Progesterone Output Research Articles

The adipokines progranulin and omentin – novel regulators of basic ovarian cell functions

The present study aimed to examine the effects of progranulin and omentin on basic ovarian cell functions. For this purpose, we investigated the effects of the addition of progranulin and omentin (0, 0.1, 1, or 10 ng/ml) on the viability, proliferation, apoptosis and steroidogenesis of cultured rabbit ovarian granulosa cells. To determine the importance of the interrelationships between granulosa cells and theca cells, we compared the influence of progranulin and omentin on progesterone and estradiol release in cultured granulosa cells and ovarian fragments containing both granulosa cells and theca cells. Cell viability, proliferation, cytoplasmic apoptosis and release of progesterone and estradiol were measured by Cell Counting Kit-8 (CCK-8), BrdU incorporation, cell death detection, and ELISA. Both progranulin and omentin increased granulosa cell viability and proliferation and decreased apoptosis. Progranulin increased progesterone release by granulosa cells but reduced progesterone output by ovarian fragments. Progranulin decreased estradiol release by granulosa cells but increased it in ovarian fragments. Omentin reduced progesterone release in both models. Omentin reduced estradiol release by granulosa cells but promoted this release in ovarian fragments. The present observations are the first to demonstrate that progranulin and omentin can be direct regulators of basic ovarian cell functions. Furthermore, the differences in the effects of these adipokines on steroidogenesis via granulosa and ovarian fragments indicate that these peptides could target both granulosa and theca cells.

Fennel affects porcine ovarian cell functions: The interrelationships with the environmental contaminant benzene

The objective of this study was to examine the direct effects of the medicinal plant fennel on basic functions of ovarian cells, including proliferation, apoptosis, and release of progesterone and insulin-like growth factor I (IGFI), as well as to prevent the influence of the environmental contaminant benzene on these cells. Porcine ovarian granulosa cells were cultured with or without fennel extract alone or in combination with benzene. The expression of the proliferation marker PCNA and the apoptosis marker bax was analyzed by quantitative immunocytochemistry and enzyme-linked immunosorbent assay (ELISA). Fennel was able to promote proliferation and IGF-I release, but to suppress apoptosis and progesterone release. Benzene promoted the accumulation of both the proliferation and apoptosis markers, as well as IGF-I release, but it inhibited progesterone secretion. The presence of fennel did not prevent the effects of benzene on any of the measured parameters, while benzene prevented the effects of fennel on cell proliferation, apoptosis, and IGF-I but not progesterone output. These observations demonstrate the direct influence of fennel and benzene on basic ovarian cell functions. Furthermore, they show the inability of fennel to prevent the effects of benzene on these cells. On the other hand, the environmental contaminant benzene can block the response of ovarian cells to the medicinal plant fennel.

Utilizing an immunostimulant (MCWF; Settle) to hasten the degradation of endometrial cups

Placentation in the horse begins at approximately 35 days of gestation, when the chorionic girdle leads to the development of endometrial cups. If embryonic loss occurs in the presence of endometrial cups, return to cyclicity is delayed due to the retainment of these structures, continued production of equine chorionic gonadotropin (eCG), and prolonged progesterone output from accessory corpus lutea. It is believed that natural degradation of endometrial cups is governed by IFNγ-producing lymphocytes at 100-120 days of gestation, but external stimulation of this process has been relatively unsuccessful. Recently, we observed an increase in IFNγ following the use of an immunostimulant (Mycobacterium cell wall fraction (MCWF); Settle) in the postpartum mare. Therefore, we hypothesize that MCWF administration following embryonic loss will increase IFNγ production, hasten the degradation of the endometrial cups, and decrease the time to return to cyclicity in mares. To investigate this, 16 mares were aborted with PGF2α at 40-45d of gestation. Following abortion, mares were divided into groups: MCWF-Treated (6mg MCWF in 20mL LRS; n=10) and Control (20mL LRS; n=6). Between 45-50 days of gestation, mares were assessed via trans-rectal ultrasonography for ovarian function and endometrial cup distribution. Hysteroscopic-guided injection of endometrial cups was performed, with 1mL placed into eachvisible endometrial cup, and the remaining volume injected peripherally. This treatment was repeated 7 days later. Mares were monitored for rectal temperature and serum was taken to assess eCG and IFNγ concentrations. Additionally, transrectal ultrasonography was performed to monitor cyclicity. Statistics were performed utilizing SAS9.4 with a general linear model of repeated measures applied for all temperature, eCG, and cytokine data. A chi-squared test was performed to assess return to cyclicity. An overall decrease in eCG concentrations was noted in the MCWF-treated group (P<0.0001) when compared to controls. This suppression was significant 7 days after onset of treatment and remained suppressed 35 days following treatment. This coincided with an increase in peripheral IFNγ in the MCWF-treated group compared to controls (P<0.01). A higher percentage of MCWF-treated mares returned to cyclicity in comparison to controls (8/10 vs. 2/6; P=0.06), and a pre-ovulatory follicle was noted 23±4 days after onset of treatment. To our knowledge, this is the first report of a successful treatment for degradation of endometrial cups following abortion, by hastening the natural immune response to these structures. This caused mares to return to cyclicity within a month following treatment onset, allowing for repeat attempts at a successful pregnancy within the confines of the breeding season.

