h incubations with FK 33-824 (1-100 nM), naloxone (10 microM; opioid receptor antagonist), and LH (100 ng/mL; a positive control) was examined. FK 33-824 at the dose 1 nM inhibited cAMP secretion during 2 h incubation, but had no effect during longer incubation. LH in a manner independent on incubation time multiplied cAMP release. Protein kinase A inhibitor, PKAi (100-2000 nM), alone and in combination with FK 33-824 (1 nM), inhibited A(4), T, and E(2) secretions by theca cells. PKA activator, 8BrcAMP (10-1000 microM), stimulated the steroid hormone release, but this stimulatory effect was diminished in the presence of FK 33-824. The results allow to suggest that opioid peptides affect porcine theca cell steroidogenesis and their acute action on the cells is connected with the inhibition of phospholipase C/protein kinase C and adenylyl cyclase/protein kinase A signal transduction systems.  相似文献   

2.
A study was conducted to characterize steroidogenesis in small ovarian follicles (1–10 mm in diameter) of the hen. The aims of our study were: 1) to determine basal estradiol-17β (E2) production by different sizes of small follicles; 2) to determine the ability of intact small follicles to utilize exogenous substrates for testosterone (T) and E2 production; and 3) to investigate the preferred steroidogenic pathway in small follicles. Small follicles which had not entered the hierarchy were isolated from ovaries obtained 2 hr after oviposition and divided into three groups: small white follicles (SWF; 1–2 mm in diameter), large white follicles (LWF; 2–4 mm in diameter), and small yellow follicles (SYF; 5–10 mm in diameter). Yolk and granulosa cells were removed from LWFs and SYFs and the remaining theca layer was called a follicle shell. Intact follicles or follicle shells (4/4 ml/tube) were incubated in avian Ringer's buffer supplemented with 10 mM HEPES and 0.1% BSA at 37°C for 3 hr with various treatments. Testosterone and E2 were measured in the medium. The SYFs and their corresponding follicle shells produced the greatest amount of E2 when E2 production was expressed per follicle. Addition of 2 mM 8-Br-cAMP to the incubation medium stimulated E2 production by all sizes of follicles and follicle shells. However, follicle shells produced lower basal- and 8-Br-cAMP-stimulated E2 secretion compared to corresponding intact follicles. There was no significant difference in E2 production in response to various concentrations of 25-hydroxycholesterol (25-OH-CHOL; 0–100 μM) by intact follicles and follicle shells. On the contrary, intact follicles and follicle shells produced T and E2 in a dose-dependent manner in response to increasing concentrations (0–100 μM) of pregnenolone (P5). Intact follicles also used progesterone (P4) and dehydroepiandrosterone (DHEA) as substrates for T and E2 production. DHEA was the preferred substrate for steroid production compared to P4. In summary, we found that: 1) steroidogenesis in small follicles is regulated by a cAMP-dependent protein kinase A second messenger system; 2) adequate amounts of endogenous cholesterol are available for steroidogenesis; and 3) both Δ5 and Δ4 pathways are functional in small follicles and the Δ5 pathway may be the preferred steroidogenic pathway. pathway.  相似文献   

3.
Oxidized(OX)-low density lipoprotein (LDL) inhibits steroidogenesis by luteal cells (LC) from regressing porcine CL. The present study was designed to investigate the mechanism of inhibition by determining whether OX-LDL inhibits basal and agonist-stimulated cAMP production in regressing LC. Collagenase-dispersed porcine LC (n = 7 animals, estrous cycle Day 12-15) were cultured (2.5 x 10(5) cells/0.5 ml) in serum-free DMEM/Hams F-12 in duplicate wells at 37 degrees C. Approximately 18 hr after plating, media were replaced and LC were immediately treated with human LDL (0, 25, or 100 microg/ml) or OX-LDL (25 or 100 microg/ml). LC were incubated for 2 hr before addition of isobutylmethylxanthine (IBMX) to inhibit phosphodiesterase activity, immediately followed by hCG (100 ng/ml), cholera toxin (CT; 0.1 microM), forskolin (FS; 50 microM), or no further treatment (controls). LC were incubated for an additional 90 min. After removal of culture media, cells were extracted with 0.1 N HCl. Cell extracts were assayed for cAMP by enzyme immunoassay (EIA). HCG, CT, and FS increased (P < 0.05) cAMP production approximately four-, 10-, and 25-fold, respectively, relative to controls. OX-LDL (25 and 100 microg/ml) inhibited (P < 0.05) cAMP production by unstimulated, hCG-, and CT-stimulated LC, but not that by FS-stimulated LC. The highest concentration of OX-LDL (100 microg/ml) reduced cAMP formation by 39.8 +/- 6.6%, 44.7 +/- 10.5%, and 67.7 +/- 4.5% in unstimulated, hCG-, and CT-stimulated LC, respectively. In contrast, unmodified LDL (25 and 100 microg/ml) did not alter cAMP production. We conclude that OX-LDL can interfere with the cAMP signaling pathway in regressing luteal cells by acting at sites proximal to adenylate cyclase activation.  相似文献   

