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1.
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   

2.
Sanguinarine (SA), a phytobiotic from Sanguinaria Canadensis, has been demonstrated to inhibit vessel growth. Current restrictions on the use of antibiotic growth promoters have motivated addition of this alkaloid as a naturally appetizing feed additive for farm animals. However, concern may araise since angiogenesis is a fundamental event in ovarian follicle growth. Therefore, the aim of this study was to evaluate the potential negative role of SA in follicular angiogenesis. For this purpose, we studied the effect of 300 nM SA on the production of vascular endothelial growth factor (VEGF) by swine granulosa cells from follicles >5 mm and on the activation of Akt, the main effector of the VEGF signalling pathway. In addition, the potential interference of SA in vessel development was tested in an in vitro angiogenesis bioassay. SA inhibited both VEGF production and VEGF-induced Akt activation in swine granulosa cells. Moreover, it was able to block vessel growth induced by VEGF. Taken together, our results suggest that SA could be detrimental to follicular angiogenesis, and therefore supplementation of feed with this alkaloid should be carefully considered.  相似文献   

3.
The aim of this study was to investigate the effects of some endocrine and intra‐ovarian factors on the activation/inhibition of apoptosis in swine granulosa cells. Upon incubation in a 10% FCS‐supplemented M199, granulosa cells from small (< 3 mm) follicles programmed their death after 24–48 h of culture; in the absence of FCS, apoptosis was reduced after 24 h of culture. Cells cultured in the presence of FCS were treated with db‐cAMP, LH, FSH, Insulin‐like Growth Factor‐I IGF‐I or PMSG to verify the role of these substances in apoptotic death: all these molecules inhibited apoptosis after 48 h of incubation. A further aim of the study was to investigate the possible involvement of nitric oxide (NO), an intra‐ovarian modulator, in the regulation of granulosa cell apoptosis and its possible role in the modulation of steroidogenesis. After a 48 h incubation with a substrate of NO synthesis ( l ‐arginine, 0.1 and 1 m m ), a NO donor [S‐nitroso‐N‐acetyl‐penicillamine (SNAP) , 0.2 and 1 m m ] or a NO synthase inhibitor [Nω‐nitro‐ l ‐arginine‐methyl‐ester (NAME, 1 and 5 m m )], the onset of apoptotic death was evaluated: l . arginine and NAME did not induce any significant variation of apoptosis, whereas 1 m m SNAP exerted a protective action. A significant stimulatory effect of l ‐arginine on NO production, associated with a suppressive action on estradiol 17β concentrations was observed; NAME exerted an inhibitory effect on NO production, associated with an increase in estradiol secretion; estradiol 17β production was markedly inhibited by SNAP. In summary, the depletion of FCS could induce a cell cycle arrest in G0 whereas apoptosis could be the consequence of cell cycle progression mediated by FCS; gonadotropins and IGF‐I could also act as survival factors. NO appeared to represent a ‘trophic’ signal for the follicle, whose involvement in the regulation of ovarian function is substantiated by its modulatory action on steroidogenesis.  相似文献   

4.
The use of herbs as additives in livestock nutrition as an alternative to antibiotics is becoming a new goal in animal production. It is known that green tea exerts antimicrobial activity owing to specific flavonoid compounds named catechins, primarily represented by epigallocatechin-3-gallate (EGCG). Remarkably, despite many potential benefits of green tea and EGCG consumption, it is also important to get an insight on the possible reproductive-related consequences of feeding supplementation. To this purpose, granulosa cells were harvested from follicles > 5mm and treated with 5 and 50 microg/ml of EGCG in order to evaluate the effects on the main parameters of granulosa cell function: steroidogenesis, by measuring progesterone and estradiol-17beta production, and proliferation, one of the major feature of ovarian follicular growth. Moreover, as the genesis of new vessels has been demonstrated to be fundamental for follicle development, we evaluated the effect of EGCG on the production of the main angiogenetic factor, VEGF, by swine granulosa cells. Finally, since reactive oxygen species (ROS) might be involved in the control of female reproductive activity, we studied the effect of EGCG on superoxide anion (O2-) and hydrogen peroxide (H2O2) production by swine granulosa cells and on the activity of the scavenging enzyme superoxide dismutase (SOD). EGCG significantly (p < 0.05) inhibited proliferation, steroidogenesis, VEGF and O2- production by swine granulosa cells; on the contrary, H2O2 levels and SOD activity were stimulated (p < 0.05) by the catechin. Therefore, since our data demonstrate that EGCG has a negative effect on reproductive performances in swine, feeding supplementation should be carefully considered.  相似文献   