Chia (Salvia hispanica L.) can directly suppress basic ovarian cell functions in two farm animal species and protect ovarian cells from the proliferation-stimulating influence of xylene.

The influence of the functional food plant chia (Salvia hispanica L.) on reproduction functions and its ability to prevent the negative effects of environmental contaminants has not yet been studied. Our study aimed to examine the effect of chia seed extract alone and in combination with xylene on the markers of proliferation, apoptosis and hormones release by cultured bovine and porcine ovarian granulosa cells. The extract of chia reduced all of the measured parameters in bovine and porcine ovarian cells but had no effect on the proliferation of porcine cells. Xylene, stimulated proliferation and IGF-I release and inhibited the release of progesterone and testosterone but not apoptosis of bovine granulosa cells. It promoted proliferation, apoptosis and progesterone output by porcine cells. Chia mitigated the stimulatory effect of xylene on proliferation but not on other parameters in both species. The present results are the first demonstration of a direct effect of chia on basic ovarian cell functions. They confirmed a direct influence of xylene on these functions and found a similar stimulatory action of xylene on bovine and porcine ovarian cell proliferation. The present observations demonstrated species-specific differences in the characteristics of xylene influences on ovarian cell apoptosis and secretory activity. Finally, the present results indicate that chia can be a natural protector against the proliferation-stimulating effects of xylene on ovarian cells in both species.

Quercetin can affect porcine ovarian cell functions and to mitigate some of the effects of the environmental contaminant toluene

The present in vitro experiments aimed to examine the effects of the plant polyphenol quercetin and the environmental contaminant toluene on basic ovarian cell functions, including the ability of quercetin to be a natural protector against the adverse effects of toluene. The influence of toluene, quercetin, and their combination on proliferation (accumulation of PCNA), apoptosis (accumulation of bax) and release of progesterone, testosterone and insulin-like growth factor I (IGFI) by cultured porcine ovarian granulosa cells was investigated. Toluene stimulated cell proliferation and inhibited progesterone, IGF-I and testosterone release but did not affect apoptosis. Quercetin, when administered alone, inhibited cell proliferation, apoptosis, IGF-I and testosterone release and stimulated progesterone output. When administered in combination with toluene, quercetin mitigated toluene's effects on proliferation and on progesterone release and induced toluene to exhibit a pro-apoptotic effect. These observations demonstrate the direct effects of both quercetin and toluene on basic ovarian functions and a protective effect of quercetin against the effects of toluene. Therefore, quercetin-containing plants could be regulators of porcine reproduction and natural protectors against the adverse effects of the environmental contaminant toluene.

Potential protective effect of puncturevine (Tribulus terrestris, L.) against xylene toxicity on bovine ovarian cell functions.