4.
Some functional properties of highly enriched turkey poult adrenocortical cells were characterized. Cells were incubated with various mammalian and avian ACTH analogues, 8-Br-cAMP, and 25-hydroxycholesterol for 2 hr. Corticosterone production and, where appropriate, cyclic AMP (cAMP) production were measured by radioimmunoassay. Human ACTH-(1-24) was the most efficacious and potent ACTH analogue for stimulating corticosterone and cAMP production, whereas turkey ACTH-(1-39) was among the least efficacious and least potent analogues. Maximal corticosterone production induced by 8-Br-cAMP and supported by 25-hydroxycholesterol was 67-109% greater than that induced by ACTH analogues. The data suggest that intracellular concentrations of cAMP-dependent factors and steroidogenic enzymes exceed those which are accessible to ACTH-activated cellular processes. In addition, there were sex-dependent contrasts in some functional parameters, despite the immature status of the birds. Basal corticosterone production of female cells was 19% greater than that of male cells, albeit maximal ACTH analogue-induced corticosterone production was not different between male and female cells. In contrast, maximal 8-Br-cAMP-induced and 25-hydroxycholesterol-supported corticosterone production of male cells were, respectively, 72 and 45% greater than that of female cells, thus suggesting greater intracellular concentrations of protein kinase A-dependent factors and steroidogenic enzymes in male cells compared to female cells. However, maximal ACTH-induced cAMP production of female cells was 21% greater than those of male cells, thus suggesting a compensatory mechanism in female cells. In addition, there were sex-dependent differences in sensitivity to ACTH as indicated by corticosterone and cAMP responses to ACTH analogues: sensitivity to ACTH as indicated by corticosterone and cAMP responses to ACTH analogues: sensitivity of male cells was 1.2-3.2 times that of female cells. A sex-dependent difference in ACTH-cell interaction, possibly including ACTH receptors, is implicated since there were no sex differences in cellular sensitivities to 8-Br-cAMP and 25-hydroxycholesterol. These data indicate that the turkey adrenal gland is amenable for the preparation of isolated adrenocortical cells that have functional integrity. Thus, turkey adrenocortical cells expand the in vitro repertoire for elucidating the regulatory mechanisms of avian adrenocortical function.  相似文献   

5.
Effects of various selective phosphodiesterase (PDE) inhibitors on muscle contractility and cyclic nucleotide contents in guinea pig taenia coli were investigated. Forskolin and sodium nitroprusside inhibited carbachol (CCh)-induced contraction in a concentration-dependent manner. Various selective PDE inhibitors, vinpocetine (type 1), erythro -9-(2-hydroxy-3-nonyl)adenine (EHNA, type 2), milrinone (type 3), Ro20-1724(type 4) and zaprinast (type 5) inhibited CCh-induced contraction in a concentration-dependent manner, but the inhibition of milrinone was noticeably smaller than that of the other PDE inhibitors. The rank order of potency was zaprinast > vinpocetine > EHNA > Ro20-1724 > milrinone. In the presence of CCh (0.3 microM), vinpocetine and Ro20-1724 both increased cAMP content, but not cGMP. By contrast, EHNA and zaprinast both increased cGMP content, but not cAMP. Pretreatment with ODQ (30 microM), a soluble guanylyl cyclase inhibitor, decreased the inhibition of CCh-induced contraction by EHNA or zaprinast. Pretreatment with SQ22536 (100 microM), an adenylyl cyclase inhibitor, decreased the inhibition of CCh-induced contraction by vinpocetine or Ro20-1724. In conclusion, it was indicated that vinpocetine- or Ro20-1724-induced relaxation was correlated with cAMP but EHNA- or zaprinast- induced relaxation was correlated with cGMP.  相似文献   

