Prostaglandin-endoperoxide synthase (PTGS; EC
1.14.99.1) is the rate-limiting step in the conversion of arachidonic acid to
prostaglandins is catalyzed by PTGS. Two isoforms of PTGS have been identified:
PTGS1 (COX1), encoded by a 2.8-kb mRNA, and a mitogen-inducible form,
PTGS2, encoded by a 4.5-kb mRNA . Both
PTGS1 and PTGS2 are expressed in cells involved in inflammatory processes.
Dramatic induction of PTGS2 mRNA in normal peripheral blood monocytes has been
documented in response to lipopolysaccharide (LPS) and phorbol myristate acetate
(PMA).///////////In addition to PGF2 acting through the PGF receptor, there are four paralogous receptors (EP1-4) for PGE2. ///////Novel contraceptive targets to inhibit ovulation: the prostaglandin E2 pathway. Duffy DM et al. (2015) Prostaglandin E2 (PGE2) is an essential intrafollicular regulator of ovulation. In contrast with the one-gene, one-protein concept for synthesis of peptide signaling molecules, production and metabolism of bioactive PGE2 requires controlled expression of many proteins, correct subcellular localization of enzymes, coordinated PGE2 synthesis and metabolism, and prostaglandin transport in and out of cells to facilitate PGE2 action and degradation. Elevated intrafollicular PGE2 is required for successful ovulation, so disruption of PGE2 synthesis, metabolism or transport may yield effective contraceptive strategies. This review summarizes case reports and studies on ovulation inhibition in women and macaques treated with cyclooxygenase inhibitors published from 1987 to 2014. These findings are discussed in the context of studies describing levels of mRNA, protein, and activity of prostaglandin synthesis and metabolic enzymes as well as prostaglandin transporters in ovarian cells. The ovulatory surge of LH regulates the expression of each component of the PGE2 synthesis-metabolism-transport pathway within the ovulatory follicle. Data from primary ovarian cells and cancer cell lines suggest that enzymes and transporters can cooperate to optimize bioactive PGE2 levels. Elevated intrafollicular PGE2 mediates key ovulatory events including cumulus expansion, follicle rupture and oocyte release. Inhibitors of the prostaglandin-endoperoxide synthase 2 (PTGS2) enzyme (also known as cyclooxygenase-2 or COX2) reduce ovulation rates in women. Studies in macaques show that PTGS2 inhibitors can reduce the rates of cumulus expansion, oocyte release, follicle rupture, oocyte nuclear maturation and fertilization. A PTGS2 inhibitor reduced pregnancy rates in breeding macaques when administered to simulate emergency contraception. However, PTGS2 inhibition did not prevent pregnancy in monkeys when administered to simulate monthly contraceptive use. PTGS2 inhibitors alone may be suitable for use as emergency contraceptives. However, drugs of this class are unlikely to be effective as monthly contraceptives. Inhibitors of additional PGE2 synthesis enzymes or modulation of PGE2 metabolism or transport also hold potential for reducing follicular PGE2 and preventing ovulation. Approaches which target multiple components of the PGE2 synthesis-metabolism-transport pathway may be required to effectively block ovulation and lead to the development of novel contraceptive options for women. Therapies which target PGE2 may also impact disorders of the uterus and could also have benefits for women's health in addition to contraception.//////////////////
NCBI Summary:
Prostaglandin-endoperoxide synthase (PTGS), also known as cyclooxygenase, is the key enzyme in prostaglandin biosynthesis, and acts both as a dioxygenase and as a peroxidase. There are two isozymes of PTGS: a constitutive PTGS1 and an inducible PTGS2, which differ in their regulation of expression and tissue distribution. This gene encodes the inducible isozyme. It is regulated by specific stimulatory events, suggesting that it is responsible for the prostanoid biosynthesis involved in inflammation and mitogenesis. [provided by RefSeq, Feb 2009]
General function
Enzyme
Comment
Paracrine and autocrine regulation of EGF-like factors in cumulus oocyte complexes (COCs) and granulosa cells: key roles for prostanglandin synthase 2 (Ptgs2) and progesterone receptor (Pgr). Shimada M et al. The molecular bridges that link the LH surge with functional changes in cumulus cells that possess few LH receptors are being unraveled. Herein we document that EGF-like factors amphiregulin (Areg), epiregulin (Ereg) and betacellulin (Btc) are induced in cumulus oocyte complexes (COCs) by autocrine and paracrine mechanisms that involve the actions of prostaglandins (PGs) and progesterone receptor (PGR). Areg and Ereg mRNA and protein levels were reduced significantly in COCs and ovaries collected from prostagandin synthase 2 (Ptgs2) null mice and Pgr null (PRKO) mice at 4 h and 8 h post-hCG, respectively. In cultured COCs, FSH/forskolin induced Areg mRNA within 0.5 h that peaked at 4 h, a process blocked by inhibitors of p38MAPK (SB203580), MEK1 (PD98059) and PTGS2 (NS398) but not PKA (KT5720). Conversely, AREG but not FSH induced Ptsg2 mRNA at 0.5 h with peak expression of Ptgs2 and Areg mRNAs at 4 h, processes blocked by the EGF receptor tyrosine kinase inhibitor AG1478, PD98059 and NS398. PGE2 reversed the inhibitory effects of AG1478 on AREG induced expression of Areg but not Ptgs2, placing Ptgs2 downstream of EGF-R signaling. PMA and adenovirally expressed PGRA synergistically induced Areg mRNA in granulosa cells. In COCs AREG not only induced genes that impact matrix formation but also genes involved in steroidogenesis (StAR, Cyp11a1) and immune cell-like functions (Pdcd1, Runx1, Cd52). Collectively, FSH mediated induction of Areg mRNA via p38MAPK precedes AREG induction of Ptgs2 mRNA via ERK1/2. PGs acting via PTGER2 in cumulus cells provide a secondary, autocrine pathway to regulate expression of Areg in COCs showing critical functional links between G-protein coupled receptor and growth factor receptor pathways in ovulating follicles.
