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1.
Polychlorinated biphenyls (PCBs) due to their lipophilic properties can be easily accumulated in animal and human body and elicit diverse effects causing impairment of reproductive processes. Since these compounds were not be able to affect directly the luteal steroidogenesis, the aim of the present study was to verify hypothesis that PCBs can impair the effect of LH on the secretory function of luteal cells. Bovine luteal cells from different stages of the oestrous cycle (days 1-5, 6-10, 11-15 and 16-18) were exposed for 72h to various congeners of PCBs (PCB 126, PCB 77 and PCB 153) at the doses of 1, 10 or 100 ng/ml, in the presence or absence of LH (100 ng/ml), to determine the possible effect of these compounds on progesterone (P4) and ovarian oxytocin (OT) secretion. Only PCB 77 on days 1-5 and 16-18 increased P4 secretion. All PCBs decreased LH-simulated secretion of P4 from luteal cells obtained from all days of luteal phase. Dioxin-like congener (PCB 126) inhibited (P<0.05) the most evidently LH effect on P4 secretion. All congeners, except the lower doses of PCB 126, increased (P<0.05) OT secretion. They can also increase LH-stimulated secretion of OT, but the effect was dependent on the congener used and on the phase of oestrous cycle. On days 1-5 and 10-15, PCB 126 diminished LH-stimulated effect on OT secretion from luteal cells. PCB 77 (mimickig both dioxin and estradiol effect) in the higher doses, amplified effect of LH-stimulated OT secretion, while on all other days it diminished LH influence. PCB 153, which has estrogen-like properties, amplified LH effect on OT secretion during all studied days of the cycle. We conclude that PCBs (supposedly via estrogen and arylhydrocarbon - AhR receptor) may directly affect LH-stimulated function of CL. This does not appear to be a direct adverse effect on luteal steroidogenesis, but rather indirect on OT secretion from or within CL.  相似文献   

2.
The current investigations were undertaken to study the mechanism of the adverse effect of phytoestrogens on the function of bovine granulosa (follicles >1< cm in diameter) and luteal cells from day 1–5, 6–10, 11–15, 16–19 of the oestrous cycle. The cells were incubated with genistein, daidzein or coumestrol (each at the dose of 1 × 10?6 m ). The viability and secretion of estradiol (E2), progesterone (P4) and oxytocin (OT) were measured after 72 h of incubation. Moreover, the expression of mRNA for neurophysin‐I/OT (NP‐I/OT; precursor of OT) and peptidyl‐glycine‐α‐amidating monooxygenase (PGA, an enzyme responsible for post‐translational OT synthesis) was determined after 8 h of treatment. None of the phytoestrogens used affected the viability of cells except for coumestrol. The increased secretion of E2 and P4 was only obtained by coumestrol (p < 0.05) from granulosa cells from follicles <1 cm in diameter and decreased from luteal cells on days 11–15 of the oestrous cycle, respectively. All three phytoestrogens stimulated (p < 0.05) OT secretion from granulosa and luteal cells in all stages of the oestrous cycle and the expression of NP‐I/OT mRNA in the both types of cells. The expression of mRNA for PGA was stimulated (p < 0.05) by daidzein and coumestrol in granulosa cells, and by genistein and coumestrol in luteal cells. In conclusion, our results demonstrate that these phytoestrogens can impair the ovary function in cattle by adversely affecting the synthesis of OT in follicles and in corpus luteum. However, their influence on the ovarian steroids secretion was less evident.  相似文献   