5.
Ovarian angiogenesis, which is currently considered to be of crucial importance in controlling the growth of developing follicles, is a physiological process driven by a variety of angiogenic factors. Among these, vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) have been recognized as key players in promoting cell growth and differentiation. Porcine granulosa cells from small (<3 mm), medium (3–5 mm) and large (>5 mm) follicles were seeded at different densities in DMEM:Ham's F12 (1:1) with or without different concentrations of VEGF or bFGF. After 48 h of culture, media were assayed for oestradiol (E2) 17β, progesterone (P4), nitric oxide (NO) and VEGF levels; in addition, cell proliferation was evaluated by 3H‐thymidine incorporation assay. Both bFGF and VEGF effects on E2 and P4 production by cultured granulosa cells resulted to be dependent on follicle size. The bFGF was always ineffective in modulating cell proliferation, while VEGF exerted an inhibitory effect on the proliferation in the small follicle group and a stimulatory one in the medium and large follicle groups. The bFGF consistently reduced NO levels in culture media. The VEGF appeared to be ineffective in modifying NO production in the small follicle group, while it was stimulatory in the medium follicle group and inhibitory in the large follicle group. Basal VEGF production was higher in cells from the large follicle as compared with the small and medium follicle groups, and it was unaffected by bFGF. These results suggest that VEGF plays a modulatory role in granulosa cell functional activity and it is possibly involved in the regulation of follicle growth; on the contrary, bFGF does not appear to represent a significant regulatory factor in our cellular model, except for an inhibitory action on the production of NO, whose anti‐angiogenic properties need to be further substantiated.  相似文献   

6.
This study was aimed at testing the hypothesis that insulin-like growth factor binding protein (IGFBP)-3 can modulate hormone-dependent differentiation of granulosa cells in vitro. Granulosa cells from small (1 to 5 mm) follicles were collected from cattle, cultured for 2 d in medium containing 10% fetal calf serum, washed, and then treated for an additional 2 d in serum-free medium with follicle-stimulating hormone (FSH) (50 ng/ml), recombinant human IGF-I (0, 1.3, 4.0, or 13.3 nM), or recombinant human IGFBP-3 (0 to 4.26 nM). In one series of experiments, IGFBP-3 (0.53 and 2.13 nM) inhibited (51% to 92% decreases; P < 0.05) progesterone and estradiol production induced by 1.3 nM of IGF-I, but did not influence (P > 0.10) granulosa cell numbers or steroidogenesis in the absence of IGF-I. Only 4.26 nM of IGFBP-3 inhibited (by 35%) the increase in granulosa cell numbers induced by 1.3 nM of IGF-I. In another series of experiments, 13.3 nM of IGF-I, but not 4.0 nM of IGF-I, was able to completely overcome the inhibitory effect of 4.26 nM of IGFBP-3 on estradiol production. The increase in cell numbers induced by 4.0 and 13.3 nM of IGF-I was attenuated (P < 0.001) by 4.26 nM of IGFBP-3. In a third series of experiments, IGFBP-3 inhibited 125I-IGF-I binding to granulosa cells. These results indicate that IGFBP-3 has a pronounced inhibitory effect on IGF-I action in cultured bovine granulosa cells, and that this inhibitory effect is likely attributable to IGFBP-3 binding/sequestering IGF-I. Thus, IGFBP-3 may play a significant role in regulating granulosa cell proliferation and steroidogenesis during follicular development in cattle.  相似文献   