The action of the medicinal plant Tribulus terrestris (TT) on bovine ovarian cell functions, as well as the protective potential of TT against xylene (X) action, remain unknown. The aim of the present in vitro study was to elucidate the influence of TT, X and their combination on basic bovine ovarian cell functions. For this purpose, we examined the effect of TT (at doses of 0, 1, 10, and 100 ng/mL), X (at 20 ?g/mL) and the combination of TT + X (at these doses) on proliferation, apoptosis and hormone release by cultured bovine ovarian granulosa cells. Markers of proliferation (accumulation of PCNA), apoptosis (accumulation of Bax) and the release of hormones (progesterone, testosterone and insulin-like growth factor I, IGF-I) were analyzed by quantitative immunocytochemistry and RIA, respectively. TT addition was able to stimulate proliferation and testosterone release and inhibit apoptosis and progesterone output. The addition of X alone stimulated proliferation, apoptosis and IGF-I release and inhibited progesterone and testosterone release by ovarian cells. TT was able to modify X effects: it prevented the antiproliferative effect of X, induced the proapoptotic action of X, and promoted X action on progesterone but not testosterone or IGF-I release. Taken together, our observations represent the first demonstration that TT can be a promoter of ovarian cell functions (a stimulator of proliferation and a suppressor of apoptosis) and a regulator of ovarian steroidogenesis. X can increase ovarian cell proliferation and IGF-I release and inhibit ovarian steroidogenesis. These effects could explain its anti-reproductive and cancer actions. The ability of TT to modify X action on proliferation and apoptosis indicates that TT might be a natural protector against some ovarian cell disorders associated with X action on proliferation and apoptosis, but it can also promote its adverse effects on progesterone release.

Plant isoflavones can affect accumulation and impact of silver and titania nanoparticles on ovarian cells.

Objectives. The application of nanoparticles is experiencing a rapid growth, but it faces a problem of their toxicity, especially adverse effects on female reproduction. Food and medicinal plants and their isoflavones can be protectors against environmental stressors, but their ability to abate the adverse effects of nanoparticles has not been studied yet. In the present study, we examined the effect of silver (AgNPs) and titanium dioxide (titania, TiO2NPs) nanoparticles alone or in combination with plant phytoestrogens/antioxidants (resveratrol, diosgenin, and quercetin) on accumulation of nanoparticles, and progesterone release by cultured porcine ovarian granulosa cells.Methods. Porcine granulosa cells were incubated in the presence of AgNPs or TiO2NPs (0.1, 1, 10 or 100 µg/ml) alone or in combination with resveratrol, diosgenin or quercetin (10 µg/ml) for 48 h. The accumulation of tested nanoparticles by granulosa cells was assessed under light microscope. Progesterone concentration in culture media was measured by ELISA kit.Results. Cells accumulated both AgNPs and TiO2NPs in a dose-dependent manner. AgNPs, but not TiO2NPs, at highest dose (100 µg/ml) resulted in a destruction of cell monolayer. Both Ag-NPs and TiO2NPs reduced progesterone release. Resveratrol, diosgenin, and quercetin promoted accumulation of both AgNPs and TiO2NPs in ovarian cells and inhibited the progesterone output. Furthermore, resveratrol and diosgenin, but not quercetin, prevented the suppressive action of both AgNPs, and TiO2NPs on progesterone release.Conclusions. These observations (1) demonstrate accumulation of AgNPs and TiO2NPs in ovarian cells, (2) confirm the toxic impact of AgNPs, and TiO2NPs on these cells, (3) confirm the inhibitory effects of plant polyphenols/phytoestrogens on ovarian steroidogenesis, (4) show the ability of these isoflavones to increase the accumulation of AgNPs and TiO2NPs, and (5) show their ability to reduce the suppressive effect of AgNPs and TiO2NPs on ovarian progesterone release. The suppressive effect of AgNPs and TiO2NPs on ovarian functions should be taken into account by their exposition. However, these adverse effects could be mitigated by some plant isoflavones.