Efficacy evaluation of low-dose aspirin in IVF/ICSI patients evidence from 13 RCTs: A systematic review and meta-analysis. Wang L et al. (2017) We conducted a systematic review and meta-analysis of existing literature to evaluate the different outcomes of low-dose aspirin on patients undergoing in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI), including clinical pregnancy rate, implantation rate, live birth rate, miscarriage rate, fertilization rate, number of oocytes retrieved, and so forth. Electronic databases including PubMed, MEDLINE, and Embase were searched between 1997 and March 2016 to identity eligible studies. The following comparisons between treatment groups were included: aspirin versus placebo; aspirin versus control group; aspirin versus aspirin + prednisolone + control. Thirteen randomized controlled trials which included 3104 participants were selected. There were no significant differences in implantation rate (RR = 1.15; 95% CI = 0.78-1.70), live birth rate (RR = 1.06; 95% CI = 0.93-1.21), miscarriage rate (RR = 1.28; 95% CI = 0.93-1.77), fertilization rate (RR = 0.91; 95% CI = 0.75-1.11), and endometrial thickness (WMD = 0.15; 95% CI = -0.38-0.67). But the research showed that aspirin treatment may improve the clinical pregnancy rate (RR = 1.16; 95% CI = 1.04-1.28) compared to placebo or no treatment, and reduce the number of oocytes retrieved (WMD = -0.68; 95% CI = -0.91-0.46). Our findings suggest that low-dose aspirin may improve the pregnancy rate in IVF/ICSI, with the recommended clinical use dose of 100 mg/day. Considering the limitation of included studies, further well-designed large-scaled RCTs are necessary to clarify whether aspirin may improve assisted reproduction outcomes in IVF/ICSI patients.//////////////////
The COX-2 inhibitor meloxicam prevents pregnancy when administered as an emergency contraceptive to nonhuman primates. McCann NC 2013 et al.
OBJECTIVE
Cyclooxygenase-2 (COX-2) inhibitors reduce prostaglandin synthesis and disrupt essential reproductive processes. Ultrasound studies in women demonstrated that oral COX-2 inhibitors can delay or prevent follicle collapse associated with ovulation. The goal of this study was to determine if oral administration of a COX-2 inhibitor can inhibit reproductive function with sufficient efficacy to prevent pregnancy in primates.
STUDY DESIGN
The COX-2 inhibitor meloxicam (or vehicle) was administered orally to proven fertile female cynomolgus macaques using one emergency contraceptive model and three monthly contraceptive models. In the emergency contraceptive model, females were bred with a proven fertile male once 2?1 days before ovulation, returned to the females' home cage, and then received 5 days of meloxicam treatment. In the monthly contraceptive models, females were cocaged for breeding with a proven fertile male for a total of 5 days beginning 2?1 days before ovulation. Animals received meloxicam treatment (1) cycle days 5-22, or (2) every day, or (3) each day of the 5-day breeding period. Female were then assessed for pregnancy.
RESULTS
The pregnancy rate with meloxicam administration using the emergency contraception model was 6.5%, significantly lower than the pregnancy rate of 33.3% when vehicle without meloxicam was administered. Pregnancy rates with the three monthly contraceptive models (75%-100%) were not consistent with preventing pregnancy.
CONCLUSIONS
Oral COX-2 inhibitor administration can prevent pregnancy after a single instance of breeding in primates. While meloxicam may be ineffective for regular contraception, pharmacological inhibition of COX-2 may be an effective method of emergency contraception for women.
IMPLICATIONS
COX-2 inhibitors can interfere with ovulation, but the contraceptive efficacy of drugs of this class has not been directly tested. This study, conducted in nonhuman primates, is the first to suggest that a COX-2 inhibitor may be effective as an emergency contraceptive.
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Impact of the prostaglandin synthase-2 inhibitor celecoxib on ovulation and luteal events in women. Edelman AB et al. Jensen 2013;; BACKGROUND: Ovarian prostaglandins are critical in normal ovulation processes; thus, their inhibition may provide contraceptive benefits. This study was performed to determine the effect of the cyclooxygenase-2 (COX2) inhibitor celecoxib on ovulation and luteal events in women. STUDY DESIGN: The study had a randomized, double-blind, crossover design. Ovulatory, reproductive-aged women underwent ovarian ultrasound and serum hormone monitoring during four menstrual cycles (control cycle, treatment cycle 1, washout cycle, treatment cycle 2). Subjects received study drug (oral celecoxib 400 mg or placebo) either (a) once daily starting on cycle day 8 and continuing until follicle rupture or the onset of next menses if follicle rupture did not occur (pre-luteinizing hormone (LH) surge dosing) or (b) once daily beginning with the LH surge and continuing for 6 days (post-LH surge dosing). Subjects were randomly assigned to one of the above treatment schemes and received the other in the subsequent treatment cycle. The main outcomes were evidence of ovulatory and luteal dysfunction as determined by inhibited/delayed follicle rupture and reduced luteal progesterone synthesis or lifespan, respectively. RESULTS: A total of 20 women enrolled and completed the study (Group 1=10, Group 2=10), with similar demographics between groups. Nineteen subjects exhibited normal ovulation in the control cycle (one had a blunted LH peak). In comparison to control cycles, treatment cycles resulted in a significant increase in ovulatory dysfunction .pre-LH treatment: 30% (6/20), p=.04; post-LH treatment: 25% (5/20), p=.04. Mean peak progesterone, estradiol, and LH levels and luteal phase length did not differ significantly between control and either treatment cycle. CONCLUSIONS: Although treatment with celecoxib before or after the LH surge increases the rate of ovulatory dysfunction, most women ovulate normally. Thus, this selective COX2 inhibitor appears to be of limited usefulness as a potential emergency contraceptive.