3.
We have previously shown that 3,3',4,4',5-pentachlorobiphenyl (PCB-126) vertically transferred from dams potentially exerts a direct effect on the ovaries of offspring and adversely affects female puberty. To investigate its toxicological targets in ovarian tissues, mRNAs encoding representative peptides that regulate follicular development in granulosa cells, theca cells, and oocytes were quantified using ovaries collected on postnatal days (PND) 5, 15, and 24 from the offspring of dams administered oral doses of 0, 1 or 3 microg/kg PCB-126 starting 2 weeks prior to mating and continuing until 20 days after delivery. Quantification using the real-time RT-PCR method revealed that PCB-126 lowered the amounts of mRNAs that encoded the inhibin alpha- and inhibin/activin beta A-subunits from PND 15 onwards; the amounts of mRNAs for inhibin/activin beta B-subunit, follicle-stimulating hormone (FSH) receptor, and aromatase on PND 15; and the amounts of luteinizing hormone receptor mRNA on PND 24 compared with those of the age-matched controls. In contrast, no differences were noted for mRNAs encoding c-kit, growth differentiation factor-9, bone morphogenetic protein-15, or kit ligand for any of the age groups examined. The serum FSH level on PND 24 was higher than that in the control. Since the earliest effects on the mRNAs in the rat ovaries were observed in those expressed in the granulosa cells of the growing follicles after the antral follicles had developed, molecules in granulosa cells but not in oocytes during the early stages of the antral follicles might be the primary targets of vertically transferred PCB-126.  相似文献   

4.
To determine if (1) levels of pregnancy-associated plasma protein-A (PAPP-A) mRNA and insulin-like growth factor binding protein (IGFBP) (-2, -3, -4 and -5) mRNAs differ between the dominant and subordinate follicles during the follicular phase of an estrous cycle, and (2) these differences are associated with differences in follicular fluid (FFL) concentrations of steroids (estradiol, androstenedione, and progesterone), total and free IGF-I, or IGFBPs, estrous cycles of non-lactating Holstein dairy cows (n = 16) were synchronized with two injections of prostaglandin (PGF2 alpha) 11 days apart. Granulosa cells and FFL were collected either 24 h or 48 h after the second injection of PGF2 alpha. FFL from dominant follicles had lower concentrations of progesterone (P < 0.08) and higher concentrations of estradiol (P < 0.05), androstenedione (P < 0.0001), estradiol:progesterone ratio (P < 0.0001), free IGF-I (P < 0.0001), and calculated percentage free IGF-I (P < 0.01) than large subordinate follicles. Levels of IGFBP-2, -4, and -5 in FFL were 3.0- (P < 0.05), 2.4- (P < 0.06), and 3.4-fold (P < 0.05) greater, respectively, in subordinate than in dominant follicles. IGFBP-3, IGFBP-4 and PAPP-A mRNA expression and IGF-II concentration did not differ (P > 0.10) between dominant or subordinate follicles. Levels of IGFBP-2 and -5 mRNA were severalfold greater (P < 0.05) in subordinate than dominant follicles. IGFBP-5 mRNA in granulosa cells decreased (P < 0.05) 62% to 92%, between 24h and 48 h post-PGF2 alpha. We conclude that decreased levels of IGFBP-2 and -5 mRNA in granulosa cells may contribute to the decrease in FFL IGFBP-2 and -5 protein levels of preovulatory dominant follicles, and that changes in granulosa cell IGFBP-3 and -4 mRNA and PAPP-A mRNA levels do not occur during final preovulatory follicular development in cattle.  相似文献   