7.
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8.
Bisphenol S (BPS) is an endocrine‐disrupting chemical with multiple potential mechanisms of action, including as an oestrogen receptor agonist. BPS is increasingly used in plastics and thermal receipts as a substitute for bisphenol A, which has been phased out due to concerns about human health implications. The ability of BPS to alter female reproductive function in mammals has not been widely studied, despite the importance of normal hormone signalling for female reproduction. The aim of this study was to investigate how BPS (in a wide range of doses, including very low doses) affects granulosa cell and theca cell steroid hormone production and cell viability in the bovine. Granulosa cell oestradiol production was stimulated when cells were exposed to 100 μM BPS under basal conditions, but there was no effect of BPS when cells were stimulated with follicle‐stimulating hormone (FSH). Additionally, there was no effect of BPS on granulosa cell progesterone production or cell viability under basal or FSH‐stimulated conditions. BPS did not affect theca cell androstenedione or progesterone production, or theca cell viability under basal or luteinizing hormone‐stimulated conditions. This study suggests for the first time that BPS may alter oestradiol production by bovine granulosa cells, albeit at a concentration that is unlikely to be physiologically relevant. Further studies are needed to determine the effects of BPS on the bovine oocyte and on other functions of follicular cells.  相似文献   

9.
Bisphenol A (BPA) is a chemical of high production volume that is used widely in many industries and is known as a xenooestrogen and anti‐thyroid hormone endocrine disrupter. There is little information regarding the effects of BPA in the presence of thyroid hormone on porcine granulosa cell development. Thus, the primary granulosa cells were treated with thyroxine (T4, 10 nM), BPA (10 µM) or T4 plus BPA; we subsequently evaluated the effects of T4 or BPA on 17β‐estradiol synthesis, cellular proliferation and apoptosis. Our data showed that BPA significantly increased the accumulation of 17β‐estradiol and promoted granulosa cell proliferation, whereas T4 significantly decreased 17β‐estradiol and had no effect on cellular proliferation. In addition, it was noteworthy that T4 treatment induced apoptosis in porcine granulosa cells and BPA co‐incubation attenuated T4‐induced apoptosis as shown from flow cytometric assay analysis. We hypothesized that BPA attenuates T4‐induced apoptosis by regulating 17β‐estradiol accumulation and oestrogen receptor‐mediated signalling pathways. In conclusion, our results demonstrated that T4 affected 17β‐estradiol accumulation and induced cellular apoptosis, but did not affect granulosa cell proliferation. Exposure to BPA increased 17β‐estradiol accumulation, promoted granulosa cell proliferation and attenuated T4‐induced apoptosis in porcine granulosa cells in vitro.  相似文献   

10.
Our work with both sheep and mouse models has revealed many of the neural substrates and signalling pathways involved in olfactory recognition memory in the main olfactory system. A distributed neural system is required for initial memory formation and its short-term retention-the olfactory bulb, piriform and entorhinal cortices and hippocampus. Following memory consolidation, after 8 h or so, only the olfactory bulb and piriform cortex appear to be important for effective recall. Similarly, whereas the glutamate-NMDA/AMPA receptor-nitric oxide (NO)-cyclic GMP signalling pathway is important for memory formation it is not involved in recall post-consolidation. Here, within the olfactory bulb, up-regulation of class 1 metabotropic glutamate receptors appears to maintain the enhanced sensitivity at the mitral to granule cell synapses required for effective memory recall. Recently we have investigated whether fluctuating sex hormone levels during the oestrous cycle modulate olfactory recognition memory and the different neural substrates and signalling pathways involved. These studies have used two robust models of social olfactory memory in the mouse which either involve social or non social odours (habituation-dishabituation and social transmission of food preference tasks). In both cases significant improvement of learning retention occurs when original learning takes place during the proestrus phase of the ovarian cycle. This is probably the result of oestrogen changes at this time since transgenic mice lacking functional expression of oestrogen receptors (ERalpha and ERbeta, the two main oestrogen receptor sub-types) have shown problems in social recognition. Therefore, oestrogen appears to act at the level of the olfactory bulb by modulating both noradrenaline and the glutamate/NO signalling pathway.  相似文献   

11.
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   

12.
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13.
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   