Oral administration of the cyclooxygenase-2 (COX-2) inhibitor meloxicam blocks ovulation in non-human primates when administered to simulate emergency contraception. Hester KE et al. BACKGROUND Prostaglandins produced via cyclooxygenase-2 (COX-2) within the periovulatory follicle are required for successful ovulation. Inhibition of follicular prostaglandin synthesis prevents timely follicle rupture and oocyte release. This study was conducted to determine if a 5-day course of oral administration of the COX-2 inhibitor meloxicam can prevent ovulation while maintaining normal menstrual cycles in non-human primates. METHODS Adult female cynomolgus monkeys were studied in each of four sequential menstrual cycles. In Cycle 1, a serum sample was obtained each day and assayed for estradiol, progesterone and luteinizing hormone; first menses was also noted to establish parameters of a normal menstrual cycle for each animal. In Cycle 2, meloxicam was administered orally once each day for 5 days beginning at either mid follicular (n = 4), late follicular (n = 4) or periovulatory (n = 4) phase of the menstrual cycle; daily serum samples and menses were assessed as for Cycle 1. In Cycle 3, the follicle-bearing ovary was removed 2 days after the expected day of ovulation (n = 3-4/treatment group). In Cycle 4, monkeys received the 5-day courses of oral meloxicam as in Cycle 2 (n = 3-4/treatment group), and the remaining ovary was removed. Ovaries were examined for the presence of an oocyte within the follicle. RESULTS Monkeys had the expected levels of changing reproductive hormones during Cycle 1. Meloxicam treatment in Cycle 2 did not alter hormone levels or the luteal phase length. Follicles of ovaries removed during Cycle 3 did not contain oocytes, indicating successful ovulation. Follicles did contain oocytes after meloxicam treatment beginning in the mid follicular (67%), late follicular (100%) or periovulatory (50%) phase of Cycle 4, indicating failure of ovulation. CONCLUSIONS A 5-day course of oral meloxicam administered around the time of ovulation reduced the rate of oocyte release without alteration of reproductive hormones or menstrual cycle length. Meloxicam may be effective as an emergency contraceptive in women.
Inhibition of ovulation by a lipoxygenase inhibitor involves reduced cyclooxygenase-2 expression and prostaglandin E2 production in gonadotropin-primed immature rats. Kurusu S et al. Potential roles of cyclooxygenase (COX) pathway of arachidonic acid (AA) metabolism are established in a murine model of induced ovulation. Pharmacological inhibition of an alternative lipoxygenase (LOX) pathway has been shown to cause defective ovulation, but the mechanism is still undefined. This study investigated the effects of two LOX inhibitors and their time dependency on ovulation and COX activity in gonadotropins (eCG and human chorionic gonadotropin (hCG))-primed immature rats. Intra-ovarian bursal treatment with a general LOX inhibitor nordihydroguaiaretic acid (NDGA) at 0 h post-hCG (hCG0h) dose dependently inhibited ovulation rate. The drug was still but less effective when treated at hCG6h. A more specific inhibitor, 3,4-dihydroxyphenyl ethanol (DPE) was also inhibitory when treated at hCG0h but not at hCG6h. Interestingly, treatment with DPE at hCG0h resulted in attenuated expression of immunoreactive PTGS2 in granulosa layers and concomitant decrease in ovarian prostaglandin E(2) (PGE(2)) content at hCG8h. NDGA treatment reduced immunoreactive PTGS2. Ovulatory impairment by both inhibitors was prevented by systemic administration of PGE(2) at hCG6h. Immunohistochemistry revealed the expression of ALOX5 and ALOX12 in both thecal and granulosa layers of preovulatory follicles and, notably, the augmented immunoreactivities during 8 h after hCG treatment. Our results indicate the probable presence of multiple LOX isoforms and that specific inhibition of LOX at an early stage of hCG-signaling led to reduced PTGS2 activity and thus defective ovulation. They reveal a probable relationship between two pathways of AA metabolism and account at least partly for the mechanism by which the LOX inhibitor causes impaired ovulation.
Liu et al reported that prostaglandin G/H synthase-2 is expressed in bovine
preovulatory follicles after the endogenous surge of
luteinizing hormone.
PGHS-2 mRNA (4.0 kilobases) and protein (74000 Mr)
were absent in preovulatory follicles isolated at the onset of estrus, low in
follicles obtained 18 h after estrus (16.0 +/- 1.2 h post-LH surge), and
markedly induced 24 h postestrus (20 +/- 0 h post-LH surge).
Immunoblot and immunohistochemical analyses revealed that PGHS-2
protein was selectively induced in granulosa cells. Follicular fluid
concentrations of PGE2 and PGF2alpha increased significantly between 0 and 24 h after estrus.
37:
Tsai SJ, et al 1997 reported that prostaglandin F2alpha induces expression of prostaglandin G/H synthase-2 in the
ovine corpus luteum and a potential positive feedback loop during luteolysis.
Pall M, et al 2001 reported delayed follicular rupture in the human by the
selective COX-2 inhibitor rofecoxib: a randomized double-blind
study.