PSII-32 In vitro effect of growth hormone on progesterone production by large preovulatory follicles depends on the hen age and follicular layers interaction

Abstract Growth hormone (GH) is an endocrine and paracrine/autocrine regulator of the hen ovarian function, with the GH receptor concentration in the granulosa layer (GL) being maximum in the largest preovulatory follicle (Lebedeva et al. 2004, Biol.Reprod. 71:1174–1181). In the present study, GH effects on in vitro production of progesterone by GL from the two largest yellow follicles (F1 and F2) were investigated due to the hen age and the presence of the theca layer (TL). Young hens with long clutch (YLC, 32–33 week-old, &amp;gt;10 eggs per clutch) and old hens with short clutch (OSC, 74–76 week-old, 3–6 eggs per clutch) were used. After isolation, GL from F1 and F2 follicles (n = 8–9) was cultured separately or jointly with the respective TL for 18 h in the presence or absence of chicken GH (25 ng/ml). Concentrations of progesterone in culture media were measured by ELISA. The data were analyzed by repeated measures ANOVA. When GL from F1 follicle cultured alone, GH did not affect progesterone production in YLC hens and decreased it from 30,5±3,4 to 20,5±2,9 ng/mg tissue (P &amp;lt; 0.01) in OSC hens. Conversely, when tested GL from F2 follicle, GH increased progesterone output from 15,8±2,4 to 20,4±2,5 ng/mg tissue (P &amp;lt; 0.05) in YLC birds and had no effect on the output in OSC birds. During co-culture of GL and TL, GH raised 1.4–1.5 times the production of progesterone in the case of F1 follicle and did not change it in the case of F2 follicle in hens of both ages. The findings indicate that the steroidogenic response of GL from the two largest preovulatory follicles to GH differs in young and old hens. However, the interaction with TL modifies the GL response and makes it similar in birds regardless the age and reproductive status. The study was supported by RFBR (19-016-00216).

Hypoxia-inducible factor (HIF1alpha) inhibition modulates cumulus cell function and affects bovine oocyte maturation in vitro†.

Various metabolic and hormonal factors expressed in cumulus cells are positively correlated with the in vitro maturation (IVM) of oocytes. However, the role of hypoxia sensing both during maturation of cumulus–oocyte complexes (COCs) as well as during the resumption of meiosis remains uncertain. HIF1alpha plays major roles in cellular responses to hypoxia, and here we investigated its role during bovine COC maturation by assessing the expression of related genes in cumulus cells. COCs were divided into the following groups: immature (control), in vitro matured (IVM/control), or matured in the presence of a blocker of HIF1alpha activity (echinomycin, IVM/E). We found an inhibition of cumulus cell expansion in IVM/E, compared with the IVM/control. Transcript levels of several factors (n = 13) were assessed in cumulus cells. Decreased expression of HAS2, TNFAIP6, TMSB4, TMSB10, GATM, GLUT1, CX43, COX2, PTGES, and STAR was found in IVM/E (P < 0.05). Additionally, decreased protein levels were detected for STAR, HAS2, and PCNA (P < 0.05), while activated-Caspase 3 remained unaffected in IVM/E. Progesterone output decreased in IVM/E. The application of PX-478, another blocker of HIF1alpha expression, yielded identical results. Negative effects of HIF1alpha suppression were further observed in the significantly decreased oocyte maturation and blastocyst rates from COCs matured with echinomycin (P < 0.05) or PX-478 (P < 0.05). These results support the importance of HIF1alpha for COC maturation and subsequent embryo development. HIF1alpha is a multidirectional factor controlling intercellular communication within COCs, steroidogenic activity, and oocyte development rates, and exerting effects on blastocyst rates.

HCG Improves Luteal Function and Promotes Progesterone Output through the Activation of JNK Pathway in the Luteal Granulosa Cells of the Stimulated IVF Cycles†.

JNK signaling pathway plays a central role in the upregulated expression of the steroidogenic enzymes StAR and 3b-HSD and augmented progesterone production by hCG/LH in human luteal granulosa cells.

Melatonin effects on ovarian follicular cells: a systematic review.