5.
In cattle, sub-luteal circulating progesterone induces an increase in the frequency of LH pulses, prolonged growth of the dominant follicle, increased peripheral estradiol and reduced fertility. The objective of this study was to examine the earliest stages of development of prolonged dominant follicles, to gain insight into the etiology of this aberrant condition. Heifers were treated with an intravaginal progesterone-releasing device (CIDR) from Day 4-8 post-estrus and PGF2alpha was injected on Day 6 and again 12h later (early prolonged dominant group). Follicular phase (CIDR: Day 4-6, with PGF2alpha) and luteal phase (CIDR: Day 4-8, without PGF2alpha) groups served as controls. As expected, peripheral progesterone in heifers of the early prolonged dominant group was intermediate between luteal and follicular phase groups after luteal regression (P<0.05). On Day 7, the frequency of LH pulses was higher in heifers of the follicular phase and early prolonged dominant groups than the luteal phase group (P<0.05). Dominant follicles (n = 4 per group) were collected by ovariectomy on Day 8 and were similar in size among groups (P>0.05). Estradiol and androstenedione concentrations in the follicular fluid at ovariectomy were higher in the follicular phase and early prolonged dominant groups versus the luteal phase group (P<0.01), whereas progesterone did not differ among groups (P>0.05). Granulosa cells and theca interna isolated from dominant follicles were incubated for 3h with or without gonadotropins or frozen for later analysis of mRNA for steroidogenic enzymes. Luteinizing doses (128 ng/ml) of LH and FSH increased secretion of progesterone (P<0.05) but did not affect secretion of estradiol by granulosa cells in all groups. Low (2 or 4 ng/ml) and luteinizing doses of LH increased secretion of androstenedione by theca interna to a similar extent among groups. Expression of mRNA for P450 side chain cleavage (P450scc), 3beta-hydroxysteroid dehydrogenase (3beta-HSD), P450 aromatase (aromatase) and Steroidogenic Acute Regulatory (StAR) protein by granulosa cells did not differ among groups (P>0.05). Levels of mRNA for P450scc, 3beta-HSD, 17alpha-hydroxylase (17alpha-OH) and StAR protein in theca interna were similar in the follicular phase and early prolonged dominant groups (P>0.05), but lower in the luteal phase group (P<0.05-0.1). In summary, the premature follicular luteinization observed in previous studies after prolonged periods of sub-luteal progesterone was absent in early prolonged dominant follicles, exposed to sub-luteal progesterone for 36 h, and their characteristics resembled those of control follicles during the follicular phase.  相似文献   

6.
To determine the effect of gonadotropins on insulin- and insulin-like growth factor (IGF-I)-induced bovine granulosa cell functions, granulosa cells from bovine ovarian follicles were cultured for 2 days in the presence of 10% fetal calf serum (FCS), and then cultured for an additional 2 days in serum-free medium with added hormones. In the presence of 0 or 1 ng/mL of insulin or IGF-I, FSH had little or no effect (P>0.05) on estradiol production by granulosa cells from both small (1–5 mm) and large (≥8 mm) follicles. However, in the presence of ≥3 ng/mL of insulin, FSH increased (P<0.05) estradiol production by granulosa cells from small and large follicles such that the estimated dose (ED50) of insulin necessary to stimulate 50% of the maximum estradiol production was decreased by 2- to 3-fold from 22 to 28 ng/mL in the absence of FSH to 7–14 ng/mL in the presence of FSH. Similarly, in the presence of ≥3 ng/mL of IGF-I, FSH increased (P<0.05) estradiol production by granulosa cells from small and large follicles such that the ED50 of IGF-I for estradiol production was decreased by 4- to 5-fold from 25 to 36 ng/mL in the absence of FSH to 5–6 ng/mL in the presence of FSH. In the presence of FSH, the maximal effect of insulin on estradiol production was much greater than that of IGF-I (137- versus 12-fold increase) and were not additive; when combined, 100 ng/mL of IGF-I completely blocked the stimulatory effect of 100 ng/mL of insulin. In the absence of FSH, the maximal effect of insulin and IGF-I on estradiol production was similar. Concomitant treatment with 30 ng/mL of LH reduced (P<0.05) insulin-stimulated estradiol production by 52% on day 1 and 19% on day 2 of treatment. Insulin, IGF-I and FSH also increased (P<0.05) granulosa cell numbers and progesterone production but their maximal effects were less (i.e., <4-fold increase) than their effects on estradiol production. In conclusion, insulin and IGF-I synergize with FSH to directly regulate ovarian follicular function in cattle, particularly granulosa cell aromatase activity.  相似文献   