14.
Gonadotropins are required for follicular growth and differentiation, but increasing amounts of evidence indicate that intrafollicular factors modulate their effects at granulosa cell level. In order to study the effect of factors present in bovine follicular fluid, a partial purification of low molecular mass factors from fluids collected from small (< 5 mm), medium (5–8 mm) and large (> 8 mm) follicles was performed and the biological activity of these peptides on steroidogenesis of granulosa cells from small and large follicles was examined. The purification was carried out by filtration through membranes in 25 and 10 kDa molecular weight cutoffs. After filtration, samples were analysed by polyacrylamide gel electrophoresis and protein concentration was measured by spectrophotometric analysis. Granulosa cells from small and large follicles were cultured in serum‐free DMEM/Ham's F12 (1 : 1) plus transferrin (5 mg/l) and selenium (5 µg/l) for 2 days. At the end of the culture period, media were renewed and follicular extracts from two different preparations (< 25 and < 10 kDa) were added at the concentrations of 1–10–100–1000 ng/ml. After 24 h the media were collected and stored until estradiol 17β (E2) and progesterone (P4) determination by validated radio‐immuno‐assays. Basal P4 production was 18.3 ± 1.4 (mean ± SEM)and 9.8 ± 1.8 ng/24 h per 3 × 104 cells from small and large follicles, respectively. Both < 10 and < 25 kDa extracts reduced P4 production by cells from both the types of follicles (p < 0.05). Basal E2 release was 671.8 ± 21.4 and 5500 ± 800 pg/24 h per 3 × 104 cells from small and large follicles, respectively. Both extracts reduced E2 production by either cells from small and large follicles (p < 0.05). No differences were observed in the inhibition of steroidogenesis by purifications obtained from large, medium or small follicles. Results of this study indicate that factors present in bovine follicular fluid can reduce steroidogenesis in granulosa cells in vitro.  相似文献   

15.
Cystic ovarian disease (COD) is one of the main causes of infertility in dairy cattle. It has been shown that intra‐ovarian factors may contribute to follicular persistence. Transforming growth factor‐beta (TGFB) isoforms are important paracrine and autocrine signalling molecules that regulate ovarian follicle growth and physiology. Considering the importance of these factors in the ovarian physiology, in this study, we examined the expression of TGFB isoforms (TGFB1, TGFB2 and TGFB3) in the ovary of healthy cows and animals with spontaneous and adrenocorticotrophic hormone (ACTH)‐induced COD. In the oestrous‐synchronized control group, the expression of TGFB1 in granulosa and theca cells was higher in spontaneous cysts than in atretic or tertiary follicles. When we compared TGFB2 expression in granulosa cells from atretic or tertiary follicles from the oestrous‐synchronized control group with that in ACTH‐induced or spontaneous follicular cysts, we found a higher expression in the latter. The expression of the TGFB isoforms studied was also altered during folliculogenesis in both the spontaneous and ACTH‐induced COD groups. As it has been previously shown that TGFB influences steroidogenesis, ovarian follicular proliferation and apoptosis, an alteration in its expression may contribute to the pathogenesis of this disease.  相似文献   

16.
This study was undertaken to elucidate the effect of ammonia‐generating diet on serum and follicular fluid ammonia and urea levels, serum oestrogen and progesterone concentrations and granulosa cell growth and secretion parameters in ewes (Ovis aries). Ewes were fed with 14% CP diet (control) or ammonia‐generating diet or ammonia‐generating diet plus soluble sugar. The serum and follicular fluid ammonia and urea level, serum oestrogen and progesterone levels and granulosa cell (obtained from ovaries of slaughtered ewes) growth parameters and secretory activities were estimated. Ammonia‐generating diet (high‐protein diet) increased the serum ammonia and urea concentration. Supplementation of soluble sugar significantly reduced the ammonia concentration in serum with comparable levels as in control group; however, the urea level in the same group was higher than that observed in control group. Supplementation of soluble sugar significantly reduced the follicular fluid ammonia concentration; however, the level was significantly higher compared to control group. Supplementation of soluble sugar brought down the follicular fluid urea level comparable to that observed in control group. Oestrogen and progesterone levels remained unchanged in ewes fed with different types of diet. Oestrogen and progesterone secretion were significantly lowered from granulosa cells recovered from ewes fed with high ammonia‐generating diet. Low metabolic activity and high incidence of apoptosis were observed in granulosa cells obtained from ovaries of ewes fed with ammonia‐generating diet.  相似文献   