6.
Luteinizing hormone (LH)-stimulated steroidogenesis in luteal cells is known to be mediated through the activation of cyclic AMP (cAMP)-dependent protein kinase, and to be also modulated by calcium-dependent mechanisms. In the present study, we tested the hypothesis that LH stimulates progesterone (P4) production in bovine luteal cells through activation of phospholipase (PL) C by using a cell culture system. Bovine mid-luteal cells (Days 8-12 of the estrous cycle) were cultured for 24 h and then exposed to a PLC inhibitor (U-73122; 10 microM) with or without LH (10 ng/ml) for 4 h. U-73122 blocked LH-stimulated P4 production without affecting cAMP accumulation. Moreover, exposure of luteal cells to PLC increased P4 production in a dose-dependent manner. These results support the hypothesis that the luteotropic action of LH in bovine luteal cells is mediated not only by activation of adenylate cyclase but also by activation of PLC.  相似文献   

7.
Structural luteolysis occurs by apoptosis of luteal cells. The present study examined the effects of activators of well-characterized second messengers on Fas and caspase-3 mRNA expression and on P4 production in luteal cells in order to trace the pro- and anti-apoptotic factors in the bovine corpus luteum (CL). Cultured bovine mid luteal cells were treated for 24 h with a cyclic AMP analogue (8-bromo cyclic AMP; 8br-cAMP; 2.5 mM), a protein kinase C (PKC) activator (phorbol 12-myristate 13-acetate; PMA; 10 microM), or calcium ionophore (A23187; 10 microM). Fas and caspase-3 mRNA expression was inhibited by 8br-cAMP and PMA but was increased by A23187 (P<0.05). In addition, P4 production by bovine luteal cells was stimulated by 8br-cAMP and PMA, whereas it was inhibited by A23187, compared with untreated controls (P<0.05). The overall results suggest that cAMP and PKC suppress apoptosis in bovine luteal cells through inhibition of Fas and caspase-3 mRNA expression and through stimulation of P4 production. Therefore, substances that activate cAMP or PKC may act as survival factors in the bovine CL. Furthermore, substances that mobilize Ca2+ may act as apoptotic factors by stimulating Fas and caspase-3 expression in the bovine luteal cells.  相似文献   

8.
Two experiments were conducted to determine the effects of pregnenolone (P5), cyclic adenosine monophosphate (cAMP), and human chorionic gonadotropin (hCG) on porcine placental and endometrial production of progesterone (P4) and estrone (E1) in vitro at days 30, 60 and 90 of gestation. Placental P4 production increased between days 30 and 90 and was enhanced by the addition of P5. A further increase in placental P4 production occurred at days 30 and 90 due to cAMP supplementation. Addition of hCG failed to increase placental P4 production at any day. Placental E1 production in vitro was biphasic and mimicked the pattern seen in maternal plasma and fetal fluids. Placental E1 production in P5-supplemented medium was enhanced by the addition of cAMP at day 90. However, hCG supplementation reduced placental E1 production at day 90. Endometrial P4 and E1 production were similar to those of the placenta at day 30 of gestation. However, unlike placental steroidogenesis, endometrial hormone production remained relatively constant over the 3 days of gestation examined. Supplemental P5 enhanced endometrial P4 and E1 production. The overall magnitude of response to supplementation was considerably less in endometrial vs placental tissue. We conclude that both porcine placental and endometrial tissues are steroidogenically competent but that placenta is the far more active and responsive tissue. The mechanism controlling placental steroidogenesis apparently does not involve LH/hCG tropic stimulation, but cAMP is an effective intracellular second messenger.  相似文献   