7.
The objective of the present studies was to determine the effect of cytokines on FSH-induced estrogen production by granulosa cells from small (1–5 mm) and large (≥ 8 mm) bovine follicles. FSH-induced estradiol production by granulosa cells from large follicles (expressed as pg estradiol/105 cells/24 hr) was not affected (P>.05) by 10 or 100 ng/ml of interleukin (IL)-1β, 10 or 100 ng/ml of tumor necrosis factor-α (TNFα) or 100 ng/ml of IL-2. In contrast, 100 ng/ml of IL-1β, IL-2 or TNFα inhibited (P<.05) FSH-induced estradiol production by 31%, 55% or 72%, respectively in cells from small follicles. Interferon-α (IFNα; 100 U/ml) inhibited (P<.05) FSH-induced estradiol production by 61% and 20% in cultures of cells from small and large follicles, respectively. Interferon-β (IFNβ; 100 U/ml), interferon γ (IFNγ; 100 U/ml) and bovine trophoblast protein-1 (bTP-1; 100 U/ml) inhibited (P<.05) estradiol production by 47%, 71% and 28%, respectively in cells from small follicles, but had no effect (P>.05) on FSH-induced estradiol production in cells from large follicles. TNFα binding protein-I blocked (P<.05) the inhibitory effect of TNFα on FSH-induced estradiol production by cells from small follicles. Viability of granulosa cells was not affected (P>.05) by the various cytokines. In summary, cytokines have little or no effect on FSH-induced estradiol production by bovine granulosa cells collected from large follicles, whereas cytokines (bTP-1 ≤ IL-1β < IL-2 = IFNβ < IFNα < IFNγ = TNFα) have potent inhibitory effects on FSH-induced estradiol production by granulosa cells collected from small follicles. Thus, it appears that less differentiated granulosa cells (small follicles) are more responsive to cytokines than are highly differentiated granulosa cells (large follicles).  相似文献   

8.
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   

9.
The aim of this study was to investigate whether functional tumor necrosis factor-alpha (TNFalpha) receptors are present in the granulosa cells and the cells of theca interna (theca cells), obtained from bovine follicles classified into one of three groups. Each group was defined as either small vesicular ovarian follicles (small follicles; 3-5 mm in diameter), preovulatory mature ovarian follicles (preovulatory follicles) or atretic follicles (12-18 mm) according to gross examination of the corpus luteum in the epsilateral or contralateral ovary and the uterus (size, color, consistency and mucus), and the ratio of progesterone (P(4)) and estradiol-17beta (E(2)) concentrations in follicular fluid. A Scatchard analysis showed the presence of a high-affinity binding site on both granulosa and theca cells from all follicles examined (dissociation constant: 4.7 +/- 0.15 to 6.9 +/- 1.40 nM). Moreover, TNFalpha receptor concentrations in granulosa and theca cells obtained from atretic follicles were significantly higher than those in the cells from preovulatory follicles (P<0.05). Exposure of cultured granulosa cells from small antral follicles to recombinant human TNFalpha (rhTNFalpha; 0.06-6 nM) inhibited E(2) secretion in a dose-dependent fashion (P<0.01), but did not affect P(4) secretion. In addition, rhTNFalpha inhibited follicle stimulating hormone-, forskolin- or dibutylyl cyclic AMP-induced P(4) and E(2) secretion by the cells (P<0.01). These results indicate the presence of functional TNFalpha receptors in bovine granulosa and theca cells in small, preovulatory and atretic follicles, and suggest that TNFalpha plays a role in regulating their secretory function.  相似文献   