Melatonin is known for its effects on both the sleep and reproductive system of mammals. The latter has melatonin receptors type 1 and 2, which act to regulate, among other things, cyclic AMP. Notwithstanding all the literature data, there is still no sound knowledge or a clear understanding of the hormone's action on the physiology of ovarian follicular cells. OBJECTIVE To review and evaluate studies about melatonin action on the ovarian granulosa/theca interna cells from the literature. METHODS The systematic review was carried out according to the PRISMA recommendations. The MEDLINE and Cochrane primary databases were consulted with the use of specific terms. There was no limitation on language or publication year. RESULTS Seven papers about melatonin action on granulosa cells were selected. The following can be attributed to the hormone's effects: a) progesterone increase in culture medium; b) increased estrogen production; c) antagonistic action on estrogen; d) improvement in cell quality resulting in improved embryo and higher pregnancy rates; e) improved cell proliferation via MAPK; f) reduction of free radicals. Nevertheless, there are contrarian papers reporting a reduction in progesterone production. Melatonin interferes in sex steroid production, boosting progesterone output. Such action may help improve oocyte quality.

The involvement of the phosphorylatable and nonphosphorylatable transcription factor CREB-1 in the control of human ovarian cell functions

The objective of our study was to elucidate the role of the transcription factor CREB-1 in controlling ovarian cell proliferation, apoptosis, and hormone release and the significance of CREB-1 phosphorylation in these processes. Human ovarian granulosa cells were transfected with a gene construct encoding wild-type CREB-1 (CREB-1 WT) or CREB-1 nonphosphorylatable mutant (CREB-1 M1). The expression of total and phosphorylated CREB-1, markers of proliferation (PCNA) and apoptosis (bax), as well as the release of progesterone, oxytocin, prostaglandin F2 alpha (PGF2), prostaglandin E2 (PGE2), and insulin-like growth factor I (IGF-I) were compared by immunocytochemistry, enzyme immunoassay (EIA), and immunoradiometric assay (IRMA). Transfection with CREB-1 WT or CREB-1 M1 increased total CREB-1 expression and proliferation and decreased the release of oxytocin, PGE2, and IGF-I by ovarian cells. CREB-1 M1, not CREB-1 WT, promoted apoptosis and inhibited progesterone output. PGF2 release was inhibited by CREB-1 WT but stimulated by CREB-1 M1 construct. Phosphorylated CREB-1 was undetected in any cell group. These observations confirm the involvement of CREB-1 in the control of ovarian cell proliferation, apoptosis, and steroid hormone release. This is the first demonstration of the involvement of CREB-1 in the regulation of the ovarian non-steroidal hormones such as oxytocin, PGF2, PGE2, and IGF-I. The absence of CREB-1 phosphorylation, similar effects exerted by CREB-1 WT and CREB-1 M1 on cell proliferation and release of oxytocin, PGE2, and IGF-I, and the influence of CREB-1 M1 on apoptosis and progesterone suggest that phosphorylation plays no role in the action of CREB-1 on the majority of analyzed functions of human ovarian cells.

Systematic Review of Ovarian Activity and Potential for Embryo Formation and Loss during the Use of Hormonal Contraception.

For this study, the authors looked at English-language research articles that focused on how hormonal birth control, such as the birth control pill, may affect very early human embryos. The authors found that abnormal ovulation, or release of an egg followed by abnormal hormone levels, may often occur in women using hormonal birth control. This may increase the number of very early human embryos who are lost before a pregnancy test becomes positive. For women who are thinking about using hormonal birth control, this is important information to consider.

FSH promotes premature progesterone output in human granulosa cells without luteinization by up-regulating the expression of 3β-HSD and inducing disproportional increases between 17α-OH and other steroidogenic enzymes

FSH promotes premature progesterone output in human granulosa cells without luteinization by up-regulating the expression of 3β-HSD and inducing disproportional increases between 17α-OH and other steroidogenic enzymes

Temporary undernutrition during early gestation, corpora lutea morphometrics, ovarian progesterone secretion and embryo survival in gilts.

The present study reports effects of severe undernutrition on luteal function and pregnancy in pigs. Gilts were inseminated and either fasted on Day 10 and 11 after conception (n=11) or fully fed throughout (n=10). Fasting did not affect LH or progesterone pulsatile secretion pattern on Day 11 in samples taken from blood vessels draining an ovary. Ultrasonographic measurements of the size of the corpora lutea did not show any effect of fasting either. However, fasted gilts had 10 to 30% lower systemic progesterone from Day 12 through Day 15 after conception (P<0.05). All gilts farrowed, but fasted gilts had fewer born piglets than fully fed gilts (8.8±0.8 vs 10.9±0.5 respectively; P<0.05). In conclusion, fasting during embryo elongation can compromise embryonic survival by affecting ovarian function in the days after fasting, without having an immediate effect on LH secretion and progesterone output by the ovaries.