9.
Background： Heifers not used as breeding stock are often implanted with steroids to increase growth efficiency thereby altering hormone profiles and potentially changing the environment in which ovarian follicles develop. Because bovine granulosa cell culture is a commonly used technique and often bovine ovaries are collected from abattoirs with no record of implant status, the objective of this study was to determine if the presence of an implant during bovine granulosa cell development impacts follicle stimulating hormone-regulated steroidogenic enzyme expression. Paired ovaries were collected from 16 feedlot heifers subjected to 1 of 3 treatments： non-implanted （n = 5）, Revalor 200 for 28 d （n = 5）, or Revalor 200 for 84 d （n = 6）. Small follicle （1 to 5 mm） granulosa cells were isolated from each pair and incubated with phosphate buffered saline （n = 16） or 100 ng/mL follicle stimulating hormone （n = 16） for 24 h. Results： Granulosa cells of implanted heifers treated with follicle stimulating hormone produced medium concentrations of progesterone similar （P = 0.22） to non-implanted heifers, while medium estradiol concentrations were increased （P 〈 0.10） at 28 and 84 d compared to non-implanted heifers indicating efficacy of treatment. Additionally, real-time PCR analysis in response to follicle stimulating hormone treatment demonstrated a decrease in steroidogenic acute regulatory protein （P = 0.05） mRNA expression in heifers implanted for 84 d and an increase in P450 side chain cleavage mRNA in granulosa cells of heifers implanted for 28 （P 〈 0.10） or 84 d （P 〈 0.05） compared to non-implanted females. However, no difference in expression of 3-beta-hydroxysteroid dehydrogenase （P= 0.57） and aromatase （P = 0.23） were demonstrated in implanted or non-implanted heifers. Conclusions： These results indicate follicles which develop in the presence of high concentrations of androgenic and estrogenic steroids via an implant tend to demonstrate an alter  相似文献   

10.
High composite somatic cell counts (CSCC) in dairy cows may develop into clinical mastitis (CM), suggesting that prevention or intervention of high CSCC may prevent CM later in lactation. The objective of this study was to quantify the relationship between high CSCC in dairy cows and the first subsequent case of CM in the same lactation. Farmer-diagnosed cases of CM and test day CSCC measurements during 1 year of 13,917 cows in 196 randomly selected Dutch dairy herds were available for analysis. Cows were followed in 1 lactation from the first test day postpartum until CM, drying off, culling or end of study. Cox proportional hazards models with time-varying CSCC levels were used to estimate the effect of high CSCC (≥200,000cells/ml) on the time until the first case of CM. A shared frailty effect was included to adjust for clustering of cows within herds. The proportion of cows developing CM after a CSCC measurement was 11%. Primiparae with a high CSCC had a 4-fold higher hazard for subsequent CM than primiparae with a low CSCC; multiparae with a high CSCC had a 2-fold higher hazard than multiparae with a low CSCC. Additionally, multiparae with a low CSCC had a 2-fold higher hazard for CM occurrence than primiparae with a low CSCC. Increasing the threshold for high CSCC showed that the risk for CM increased. If the last CSCC before CM was low, CSCC information of 2 preceding test days was more predictive than CSCC information from only the last test day. When the last CSCC was high, CSCC information of 2 preceding test days did not have added predictive value. This study identified that approximately 25% of first subsequent CM cases after a CSCC measurement can potentially be prevented when cows are prevented to get high CSCC or when high CSCC cows are removed from the population. This corresponded with a decrease in the proportion of lactating cows with CM after a CSCC measurement from 11% to 7%.  相似文献   

11.
Early follicular development is closely related to oocyte‐granulosa cells‐ovarian stromal cells/theca cells. The aim of the present study was to investigate the effects of ovarian cortical, medullary stromal and theca cells on oestradiol and progesterone biosynthesis, proliferation and apoptosis of goat ovary granulosa cells in vitro. Using Transwell coculture system, we evaluated steroidogenesis, cell proliferation and apoptosis, and some molecular expressions regarding steroidogenic enzyme, luteinizing hormone receptor and apoptosis‐related genes in granulosa cells. The results indicated that ovarian stromal/theca cells were able to stimulate oestradiol and progesterone production, promote cell proliferation and inhibit apoptosis of granulosa cells. Among all the three kinds of cells, theca cells affected strongly on granulosa cell function, and ovarian medullary stromal cells had the weakest effect on granulosa cells. These findings would provide an important knowledge of cell interaction among follicular cells during follicular development.  相似文献   