10.
There is increasing evidence that inflammatory cytokines regulate corpus luteum (CL) function in many species. The purpose of the present study was to determine whether interleukin (IL)-4 and IL-6 are expressed in the porcine CL, and whether these cytokines influence porcine luteal steroidogenesis. The gene expressions of IL-4, IL-6 and their specific receptors were determined in the CL of Chinese Meishan pigs during the estrous cycle. Moreover, the effects of these cytokines on progesterone (P(4)), estradiol-17beta (E(2)) and prostaglandin (PG) F2alpha secretion by cultured luteal cells were investigated. IL-4 and IL-6 mRNAs were detected in the CL at all luteal stages. Furthermore, mRNAs of the receptors for IL-4 and IL-6 were clearly expressed in the CL throughout the estrous cycle. Real-time PCR analysis revealed that IL-6 receptor (IL-6R) mRNA expression was higher in the regressed CL (days 19-21 after ovulation) than in the CL at other stages (P<0.01). Exposure of cultured luteal cells obtained from mid-stage CL (days 8-11) to IL-6 (1-100 ng/ml), it inhibited P(4) and E(2) secretion by the cells (P<0.05). Although IL-4 (1-100 ng/ml) did not significantly alter P(4) secretion, it inhibited E(2) secretion by the cells (P<0.05). Neither IL-4 nor IL-6 had any effect on PGF2alpha secretion by the cells. These results suggest that IL-4 and IL-6 are locally produced in the porcine CL, and that they inhibit steroid production from luteal cells via their specific receptors. Collectively, both IL-4 and IL-6 may play roles in regulating porcine CL function throughout the estrous cycle.  相似文献   

11.
The objectives of this study were to characterize the relationship between estradiol and transforming growth factor-β1(TGF-β1) concentrations in follicular fluid of growing bovine ovarian follicles, and to examine the effect of TGF-β1 on FSH-stimulated estradiol secretion in cultured bovine granulosa cells. Follicular fluid was collected from individual follicles >5 mm in diameter by ultrasound-guided transvaginal puncture (n = 12 heifers). Follicles were sampled at four different stages of development of the first post-ovulatory wave during selection of the single dominant follicle. Estradiol, progesterone and total TGF-β1 were measured in follicular fluid of the three or four largest follicles sampled when the largest follicle (F1) had reached either 6.5, 7.5, 8.5 or 9.5 ± 0.5 mm stage of development. There was a significant negative relationship between follicular fluid TGF-β1 and estradiol concentrations (R2 = 0.44; p < 0.002), and between TGF-β1 concentrations and follicle diameter (R2 = 0.23; p < 0.01) in cohort follicles at the 6.5 mm stage, but not at any later stage of development of the follicle wave. There was no correlation between progesterone and TGF-β1 concentrations at any stage. To assess the causal relationship between TGF-β1 and estradiol, granulosa cells from follicles measuring 2–5 mm at dissection were placed in serum-free culture. TGF-β1 caused a dose-dependent decrease in FSH-stimulated estradiol secretion (P < 0.001). These findings suggest that TGF-β1 has an inhibitory effect on estradiol secretion in FSH-stimulated follicles and that a reduction in TGF-β1 inhibition may be part of the mechanism of selection of a single dominant follicle.  相似文献   

12.
The influence of phthalate esters di-2-ethylhexyl phthalate (DEHP) and mono-2-ethylhexyl phthalate (MEHP) on uterine prostaglandin (PGF2α and PGE2) and ovarian oxytocin secretion was investigated. Endometrial, granulosa, and luteal cells from cows on days 8–12 of the estrous cycle were treated with DEHP or MEHP (0.1, 1, or 10 ng/mL). We found that DEHP and MEHP stimulated (P < 0.05) secretion of PGF2α and inhibited (P < 0.001) secretion of PGE2 from endometrial cells. The ratio of PGF2α to PGE2 was markedly altered. The endocrine disrupting chemicals also enhanced secretion of oxytocin (P < 0.05) from ovarian cells. Our results indicated that DEHP and its metabolite MEHP could affect the process of the estrous cycle by impairing secretion of prostaglandin from the uterus and oxytocin from the ovary.  相似文献   