17.
In the pig, conceptus-derived oestrogens (days 11 and 12 of pregnancy) seem to be a critical component of the signalling mechanism for maternal recognition of pregnancy. Embryonic oestrogens can mediate effects on endometrial function by interactions with epithelial and stromal oestrogen receptors (ER). Recent data demonstrate that cell membrane ER interacts with the phosphatidylinositol 3-kinase/Akt pathway in several types of cells. The protein kinase Akt is involved in the control of cell growth, survival and proliferation. One distinct function of the Akt signalling cascade is its ability to phosphorylate the eukaryotic initiation factor-4E (eIF-4E)-binding protein 1 (4E-BP1). This phosphorylation suppresses the inhibitory effect of 4E-BP1 on the translation initiation factor eIF4E and in such a way potentially stimulates gene expression at the level of translational initiation. The aim of the present study was to examine if embryonic oestradiol (E(2)) transmits its effect by such a mechanism. Endometrial cells of cyclic gilts (day 13 of the oestrous cycle, n = 4) were cultured and supplemented with vehicle (control), E(2) (50 and 100 pm/l) or with the selective ER modulator raloxifen (10 and 1000 nm/l), and incubated for 24 h. The cell viability was detected by MTT assay, the abundance and phosphorylation of Akt, 4E-BP1 and ERalpha was analysed by Western blotting. Incubation with E(2) or raloxifen did not alter endometrial cell viability. The phosphorylation of Akt at Ser(473) seems to be increased by E(2) (p < 0.05) and decreased by raloxifen (p > 0.05). Raloxifen (1000 nm/l) induced a band shift in 4E-BP1 to the highest electrophoretic mobility which reflects a decrease in phosphorylation (p < 0.05), whereas an influence of E(2) on 4E-BP1 phosphorylation could not be detected. The decrease (p < 0.05) of the abundance of the 80 kDa ERalpha form both by E(2) and raloxifen indicates that the E(2)-stimulated Akt phosphorylation and the inhibition of 4E-BP1 phosphorylation by raloxifen is an E(2) ER-transmitted process. Therefore, embryonic oestrogens can potentially transmit their effect by influencing signalling cascades which modulate gene expression at the level of translational initiation.  相似文献   

18.
It has been hypothesized that the physiological basis of follicle selection is the differential expression of factors, which modulate the action of gonadotrophins on follicular cells, at key points during the process of follicle development. The aim of this research was to test this hypothesis by identifying factors that can enhance or attenuate the action of the gonadotrophins in stimulating follicle development using both in vivo and in vitro models. Experiments in vivo utilized sheep with an ovarian autotransplant to allow intra-arterial infusion of putative local factors and exposure of the ovary to high local concentrations. Experiments in vitro utilized physiological serum-free cell culture systems for both granulosa and theca cells that allow gonadotrophin-induced differentiation in vitro. The putative local factors tested included insulin-like growth factor-I (IGF-I LR3 analogue), transforming growth factor alpha (TGF alpha) or epidermal growth factor (EGF) and inhibin A. IGF-I stimulated both cellular proliferation and hormone production by both granulosa and theca cells in vitro and similarly stimulated ovarian follicle development and ovarian androgen and oestradiol secretion in vivo. Both TGF alpha and EGF stimulated granulosa and thecal cell proliferation in vitro in a dose-responsive manner and concomitantly inhibited hormone production, whereas intra-arterial infusion of TGF alpha in vivo resulted in induction of atresia in large antral follicles and an acute fall in ovarian hormone secretion. Inhibin A in vitro augmented gonadotrophin stimulated androgen and oestradiol production by thecal and granulosa cells, respectively, but had no effect on cell number. Paradoxically, intra-arterial infusion of inhibin A resulted in an acute depression in ovarian steroid secretion. This depression, however, was also associated with an acute depression in circulating FSH concentrations. In conclusion, these data provide strong support for the hypothesis that factors can modulate the action of gonadotrophins on follicular cells to augment (IGF-I, inhibin A) or inhibit (TGF alpha/EGF) granulosa and thecal cell differentiation. The challenge for the future in this area of research is to understand how these factors interact to enable one follicle to be selected from an ovulatory cohort.  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.
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   

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