12.
Intra-ovarian factors, such as activin, are implicated in multiple aspects of follicular development in mammalian ovaries. This study was conducted to investigate a possible effect of activin-A on steroidogenesis in sheep granulosa cells in vitro. Sheep granulosa cells were obtained from medium antral follicles and cultured in a chemically defined RPMI -1640. Oestradiol and progesterone production, secreted by the cultured cells, was evaluated by enzyme-linked immunosorbent assay. In order to determine the dose effect of activin-A on steroidogenesis, granulosa cells were cultured in the presence of increasing concentrations of activin-A (0, 0.5, 5 and 50 ng ml(-1)) for 48 hours. The results revealed that activin-A exerts a differential effect on steroidogenesis in granulosa cells in such a way that it significantly (P < 0.05) suppressed progesterone production and enhanced oestradiol production. These results were confirmed by the time effect of activin-A on oestradiol and progesterone production in granulosa cells. In the absence of activin-A treatment, granulosa cells showed enhanced capacity to produce progesterone, but not oestradiol, as the time progressed from 12 to 48 hours. Treatment of sheep granulosa cells with 25 ng ml(-1)activin-A for 12, 24 and 48 hours significantly stimulated oestradiol production but inhibited progesterone production. These results suggest that activin-A is a local regulator of sheep folliculogenesis that might act to support differentiation in granulosa cells and suppress luteinisation.  相似文献   

13.
Gossypol, a polyphenolic aldehyde found in cottonseed, has been shown to perturb steroidogenesis in granulosa and luteal cells of rats, pigs and cattle. However, little is known about the direct effect of gossypol on theca cell functions in any species. The present study was conducted to investigate the effect of gossypol on the steroidogenesis and the expression of genes involved in it in cultured bovine theca cells. Theca cells were isolated from healthy preovulatory follicles and were cultured in the presence of luteinizing hormone (LH) for up to 7 days. During the culture period, main steroid products of the theca cells shifted from androstenedione (A4) at day 1 to progesterone (P4) from day 2 onward. At days 1 and 7, theca cells were treated with gossypol (0‐25 μg/mL) for 24 h. Gossypol inhibited LH‐stimulated theca cell A4 and P4 production in a dose‐dependent manner at both occasions. The viability of theca cells was not affected by gossypol at any doses used. Gossypol down‐regulated expressions of steroidogenic enzymes CYP11A1, HSD3B1 and CYP17A1, but not that of LHR. These results indicate that gossypol inhibits thecal steroidogenesis through down‐regulating gene expressions of steroidogenic enzymes but without affecting cell viability in cattle.  相似文献   

14.
The present investigation was undertaken to verify if the two nitric oxide synthase isoforms, eNOS and iNOS, are present in swine granulosa cells and whether the enzyme soluble guanylate cyclase is functionally active in the same cells and can account for NO effects. Using western blotting, the presence of endothelial NO synthase was demonstrated in freshly collected cells; on the contrary, iNOS expression was not observed in the same cells either before or after culture with the inflammatory cytokine hTNF-. The treatment with a strong NO donor (S-Nitroso-L-acetyl penicillamine, SNAP) determined an increase of cGMP levels in culture media, which was attenuated by the combined treatment with an inhibitor of NO-sensitive soluble guanylate cyclase, 1H-[1,2,3]oxadiaziolo [4,3a]quinoxaline −1-one (ODQ). The cGMP analog, 8 bromo-cGMP, mimicked the strong inhibitory effect exerted by SNAP on estradiol 17 β and progesterone production, while ODQ did not modify steroids concentrations in culture media. These observations demonstrate the presence of a follicular NO-generating system, which in swine granulosa cells seems to include only the endothelial NOS isoform. Furthermore, the nitric oxide/cyclic GMP system seems to be functionally active in these cells, since cGMP appears to mediate NO action, even if it cannot account completely for NO inhibitory effect on steroidogenesis.  相似文献   