13.
Dairy cattle are susceptible to heat stress-induced reductions in fertility; however, direct effects of hyperthermia on specific reproductive functions are difficult to determine in vivo. The objective of this experiment was to examine the effect of elevated temperature in vitro on follicular steroidogenesis, to gain insight into specific follicular responses associated with heat stress. Dominant follicles were obtained from Holstein heifers on day 6 post-estrus (luteal phase; n = 4) or day 8, 36 h after an injection with 25 mg PGF(2alpha) to induce regression of the corpus luteum (follicular phase; n = 4). Pieces of follicle wall were isolated from dominant follicles and cultured for 96 h with 0, 2 or 100 ng/ml LH or FSH at 37, 39 or 41 degrees C. Concentrations of androstenedione, estradiol and progesterone were determined in culture media collected every 24h. During the last 48 h of culture, basal secretion of androstenedione and estradiol by pieces of follicle wall was lower at 41 degrees C than at 37 or 39 degrees C (P < 0.05). In contrast, cumulative secretion of progesterone by pieces of follicle wall in medium alone was higher at 41 degrees C than at 37 or 39 degrees C (P < 0.05). Pieces of follicle wall responded to treatment with both low (2 ng/ml) and high (100 ng/ml) doses of gonadotropins at all temperatures. However, gonadotropin-induced secretion of androstenedione and estradiol was generally lower, whereas gonadotropin-induced secretion of progesterone was higher at 41 degrees C and sometimes at 39 than at 37 degrees C. The changes in basal steroidogenesis and in responses to gonadotropins suggest that follicular cells begin to luteinize at elevated temperatures in vitro. Premature luteinization of follicular cells in vivo has been associated with reduced fertility in cattle with persistent follicles, suggesting that the premature differentiation of follicular cells observed in the current study may be responsible, in part, for the reduced fertility of dairy cattle under heat-stressed conditions.  相似文献   

14.
Oxytocin (OT) stimulates pulsatile secretion of uterine prostaglandinF2alpha in ruminants, but the role of OT in the estrous cycle regulation of pigs is not clear. These studies were performed to determine the effect of exogenous OT on interestrous interval of intact cyclic gilts. Intrauterine infusion of 80 USP units three times daily on d 10 to 16 after estrus did not decrease interestrous interval (24.5+/-1.3 d) compared with control gilts (22.5+/-1.3 d). In contrast, i.m. injections of 20 USP units of OT twice daily or 80 USP units of OT three times daily on d 10 to 16 after estrus decreased (P < 0.05) interestrous interval (20.0+/-0.3 or 19.5+/-0.4 d, respectively) compared with control gilts (20.5+/-0.3 d). When gilts received a single i.m. injection of 0 or 1 mg of estradiol valerate on d 11 and twice daily i.m. injections of 0 or 20 USP units OT on d 10 to 16 after estrus, an interaction (P = 0.05) between OT and estradiol valerate was detected. In the absence of exogenous estradiol valerate, injection of OT decreased interestrous interval (19.0+/-0.5 d) compared with injection of vehicle (20.4+/-0.5 d). However, when gilts were injected with 1 mg of estradiol valerate to inhibit luteolysis, OT did not prevent the increase in interestrous interval (25.4+/-0.5 d) compared with injections of vehicle (24.7+/-0.5 d). These results indicate that 1) exogenous OT administered by intrauterine infusion on d 10 to 16 did not decrease interestrous interval of intact cyclic gilts, 2) exogenous OT administered i.m. on d 10 to 16 shortened interestrous interval, and 3) exogenous OT did not prevent the increase in interestrous interval induced by estradiol valerate.  相似文献   

15.
The use of soy-based products in pig diets had raised concerns regarding the reproductive toxicity of genistein, the predominant isoflavone in soybeans. Genistein was reported to exhibit weak estrogenic activity but its mechanism of action is not fully recognized. The aim of the study was to examine the in vitro effects of genistein on (1) progesterone (P4) and estradiol (E2) secretion by porcine granulosa cells harvested from medium follicles, (2) the viability of cultured granulosa cells, and (3) the mRNA and protein expression of estrogen receptors α and β (ERα and ERβ) in these cells. In addition, to verify the role of protein tyrosine kinase (PTK)–dependent mechanisms possibly involved in genistein biological action, we tested the effects of lavendustin C, the nonsteroidal PTK inhibitor, on granulosa cell steroidogenesis. We found that genistein inhibited (P < 0.05) basal P4 secretion by granulosa cells harvested from medium follicles of pigs. In contrast, lavendustin C did not affect basal P4 secretion by the cells. Moreover, genistein increased (P < 0.05) basal granulosal secretion of E2. In contrast, lavendustin C did not alter basal E2 secretion by porcine granulosa cells. In addition, we demonstrated that genistein increased mRNA and protein expression of ERβ (P < 0.05) in the examined cells. The expression of ERα mRNA was not affected by genistein and ERα protein was not detected in the cultured granulosa cells of pigs. In summary, the genistein action on follicular steroidogenesis in pigs involved changes in the granulosal expression of ERβ. However, the genistein action on P4 and E2 production by granulosa cells harvested from medium follicles did not seem to be associated with PTK.  相似文献   