Thirteen healthy women,
30-40 years of age, without hormonal treatment and with regular menstrual cycles
(27-34 days), were given the selective COX-2 inhibitor rofecoxib (n = 6) or
placebo (n = 7) in a random double-blind fashion. In an initial control cycle,
serial hormonal analyses, detection of a measurable mid-cycle urine LH peak and
transvaginal ultrasound scans were performed to confirm normal ovulatory and
endocrinological cyclic patterns, in all participating women. During the
subsequent treatment cycle, serial ultrasound scans were performed. When the
dominant follicle reached 14-16 mm in diameter, 25 mg rofecoxib or placebo was
taken orally, once daily for 9 consecutive days, during which follicle size was
monitored daily by ultrasound scans and serial hormone analyses were performed.
Four of the six women who received rofecoxib demonstrated delayed
follicle rupture, >48 h after the LH peak, when compared with the placebo group,
who all had follicular rupture >36 h after the detected LH peak. No differences in
peripheral serum concentrations of progesterone, oestradiol, LH and FSH were
observed between placebo and rofecoxib groups, when analysed at specified time
intervals. This study suggests that selective COX-2 inhibition
has a negative, local effect on human ovulation, resulting in delayed follicular
rupture, without affecting peripheral hormonal cyclicity.
Effects of indomethacin on follicular structure, vascularity, and function over the periovulatory period in women. Athanasiou S et al. OBJECTIVE: To assess the effect of indomethacin (taken at defined times) on follicular rupture, indexes of intrafollicular blood flow, and steroidogenesis. PARTICIPANTS: Six healthy volunteers awaiting treatment for infertility by IVF-ET during subsequent natural cycles. INTERVENTIONS: All women were examined (at least every 8 hours) by transvaginal ultrasonography with color Doppler imaging and had samples of blood taken for hormone analysis. A self-test for urinary LH was performed before each scan. Indomethacin was first taken (50 mg three times per day) according to the maximum follicular diameter (first four women) or when the LH dipstick gave a positive result; the drug was taken for > or = 3 days. RESULTS: Follicular rupture was delayed in five of six cases (by 2 to 12 days). There was a reduction in intrafollicular peak systolic velocity before and after the positive urinary LH test compared with historical controls. Three follicles (50% of women) with the highest peak systolic velocity had an hemorrhagic appearance and persisted longer. There was no significant effect on menstrual cycle length or the levels of circulating FSH, E2, LH, or P. CONCLUSION: Indomethacin administered at the time of a positive self-test for urinary LH can delay follicular rupture with an associated reduction in intrafollicular blood flow but with no apparent effects on hormonal or menstrual status.
Cyclooxygenase-2 derived prostaglandin E2 directs oocyte maturation by differentially influencing multiple signaling pathways. Takahashi T et al. The process of oocyte maturation which impacts ovulation and fertilization is complex and requires an integration of the endocrine, paracrine, juxtacrine and autocrine signaling pathways. This process involves an intimate interaction between the oocyte and encircling cumulus cells within a follicle, a unique venue for somatic and germ cell communication. Cumulus cell expansion and resumption of meiosis with germinal vesicle breakdown (GVBD) are major events in oocyte maturation. Cyclooxygenase-2 (COX-2) derived prostaglandin E2 (PGE2) is a known critical mediator of oocyte maturation, but the diverse function of this lipid mediator in oocyte maturation, ovulation and fertilization has not been fully appreciated. We show here that gonadotropins in coordination with PGE2 signaling via its cell surface G-protein coupled EP2 and EP4 receptor subtypes directs cumulus cell expansion and survival, and oocyte meiotic maturation by differentially impacting PKA, MAPK, NF-B and PI3K/Akt pathways. This study is unique in the sense that it provides evidence for new site- and event-specific involvement of these signaling pathways under the influence of COX-2 derived PGE2 during the critical stages of this somatic-germ cell interaction, an absolute requirement for oocyte maturation.
Oocytes determine cumulus cell lineage in mouse ovarian follicles. Diaz FJ et al. The two principal functions of ovarian follicles are developmental and endocrine. The cumulus cells surrounding the oocyte are specialized to serve the development of the oocyte and steroidogenesis is a principal role of mural granulosa cells that line the follicle wall. The findings in this report demonstrate that oocytectomy or treatment with an inhibitor of SMAD2/3 activation results in decreased cumulus marker mRNA transcript levels and allows FSH to induce mural marker transcripts in cumulus cells. In addition, SMAD2/3 signaling is involved in enabling cumulus expansion and EGF-induced increases in Ptx3, Ptgs2 and Has2 mRNA levels. By contrast, follicle-stimulating hormone (FSH) stimulated expression of mural transcripts, but suppressed levels of cumulus transcripts. Thus, FSH and oocyte-stimulated SMAD2/3 signaling establish opposing gradients of influence in the follicle. These specify the mural and cumulus granulosa cell phenotypes that are pivotal for appropriate endocrine function and oocyte development.