15.
The present studies were conducted: (1) to determine which beta-adrenoceptor subtypes are involved in progesterone and oxytocin (OT) secretion, (2) to examine whether noradrenaline (NA) acts directly on the cytochrome P-450scc and 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD), and (3) to study the effect of prostaglandin F2 alpha (PGF2 alpha) on NA-stimulated steroidogenesis in luteal cells. The effect of NA on progesterone secretion from luteal slices of heifers on days 8-12 of the oestrous cycle was blocked by both atenolol (beta 1-antagonist) and ICI 118.551 hydrochloride (beta 2-antagonist). OT secretion was blocked only after treatment with ICI 118.551 hydrochloride (P < 0.05). Dobutamine (10(-4)-10(-6) M), a selective beta 1 agonist and salbutamol (10(-4)-10(-6) M), a selective beta 2 agonist, both increased progesterone production (P < 0.01) with an efficiency comparable to that produced by NA (P < 0.01). The increase of OT content in luteal slices was observed only after treatment with salbutamol at the dose of 10(-5) M (P < 0.01). Dobutamine had no effect on OT production at any dose. A stimulatory effect of NA on cytochrome P-450scc activity (P < 0.05) was demonstrated using 25-hydroxycholesterol as substrate. 3 beta-HSD activity also increased following NA (P < 0.01) or pregnenolone (P < 0.05) and in tissue treated with pregnenolone together with NA (P < 0.01). PGF decreased progesterone synthesis (P < 0.05) and 3 beta-HSD activity (P < 0.01) in tissue treated with NA. We conclude that NA stimulates progesterone secretion by luteal beta 1- and beta 2-adrenoceptors, while OT secretion is probably mediated only via the beta 2-receptor. NA also increases cytochrome P-450scc and 3 beta-HSD activity. PGF inhibits the luteotropic effect of NA on the luteal tissue.  相似文献   

16.

Background

Heifers not used as breeding stock are often implanted with steroids to increase growth efficiency thereby altering hormone profiles and potentially changing the environment in which ovarian follicles develop. Because bovine granulosa cell culture is a commonly used technique and often bovine ovaries are collected from abattoirs with no record of implant status, the objective of this study was to determine if the presence of an implant during bovine granulosa cell development impacts follicle stimulating hormone-regulated steroidogenic enzyme expression. Paired ovaries were collected from 16 feedlot heifers subjected to 1 of 3 treatments: non-implanted (n?=?5), Revalor 200 for 28 d (n?=?5), or Revalor 200 for 84 d (n?=?6). Small follicle (1 to 5 mm) granulosa cells were isolated from each pair and incubated with phosphate buffered saline (n?=?16) or 100 ng/mL follicle stimulating hormone (n?=?16) for 24 h.

Results

Granulosa cells of implanted heifers treated with follicle stimulating hormone produced medium concentrations of progesterone similar (P?=?0.22) to non-implanted heifers, while medium estradiol concentrations were increased (P?<?0.10) at 28 and 84 d compared to non-implanted heifers indicating efficacy of treatment. Additionally, real-time PCR analysis in response to follicle stimulating hormone treatment demonstrated a decrease in steroidogenic acute regulatory protein (P?=?0.05) mRNA expression in heifers implanted for 84 d and an increase in P450 side chain cleavage mRNA in granulosa cells of heifers implanted for 28 (P?<?0.10) or 84 d (P?<?0.05) compared to non-implanted females. However, no difference in expression of 3-beta-hydroxysteroid dehydrogenase (P?=?0.57) and aromatase (P?=?0.23) were demonstrated in implanted or non-implanted heifers.

Conclusions

These results indicate follicles which develop in the presence of high concentrations of androgenic and estrogenic steroids via an implant tend to demonstrate an altered capacity to respond to follicle stimulating hormone stimulation. Thus, efforts should be made to avoid the use of implanted heifers to study steroidogenesis in small follicle granulosa cell culture systems.  相似文献   

17.
Granulosa cells (GCs) play important roles in the regulation of ovarian functions, and in vitro culture is a relevant model for the study of steroidogenesis in ovarian follicles. Thus, growth factors secreted by the oocyte, like Growth and Differentiation Factor 9 (GDF9) and Bone Morphogenetic Protein 15 (BMP15), play an important part in the luteinization of granulosa cells. The aim of this work was to express GDF9 and BMP15 genes in bovine GCs in vitro and evaluate their effects on the luteinization process. Samples of culture medium and GCs transfected with GDF9 and BMP15 were obtained for 21 consecutive days to analyse the steroidogenic hormones' concentration (progesterone (P4) and estradiol (E2)) and the expression of STAR, GDF9 and BMP15 and their respective receptors. The results demonstrated an inhibitory effect of GDF9 and BMPF15 on P4 secretion in bovine GCs cultured in vitro. Moreover, our study demonstrated the entire expression of their respective receptors (TGFBR1, BMPR1B and BMPR2) and the inhibition of the steroidogenic marker, STAR gene. This work sheds light on a novel biological function of BMP15 and GDF9 in bovine GCs physiology, which could elucidate a non-described biological role for GDF9 and BMP15 in bovine granulosa cells' metabolism.  相似文献   