16.
Because IGFBP inhibit IGF-stimulated cellular proliferation and differentiation, it is hypothesized that variations among IGFBP in individual follicles might contribute to the regulation of recruitment, selection, dominance, and turnover of ovarian follicles. Sources of IGFBP in fluid of bovine follicles are not well established; thus, objectives of this study were to determine levels of IGFBP binding activities and messenger RNA (mRNA) in granulosa and theca interna cells at different stages of follicular development (small [< 6 mm], medium [6 to < 8 mm], and large [> or = 8 mm]) and to characterize associations of these levels measured in the cells with levels of IGFBP and steroids in follicular fluid. Thecal and granulosa cells from large healthy follicles contained two- to twentyfold less (P < 0.05) IGFBP-2, -3, and -5 than cells from small, medium, and large atretic follicles. Thecal cells from small, medium, and large atretic follicles contained more (P < 0.05) IGFBP-3 and -4 than granulosa cells from these follicles, whereas granulosa cells from these follicles contained more IGFBP-2 activity than thecal cells. Differences in IGF binding activity were paralleled by differences in levels of mRNA for the respective IGFBP. Developmental differences in IGFBP activity in follicular fluid were positively associated with activity in granulosa and/or thecal cells, with the exception of IGFBP-4, which was low in fluid from large healthy follicles but markedly increased (mRNA and binding activity) in granulosa cells from these follicles. It is concluded that developmental changes in follicular fluid IGFBP-2 and -5 binding activities seem to be controlled in part by alterations in synthesis of these IGFBP by granulosa and thecal cells, whereas diminished IGFBP-4 in fluid from large healthy follicles occurs concomitantly with increased levels of IGFBP-4 mRNA and activity in granulosa cells, implicating posttranslational regulation by specific proteases.  相似文献   

17.
This study examined the relationship among growth, steroid production and transforming growth factor-beta 1 (TGF-beta 1) immunolocalization in the mouse follicles cultured in vitro to evaluate the hypothesis that normally developing follicles should express TGF-beta 1 in the granulosa cells around the time of antrum formation. Preantral follicles with 151-175 microns (large category) and 125-150 microns (small category) of initial diameters were used as models for normal and retarded follicles, respectively. Growth rate and timing of antrum formation in both categories were comparable to those of in-vivo grown follicles. At the time of antrum formation, follicular diameters were similar between the two follicle categories; however, antral follicles from the large category showed larger number of granulosa cells, higher estradiol production and proportion of follicles with TGF-beta 1 positive granulosa cells. Two days after antrum formation, there were no differences in the number of granulosa cells and the proportions of follicles with TGF-beta 1 positive granulosa or theca cells between the two categories. Temporal association in large follicles between the increase in estradiol production and proportion of follicles with TGF-beta 1 positive granulosa cells at the time of antrum formation supports our hypothesis. Furthermore, this study demonstrated the usefulness of the follicle culture system in the investigations of follicular physiology.  相似文献   