Inhibition of cyclooxygenase-2 (COX-2) by meloxicam decreases the incidence of ovarian hyperstimulation syndrome in a rat model. Quintana R et al. OBJECTIVE: To investigate the effects of selective cyclooxygenase-2 (COX-2) inhibition on the ovarian hyperstimulation syndrome (OHSS) in an experimental model. DESIGN: Controlled laboratory study. SETTING: University-affiliated fertility center. ANIMAL(S): Female Wistar rats. INTERVENTION(S): Female Wistar rats (22 days old) were divided into four groups: group 1 (control group; n = 10) received 0.1 mL of intraperitoneal (IP) saline from days 22-26; group 2 (mild-stimulated group; n = 10) received 10 IU of pregnant mare serum gonadotropin (PMSG) on day 24 and 10 IU of hCG 48 hours later (day 26); group 3 (OHSS group; n = 10) was given 10 IU of PMSG for 4 consecutive days from day 22 and 30 IU hCG on the fifth day to induce OHSS; group 4 was treated the same as group 3, but received 2 muL (15 mg/mL) of meloxicam 2 hours before the PMSG injection for 4 consecutive days, and 2 hours before the hCG injection on the fifth day. All groups were killed on day 26. MAIN OUTCOME MEASURE(S): Number of antral and luteinized follicles, ovarian weight, semiquantitative vascular endothelial growth factor (VEGF) and COX-2 immunohistochemistry. RESULT(S): There were no differences in the ovarian weight between groups 1 and 2. Group 3 showed significantly increased ovarian weight that was suppressed, in group 4, by meloxicam. There was no difference in the number of antral follicles among the four groups. In the mild-stimulated and OHSS groups, the granulosa cells (GC) of preovulatory follicles and the stromal cells showed intense VEGF immunoreactivity. The ovaries from the meloxicam-treated group showed less immunoreactivity than the OHSS group, indicating diminished VEGF expression associated with meloxicam treatment. Group 3 (OHSS group) showed increased COX-2 immunoreactivity that was diminished in the meloxicam-treated group. Meloxicam treatment did not affect the hormone-induced increase in serum E(2) levels seen in OHSS rats. CONCLUSION(S): Our results in a rat model suggest that meloxicam has a beneficial effect on OHSS by reducing the increases in ovarian weight and VEGF expression associated with OHSS. These effects may be mediated by the COX-2 inhibitory capacity of meloxicam.
CCN2 Mediates S1P-Induced Upregulation of COX2 Expression in Human Granulosa-Lutein Cells. Hu LL et al. (2019) CCN1 and CCN2 are members of the CCN family and play essential roles in the regulation of multiple female reproductive functions, including ovulation. Cyclooxygenase-2 (COX2) is a critical mediator of ovulation and can be induced by sphingosine-1-phosphate (S1P) through the S1P1/3-mediated Yes-associated protein (YAP) signaling. However, it is unclear whether CCN1 or CCN2 can mediate S1P-induced upregulation of COX2 expression and increase in prostaglandin E2 (PGE2) production in human granulosa-lutein (hGL) cells. In the present study, we investigated the effects of S1P on the expressions of CCN1 and CCN2 in hGL cells. Additionally, we used a dual inhibition approach (siRNA-mediated silencing and small molecular inhibitors) to investigate the molecular mechanisms of S1P effects. Our results showed that S1P treatment significantly upregulated the expression of CCN1 and CCN2 in a concentration-dependent manner in hGL cells. Additionally, inhibition or silencing of S1P1, but not S1P3, completely abolished the S1P-induced upregulation of CCN2 expression. Furthermore, we demonstrated that S1P-induced nuclear translocation of YAP and inhibition or silencing of YAP completely abolished the S1P-induced upregulation of CCN1 and CCN2 expression. Notably, silencing of CCN2, but not CCN1, completely reversed the S1P-induced upregulation of COX2 expression and the increase in PGE2 production. Thus, CCN2 mediates the S1P-induced upregulation of COX2 expression through the S1P1-mediated signaling pathway in hGL cells. Our findings expand our understanding of the molecular mechanism underlying the S1P-mediated cellular activities in the human ovary.//////////////////
ATF4 Contributes to Ovulation via Regulating COX2/PGE2 Expression: A Potential Role of ATF4 in PCOS. Di F et al. (2018) Ovulatory disorder is common in patients with hyperprolactinemia or polycystic ovary syndrome (PCOS). Previous studies have shown that ATF4 plays critical role in apoptosis and glucose homeostasis, but its role in regulating reproductive function was not explored. The present study investigated the role of ATF4 in ovarian ovulatory function. Human granulosa cells (hGCs) from 48 women newly diagnosed with PCOS and 37 controls were used to determine ATF4 expression. In vitro cultured hGCs were used to detect the upstream and downstream genes of ATF4. A shRNA- Atf4 lentiviral vector (shAtf4) was injected into rat ovaries to establish an in vivo gene knockdown model to further assess the in vivo relevance of the results from PCOS women. We found that ATF4 expression was lower in hGCs from PCOS patients than in hGCs from non-PCOS women. Many pivotal transcripts involved in cumulus-oocyte complex (COC) expansion, extracellular matrix (ECM) remodeling, and progesterone production were significantly down-regulated after ATF4 knockdown. ChIP-qPCR assays indicated that ATF4 could directly bind to the COX2 promoter and that ATF4 knockdown could attenuate human chorionic gonadotropin (hCG)-induced COX2 expression and PGE2 production. The in vivo study showed that shRNA-lentivirus mediated Atf4 knockdown in rat ovaries led to reduced number of retrieved oocytes. Collectively, these findings suggested previously unknown roles of ATF4 in ovulation. Furthermore, ATF4 malfunction in PCOS patients may impact the ovulation process, which could contribute, in part, to the pathogenesis of PCOS.//////////////////