18.
The period of spring transition, from the anovulatory to the ovulatory season, is characterized in many mares by cyclical growth and regression of large dominant follicles. These follicles produce only low concentrations of estradiol and it is thought that acquisition of steroidogenic competence by large follicles during spring transition is prerequisite in stimulating LH prior to first ovulation. In situ hybridization was used to localize and quantify expression of factors that play a key role in follicular steroidogenesis: StAR, P450scc (CYP11A1), P450c17 (CYP17), P450arom (CYP19), and LH receptor (LHr). One ovary was obtained from mares on the day after detection of an actively growing 30 mm transitional anovulatory follicle (defined as the transitional follicle), and the remaining ovary was removed at the third estrus of the breeding season on the day after the preovulatory follicle reached 30 mm in diameter (defined as the preovulatory follicle). Messenger RNAs encoding StAR, CYP11A1, and CYP17 were detected only in theca cells and CYP19 mRNA was confined to the granulosa layer. There was significantly lower expression of mRNAs for the steroidogenic enzymes, StAR (P<0.001) and LHr (P<0.05) in transitional follicles than in preovulatory follicles. In conclusion, large equine follicles during spring transition have low levels of mRNA encoding steroidogenic enzymes, StAR and LHr which will contribute to the steroidogenic incompetence of dominant follicles during spring transition and their subsequent regression.  相似文献   

19.
Gossypol is a polyphenol isolated from the seed, roots and stem of cotton plant (Gossypium sp.) It has been associated with adverse effects on female reproduction, but recently also shown having promising effects against several malignancies. Its mechanisms of action are however still not fully understood. This study was therefore conducted to investigate the effect of 5 or 25 μg/mL gossypol on swine granulosa cell steroidogenic activity, redox status and Vascular Endothelial Growth Factor (VEGF) production. Study demonstrated that gossypol significantly (P < 0.001) inhibited granulosa cell estradiol 17β and progesterone production, an effect that could be at least partially mediated by an increase (P < 0.05) of nitric oxide and superoxide anion production as a consequence of superoxide dismutase inhibition. Moreover, gossypol stimulates (P < 0.001) VEGF production. In conclusion, study has demonstrated effecs of gossypol on swine granulosa cell function in vitro. Effects on female swine fertility can not be excluded.  相似文献   

20.
Bone morphogenetic proteins (BMPs) are emerging as a family of proteins crucial in the regulation of fertility and ovulation rate. We have shown that porcine theca cells express BMP receptors, however, there is a paucity of information regarding the effect(s) of BMPs on theca cell function. The purpose of this study was to investigate the effects of BMP-2 and -6 on theca cells cultured under serum-free conditions in terms of steroidogenesis, cAMP release and proliferation. The study was further extended to determine whether BMP responses in theca cells are affected by the addition of granulosa cells to the culture system. Both BMPs suppressed progesterone and androstenedione synthesis by theca cells (P < 0.05) after 144 h in culture. Oestradiol synthesis was suppressed (P < 0.05) by BMP-2, but not BMP-6, and theca cell proliferation was stimulated (P < 0.05) by BMP-6, but not BMP-2, after 144 h in culture. Both BMP-2 and -6 inhibited cAMP release (P < 0.05) by theca cells. Furthermore, progesterone and androstenedione synthesis by co-cultured theca and granulosa cells were suppressed (P < 0.05) whereas cell proliferation was stimulated (P < 0.05). These results provide strong evidence for a functional BMP system in the porcine ovary and that theca cells are responsive to BMPs in terms of steroidogenesis and proliferation. BMP-2 and -6 may have a role as luteinisation inhibitors in this polyovular species.  相似文献   

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