18.
IGF-1 plays a key role in the proliferation and differentiation of granulosa cells. However, the molecular mechanism of IGF-1 action in avian granulosa cells during follicle maturation is unclear. Here, we first studied IGF-1 receptor (IGF-1R) expression, IGF-1-induced progesterone production and some IGF-1R signaling pathways in granulosa cells from different follicles. IGF-1R (mRNA and protein) was higher in fresh or cultured granulosa cells from the largest follicles (F1 or F2) than in those from smaller follicles (F3 or F4). In vitro, IGF-1 treatment (10(-8)M, 36h) increased progesterone secretion by four-fold in mixed F3 and F4 (F3/4) granulosa cells and by 1.5-fold in F1 granulosa cells. IGF-1 (10(-8)M, 30min)-induced increases in tyrosine phosphorylation of IGF-1R beta subunit and phosphorylation of ERK were higher in F1 than in F3/4 granulosa cells. Interestingly, IGF-1 stimulation (10(-8)M, 10min) decreased the level of AMPK Thr172 phosphorylation in F1 and F3/4 granulosa cells. We have recently showed that AMPK (AMP-activated protein kinase) is a protein kinase involved in the steroidogenesis in chicken granulosa cells. We then studied the effects of AMPK activation by AICAR (5-aminoimidazole-4-carboxamide ribonucleoside), an activator of AMPK, on IGF-1-induced progesterone secretion by F3/4 and F1 granulosa cells. AICAR treatment (1mM, 36h) increased IGF-1-induced progesterone secretion, StAR protein levels and decreased ERK phosphorylation in F1 granulosa cells. Opposite data were observed in F3/4 granulosa cells. Adenovirus-mediated expression of dominant negative AMPK totally reversed the effects of AICAR on IGF-1-induced progesterone secretion, StAR protein production and ERK phosphorylation in both F3/4 and F1 granulosa cells. Thus, a variation of energy metabolism through AMPK activation could modulate differently IGF-1-induced progesterone production in F1 and F3/4 granulosa cells.  相似文献   

19.
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20.
The effects of estradiol, insulin, and gonadotropins on levels of insulin-like growth factor binding protein (IGFBP)-2, -3, -4, and -5 mRNA levels in bovine granulosa and theca cells were evaluated in vitro using serum-free medium containing various hormone treatments arranged in four different experiments. Amounts of IGFBP-2, -3, -4 and -5 mRNA were quantitated using fluorescent quantitative real-time RT-PCR. In small-follicle (1-5 mm) granulosa cells, follicle-stimulating hormone (FSH) in the presence or absence of insulin increased (P<0.05) IGFBP-3 mRNA but did not change IGFBP-2, -4, or -5 mRNA levels; estradiol was without effect on IGFBP-2, -3, -4, or -5 mRNA levels in the absence of insulin but increased (P<0.05) IGFBP-2 mRNA levels in the presence of insulin. Luteinizing hormone (LH) in the absence (but not presence) of insulin increased (P<0.05) small-follicle granulosa cell IGFBP-3 mRNA levels. In large-follicle (>7.9 mm) granulosa cells, insulin alone increased (P<0.05) IGFBP-2 gene expression while LH, FSH, and estradiol were without effect (P>0.10). Estradiol (3 and 300 ng/ml) decreased (P<0.05) IGFBP-5 mRNA levels in large-follicle granulosa cells. In theca cells, insulin decreased (P<0.05) IGFBP-4 expression, but had no effect (P>0.10) on IGFBP-2, -3, or -5 mRNA levels. Estradiol decreased (P<0.05) IGFBP-2, -3, and -4 mRNA levels but had no effect on IGFBP-5 mRNA levels in theca cells. LH had no effect on levels of IGFBP-2, -3, -4, or -5 mRNA in theca cells. These results indicate that expression of IGFBP-2, -3, -4, and -5 mRNA by granulosa and theca cells are differentially regulated by estradiol, insulin and gonadotropins, therefore discretely modulating the amount of bioavailable IGFs to these cells depending upon the specific hormonal stimuli. In particular, these studies are the first in cattle to show that estradiol selectively inhibits IGFBP-2, -3, and -4 gene expression in theca cells, inhibits IGFBP-5 gene expression in large-follicle granulosa cells, and stimulates IGFBP-2 gene expression in small-follicle granulosa cells.  相似文献   

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