Activin A increases PTGS2 expression and PGE2 production in human granulosa-lutein cells. Liu PP et al. (2016) Activin A is one of the members of transforming growth factor-β superfamily that is expressed in human large luteal cells, and may act in an autocrine/paracrine manner to regulate luteal function. Prostaglandin-endoperoxide synthase 2 (PTGS2) enzyme and its derivative, prostaglandin E2 (PGE2), play significant roles in the regulation of corpus luteum formation and maintenance. To date, whether activin A can induce the expression of PTGS2 and the production of PGE2 in human granulosa-lutein cells is largely unknown. The aim of this study was to examine the effects of activin A on the regulation of PTGS2 expression and PGE2 production in human granulosa-lutein cells, and to investigate the underlying signal transduction mechanisms. In this study, the immortalized (SVOG cells) and primary human granulosa-lutein cells were used as the cell models. A TGF-β/activin type I receptor inhibitor, SB431542 and small interfering RNAs were used to investigate the activin A-induced downstream signaling pathway. We have demonstrated that activin A up-regulated the expression of PTGS2 and increased the production of PGE2 via an ACVR1B-mediated SMAD2/3-SMAD4 signaling pathway. Our results suggest that activin A may be involved in the modulation of human corpus luteum formation via the induction of PTGS2 expression and PGE2 production.//////////////////
Prokineticin ligands and receptors are expressed in the human fetal ovary and regulate germ cell expression of COX2. Eddie SL et al. (2015) Fetal ovarian development and primordial follicle formation underpin future female fertility. Prokineticin (PROK) ligands regulate cell survival, proliferation and angiogenesis in adult reproductive tissues including the ovary. However, their expression and function during fetal ovarian development remains unclear. To investigate expression and localization of the PROK ligands, receptors and their downstream transcriptional targets in the human fetal ovary. This study was conducted at the University of Edinburgh. Ovaries were collected from 37 morphologically normal human fetuses. mRNA and protein expression of PROK ligands and receptors was determined in human fetal ovaries using qRT-PCR, immunoblotting and immunohistochemistry. Functional studies were performed using a human germ tumour cell line (TCam-2) stably transfected with PROKR1. Expression of PROK1 and PROKR1 was significantly higher in mid-gestation ovaries (17-20 weeks) than at earlier gestations (8-11 and 14-16 weeks). PROK2 significantly increased across the gestations examined. PROKR2 expression remained unchanged. PROK ligand and receptor proteins were predominantly localised to germ cells (including oocytes within primordial follicles) and endothelial cells, indicating these cell types to be the targets of PROK signalling in the human fetal ovary. PROK1 treatment of a germ cell line stably-expressing PROKR1 resulted in ERK phosphorylation, and elevated COX2 expression. Developmental changes in expression and regulation of COX2 and pERK by PROK1 suggest that PROK ligands may be novel regulators of germ cell development in the human fetal ovary, interacting within a network of growth and survival factors prior to primordial follicle formation.//////////////////
TGF-? induces COX-2 expression and PGE2 production in human granulosa cells through Smad signaling pathways. Fang L 2014 et al.
Context: Cyclooxygenase-2 (COX-2) expression and prostaglandin E2 (PGE2) production have been shown to play key roles in the regulation of ovulation. The transforming growth factor-beta (TGF-? superfamily members are important molecules that regulate many ovarian functions under normal physiological and pathological conditions. TGF-? and its receptors are expressed in human granulosa cells. However, to date, whether TGF-? can regulate COX-2 expression and PGE2 production, which in turn contribute to the process of ovulation, remains unknown. Objective: To investigate the effects of TGF-? on COX-2 expression and PGE2 production in human granulosa cells. Design: SVOG cells are human granulosa cells that were obtained from women undergoing in vitro fertilization (IVF) and immortalized with SV40 large T antigen. SVOG cells were used to investigate the effect of TGF-? on COX-2 expression and PGE2 production. Setting: An academic research center. Main Outcome Measures: mRNA and protein levels were examined by RT-qPCR and western blotting, respectively. The concentrations of PGE2 in the culture medium were measured by enzyme-linked immunosorbent assay (ELISA). Results: TGF-? treatment induced COX-2 expression and PGE2 production. The inductive effects of TGF-? on COX-2 and PGE2 were abolished by inhibition of TGF-?type I receptor (T?I). In addition, treatment with TGF-? activated Smad2 and Smad3 signaling pathways. Inhibition of Smad signaling pathways by siRNA-mediated approaches attenuated TGF-?-induced COX-2 expression and PGE2 production. Conclusion: TGF-? induced PGE2 production by inducing COX-2 expression through a Smad-dependent signaling pathway in human granulosa cells.
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EGF-like growth factors induce COX-2-derived PGE2 production through ERK1/2 in human granulosa cells. Fang L 2013 et al.
Context:Aberrant regulation of ovulation is one of the major causes of infertility. In animal models, three epidermal growth factor (EGF)-like growth factors, amphiregulin (AREG), betacellulin (BTC) and epiregulin (EREG), have been shown to be involved in ovulation by regulating cyclooxygenase-2 (COX-2) expression and prostaglandin E2 (PGE2) production. However, whether the same is true in humans remains largely unknown.Objective:To investigate the effects of AREG, BTC and EREG on COX-2 expression and PEG2 production in human granulosa cells.Design:SVOG cells are human granulosa cells that were obtained from women undergoing in vitro fertilization (IVF) and immortalized with SV40 large T antigen. SVOG cells were used to investigate the effect of AREG, BTC and EREG on ovulation-related functions.Setting:Academic research center.Main Outcome Measures:Levels of mRNA and protein were examined by RT-qPCR and western blotting, respectively. The protein levels of PGE2 were measured by enzyme-linked immunosorbent assay (ELISA).Results:LH treatment up-regulated AREG, BTC and EREG and COX-2. Knockdown of EGFR attenuated LH-induced COX-2 expression and PGE2 production. Treatment with AREG, BTC and EREG up-regulated COX-2 expression and PGE2 production. The stimulatory effects of AREG, BTC and EREG on COX-2 expression and PGE2 production were blocked by inhibition of EGFR activity and expression. AREG-, BTC- and EREG-activated ERK1/2 signaling, but not Akt signaling, was required for AREG-, BTC- and EREG-induced COX-2 expression and PGE2 production.Conclusion:AREG, BTC and EREG induced PGE2 production by up-regulating COX-2 expression through ERK1/2 signaling in human granulosa cells.