Effects of in vitro exposure to butylparaben and di-(2 ethylhexyl) phthalate, alone or in combination, on ovarian function.

Parabens and phthalates are commercial chemicals widely used in the manufacture of industrial and consumer products frequently found as contaminants in biological fluids. We evaluated the effects of di-(2-ethylhexyl) phthalate (DEHP) (ranging from 10(-9) to 10(-7) m [1-100 nm; 0.39-39 ng ml(-1) ]) and butylparaben (BP) (ranging from 10(-8) to 10(-5) m [10 nm-10 μm; 1.9 ng ml(-1) to 1.9 μg ml(-1) ]), alone and in combination, on isolated mouse preantral follicle and human granulosa cell (hGC) cultures to study direct effects on follicle growth and ovarian steroidogenesis. Our results revealed that, in follicle culture, DEHP and BP attenuate estradiol output but only when present together. DEHP decreases progesterone concentrations in the spent media of hGC cultures, an effect that was attenuated when BP was added together with DEHP. Although changes in steroidogenesis were observed, no effects on follicular development or survival were noted in the culture systems. We suggest that BP and DEHP act with additive effect to decrease estradiol production whereas at later stages of follicle development BP blocks the effect of DEHP in hGCs resulting in decreased progesterone output. Taken together our results suggest that DEHP and BP adversely affect steroidogenesis from the preantral stage onward and the effects of these chemicals are both stage-dependent and modified by co-exposure. Copyright © 2016 John Wiley & Sons, Ltd.

Aging influences steroid hormone release by mink ovaries and their response to leptin and IGF-I.

ABSTRACTThe aim of our study was to understand whether ovarian steroid hormones, and their response to the metabolic hormones leptin and IGF-I leptin, could be involved in the control of mink reproductive aging via changes in basal release of ovarian progesterone and estradiol. For this purpose, we compared the release of progesterone and estradiol by ovarian fragments isolated from young (yearlings) and old (3-5 years of age) minks cultured with and without leptin and IGF-I (0, 1, 10 or 100 ng/ml). We observed that isolated ovaries of older animals produced less progesterone but not less estradiol than the ovaries of young animals. Leptin addition stimulated estradiol release by the ovarian tissue of young animals but inhibited it in older females. Leptin did not influence progesterone output by the ovaries of either young or older animals. IGF-I inhibited estradiol output in young but not old animals, whereas progesterone release was inhibited by IGF-I irrespective of the animal age. Our observations demonstrate the involvement of both leptin and IGF-I in the control of mink ovarian steroid hormones release. Furthermore, our findings suggest that reproductive aging in minks can be due to (a) reduction in basal progesterone release and (b) alterations in the response of estradiol but not of progesterone to leptin and IGF-I.

Pregnancy losses in cattle: potential for improvement.

For heifers, beef and moderate-yielding dairy cows, it appears that the fertilisation rate generally lies between 90% and 100%. For high-producing dairy cows, there is a less substantive body of literature, but it would appear that the fertilisation rate is somewhat lower and possibly more variable. In cattle, the major component of embryo loss occurs in the first 16 days following breeding (Day 0), with emerging evidence of greater losses before Day 8 in high-producing dairy cows. In cattle, late embryo mortality causes serious economic losses because it is often recognised too late to rebreed females. Systemic concentrations of progesterone during both the cycle preceding and following insemination affect embryo survival, with evidence of either excessive or insufficient concentrations being negatively associated with survival rate. The application of direct progesterone supplementation or treatments to increase endogenous output of progesterone to increase embryo survival cannot be recommended at this time. Energy balance and dry matter intake during the first 4 weeks after calving are critically important in determining pregnancies per AI when cows are inseminated at 70-100 days after calving. Level of concentrate supplementation of cows at pasture during the breeding period has minimal effects on conception rates, although sudden reductions in dietary intake should be avoided. For all systems of milk production, more balanced breeding strategies with greater emphasis on fertility and feed intake and/or energy must be developed. There is genetic variability within the Holstein breed for fertility traits, which can be exploited. Genomic technology will not only provide scientists with an improved understanding of the underlying biological processes involved in fertilisation and the establishment of pregnancy, but also, in the future, could identify genes responsible for improved embryo survival. Such information could be incorporated into breeding objectives in order to increase the rate of genetic progress for embryo survival. In addition, there is a range of easily adoptable management factors, under producer control, that can either directly increase embryo survival or ameliorate the consequences of low embryo survival rates. The correction of minor deficits in several areas can have a substantial cumulative positive effect on herd reproductive performance.

Tumor necrosis factor-α inhibits the stimulatory effect of luteinizing hormone and prostaglandin E 2 on progesterone secretion by the bovine corpus luteum

Tumor necrosis factor-α (TNF-α) is involved in the tissue remodeling that occurs in the corpus luteum (CL) during its development and regression. This cytokine is also implicated in the regulation of reproduction by its actions on ovarian steroidogenic cells. The aim of this study was to examine the influence of TNF-α on (1) progesterone (P 4) output by the bovine CL and on (2) the responsiveness of the CL to LH or prostaglandin E 2 (PGE 2) in vitro. In experiment 1, CL (days 8 to 10 of the estrous cycle) were perfused by using an in vitro microdialysis system with TNF-α (0.1, 0.5, or 1 μg/mL) alone or with TNF-α (1 μg/mL) followed by LH (1000 ng/mL) or PGE 2 (2 × 10 −5 M). Basal P 4 release ( P < 0.05) was increased by TNF-α (0.5 or 1 μg/mL). Moreover, TNF-α (1 μg/mL) inhibited the stimulatory effect of LH or PGE 2 on P 4 output ( P < 0.05). In experiment 2, 4 h after intrauterine infusion of TNF-α (0.01 μg/mL or 1 μg/mL), CL (days 8 to 10 of the estrous cycle) were collected by colpotomy, cultured, and stimulated with LH (10 ng/mL) or PGE 2 (10 −6 M). Intrauterine infusion of TNF-α at a concentration of 1 μg/mL increased basal P 4 output by CL ( P < 0.05). Moreover, the intrauterine infusion of TNF-α at a concentration of 0.01 μg/mL inhibited the stimulatory effect of LH or PGE 2 on P 4 output ( P < 0.05). These results indicate that TNF-α (1) does not have an effect on the autonomous, pulsatile release of P 4; (2) increases P 4 secretion by bovine CL with increasing doses, and (3) reduces in a dose-dependent manner the responsiveness of CL to luteotropic factors both directly (after infusion to CL) and indirectly (after intrauterine infusion).

Effect of in vitro exposure to benzo[a]pyrene, a component of cigarette smoke, on folliculogenesis, steroidogenesis and oocyte nuclear maturation

We previoulsy quantified the concentration of benzo[a]pyrene (B[a]P) in the follicular fluid of women exposed to mainstream and/or sidestream cigarette smoke. The objective of this study was to quantify the effects of B[a]P-exposure, at concentrations representative of follicluar fluid concentrations, on folliculogenesis, on gonadal steroid and anti-müllerian hormone (AMH) output, oocyte growth, and nuclear maturation. Follicles (100–130 μm) isolated from ovaries of F1 hybrid (C57BL/6j×CBA/Ca) mice were cultured for 13 days in increasing concentrations of B[a]P (0 ng/ml (control) to 45 ng/ml). B[a]P treatment inhibited ( p < 0.05) antral follicle development, decreased estradiol output and follicle survival at the 45.0 ng/ml dose. B[a]P exposure decreased AMH output overall during preantral ( p = 0.014) and antral ( p = 0.026) follicle development but had no effect on progesterone output or oocyte growth and nuclear maturation in surviving follicles. These data suggest that B[a]P is an important toxic component of cigarette smoke that adversely affects follicular development and survival.