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Androgen/Androgen Receptor Pathway Regulates Expression of the Genes for Cyclooxygenase-2 and Amphiregulin in Periovulatory Granulosa Cells. Yazawa T et al. It is well known that the androgen/androgen receptor (AR) pathway is involved in both male and female fertility in mammals. AR knockout female mice are reported to exhibit various abnormalities in follicle development, and a subfertile phenotype. In exogenous gonadotropin-induced superovulation, serum androgen levels were robustly elevated in female mice at the periovulatory stage after human chorionic gonadotropin (hCG) treatment. At this stage, ovarian AR proteins were strongly expressed in cumulus cells. Because these results suggested that the androgen/AR pathway is involved in ovulation, we investigated the expression of ovulation-related genes in the mouse ovary treated with the nonaromatizable androgen, 5a-dihydrotestosterone (DHT). DHT treatment induced the expression of the genes for cyclooxyganase-2 (Cox-2 or prostaglandin endoperoxidase synthase 2) and the epidermal growth factor-like factor, amphiregulin (Areg), in the ovary, whereas their hCG-induced expression was suppressed by the AR antagonist flutamide. These genes were also induced by DHT in AR-expressing primary granulosa and granulosa tumor-derived cells. Reporter assays, electrophoretic shift mobility assays and chromatin immunoprecipitation assays demonstrated that androgen response sequence(s) existing upstream of each gene were responsible for androgen responsiveness and were occupied by the AR in periovulatory granulosa cells. Our results suggest that the androgen/AR pathway is involved in the ovulatory process via expression of the Cox-2 and Areg genes in periovulatory granulosa cells.
Gonadotropin-induced expression of mRNA for cyclooxygenase-2 (COX-2) and production of prostaglandins E and F2{alpha} in bovine preovulatory follicles are regulated by the progesterone receptor. Bridges PJ et al. Follicular production of prostaglandins (PGs) is essential for ovulation, but the factors mediating gonadotropin-induced secretion of PGE and PGF2alpha remain largely unknown. We tested the hypothesis that gonadotropin-induced changes in progesterone and its receptor (PR) mediate the increase in periovulatory PGs. Heifers were treated with PGF2alpha and GnRH to induce luteolysis and the LH/FSH surge (ovulation occurs approximately 30 h post-GnRH). Since there are two increases in intrafollicular progesterone/PR mRNA during the bovine periovulatory period, we first examined the temporal pattern of PG production by follicles collected at 0, 3.5, 6, 12, 18, and 24 h post-GnRH. Although PGs did not increase in the follicular fluid until 24 h post-GnRH, acute secretion of PGs by follicle wall (theca + granulosa cells) was initiated by 18 h and had increased many-fold by 24 h post-GnRH. In vitro, FSH and LH induced dramatic transient increases in PG production by follicle wall and granulosa, but not theca, cells isolated from preovulatory follicles (0 h post-GnRH). PG accumulation peaked on day 2 of culture, mimicking the secretion pattern following a gonadotropin surge in vivo. In cultures of follicle wall and granulosa cells, the PR antagonist mifepristone (MIFE, 1 microM) inhibited LH-induced PG secretion and the progestin medroxyprogesterone acetate (MPA, 1 or 10 microM), but not the glucocorticoid dexamethasone (1 or 10 microM), overcame the effect of MIFE on PGs. Semi-quantitative RT-PCR revealed that MIFE inhibited LH-induced expression of COX-2 mRNA in granulosa cells in vitro. Again, treatment with MPA overcame the effect of MIFE. Together, these results provide strong evidence that periovulatory increases in COX-2 mRNA, PGE, and PGF2alpha are mediated by gonadotropin-induced increases in progesterone/PR, indicating that in some species there is an important functional relationship between these pathways in the ovulatory cascade. Morris JK, et al 1995 reported that luteinizing hormone induces prostaglandin endoperoxide synthase-2 and
luteinization in vitro by A-kinase and C-kinase pathways.
Narko K, et al 1997 reported the induction of cyclooxygenase-2 and prostaglandin F2alpha receptor expression by
interleukin-1beta in cultured human granulosa-luteal cells.
Ultrastructural morphology of hypomelanosis in equine cutaneous papilloma. Hamada M et al. The morphology of hypomelanosis occurring in experimentally induced equine papillomas was investigated. Histologically, dopa-positive functioning melanocytes were decreased in number from the basal layer in the epidermis. Electron-microscopically, melanogenic organelles in the melanocytes were degenerate and melanosomes were decreased in number and size. In addition, the melanocytes had some abnormal melanosomes including melanosome complexes and giant melanosomes. Some abnormal melanosomes were also present in the keratinocytes. The hypomelanosis seemed to be related to a disturbance in melanin synthesis and melanocytic-keratinocytic interaction in the epidermal melanin unit.
Joyce et al 2001 reported the regulation of prostaglandin-endoperoxide synthase 2 messenger ribonucleic acid expression in mouse granulosa cells during ovulation. COx2 is expressed in cumulus and mural granulosa cells and the levels are highest at 4h after hCG treatment of PMSG-primed rats.
Gene Silencing of Cyclooxygenase-2 mRNA by RNA Interference in Bovine Cumulus-Granulosa cells. Kobayashi SI et al. Inhibition of specific gene expression using RNA interference (RNAi) is a valuable tool for functional analysis of a target gene. However, there is little information available concerning RNAi for analysis of gene function in relation to the reproductive physiology of follicular cells in ruminants. Thus, the aim of this study was to evaluate the interfering effect of small interference RNA (siRNA) on expression of cyclooxygenase-2 (Cox-2) mRNA and prostagrandin F(2alpha) (PGF (2alpha)) production in bovine cumulus-granulosa (CG) cells. Bovine CG cells were collected from aspirated follicles and cultured. After reaching confluency, two experiments were conducted. In experiment 1, to investigate the effective concentration of siRNA, 0, 100, 250 and 500 pM of Cox-2 siRNA was introduced into the CG cells, respectively. After 24 h, the amount of Cox-2 mRNA expression was measured by RT-PCR and real-time PCR. In experiment 2, to investigate the time required for effective interference of siRNA and Cox-2 activity, 250 pM siRNA was introduced for 0, 3, 6, 12 and 24 h. After culture, the amount of Cox-2 mRNA expression was measured and the culture medium was collected to determine the PGF(2) (alpha) concentration by enzyme immunoassay. The Cox-2 mRNA expression was not affected by introduction of 100 pM siRNA into CG cells for 24 h, but 250 and 500 pM Cox-2 siRNA significantly reduced the Cox-2 mRNA expression. Moreover, the significant suppressive effect of 250 pM siRNA was observed 6h after introduction, and the reduction of mRNA expression by RNAi became more obvious over 12 h. On the other hand, the PGF(2alpha) concentration in the culture medium was not significantly different 12 h after siRNA introduction; however, the PGF(2alpha) concentration 24 h after siRNA introduction was significantly decreased compared with the control at the same time point. These results suggest that gene silencing of Cox-2 with siRNA is capable of analyzing the function and expression of specific genes in bovine CG cells.
Prostaglandin E2 as a regulator of germ cells during ovarian development. Bayne RA et al. Context: The formation of primordial follicles occurs during fetal life yet is critical to the determination of adult female fertility. Prior to this stage, germ cells proliferate, enter meiosis, and associate with somatic cells. Growth and survival factors implicated in these processes include activin A (INHBA), the neurotrophins BDNF and NT4 (NTF5) and MCL1. The prostaglandins have pleiotrophic roles in reproduction, notably in ovulation and implantation, but there are no data regarding roles for prostaglandins in human fetal ovarian development. Objective: To investigate a possible role for Prostaglandin E2 in human fetal ovary development. Design: In vitro analysis of ovarian development between 8 and 20 weeks gestation. Main Outcome Measure(s): The expression patterns of prostaglandin synthesis enzymes and the PGE2 receptors EP2 and EP4 in the ovary were assessed and downstream effects of PGE2 on gene expression analysed. Results: Ovarian germ cells express the prostaglandin synthetic enzymes COX2 and PTGES as well as the EP2 and EP4 receptors, whereas COX1 is expressed by ovarian somatic cells. Treatment in vitro with PGE2 increased the expression of BDNF mRNA 1.7+/-0.16 fold (P=0.004), INHBA mRNA 2.1+/-0.51 fold (P=0.04) and MCL1 mRNA 1.15+/-0.06 fold (P=0.04) but not that of OCT4, DAZL, VASA, NTF5 or SMAD3. Conclusions: These data indicate novel roles for PGE2 in the regulation of germ cell development in the human ovary and that these effects may be mediated by the regulation of factors including BDNF, activin A and MCL1.
Follicle stages
Preovulatory, Corpus luteum
Comment
The development of the corpus luteum (CL), which involves angiogenesis, is essential for the establishment of early pregnancy. Sakurai T,et al investigated the roles of the prostaglandin synthases cyclooxygenase (COX) I and COX-II in angiogenesis and progesterone production in the newly formed CL, using inhibitors of the COX enzymes and the gonadotropin-induced pseudopregnant rat as a model. Injection of indomethacin, a nonselective COX inhibitor, on the day of ovulation and the following day decreased serum levels of progesterone, as did injection of the selective COX-II inhibitor NS-398. In contrast, a selective COX-I inhibitor, SC-560, had no effect on serum progesterone concentrations. None of the inhibitors had any effect on the weight of the superovulated ovaries or on the synthesis of progesterone by cultured luteal cells. To determine whether changes in angiogenesis are responsible for the decrease in progesterone synthesis, we measured hemoglobin and CD34 levels in luteinized ovaries following injection of COX inhibitors and measured the relative frequency of cells positive for platelet-endothelial cell adhesion molecule as a specific marker for endothelial cells. All of these parameters were reduced by the COX-II inhibitors, suggesting that changes in the vasculature are responsible for the decrease in serum progesterone. Histological examination of ovarian corrosion casts indicated that NS-398 inhibited the establishment of luteal capillary vessels following the injection of hCG. The results are consistent with the hypothesis that the activity of COX-II is associated with the formation of functional CL via its stimulation of angiogenesis.
Phenotypes
Mutations
3 mutations
Species: mouse
Mutation name: None
type: null mutation fertility: infertile - ovarian defect Comment: Multiple female reproductive failures in cyclooxygenase 2-deficient mice. Lim H et al. Cyclooxygenase (COX) is the rate-limiting enzyme in the synthesis of prostaglandins (PGs) and exists in two isoforms, COX-1 and COX-2. In spite of long-standing speculation, definitive roles of PGs in various events of early pregnancy remain elusive. We demonstrate herein that the targeted disruption of COX-2, but not COX-1, in mice produces multiple failures in female reproductive processes that include ovulation, fertilization, implantation, and decidualization. Using multiple approaches, we conclude that these defects are the direct result of target organ-specific COX-2 deficiency but are not the result of deficiency of pituitary gonadotropins or ovarian steroid hormones, or reduced responsiveness of the target organs to their respective hormones. COX-2(−/−) female mice are infertile and this was attributed to ovulation failure in spite of apparently normal follicular development ([9]). We reexamined in more detail the ovulation and subsequent events in these mice. As shown in Figure 1A, 64% (47/73, n = 11) of eggs recovered from wild-type females were fertilized. In contrast, only 19 eggs were recovered from 5 of 7 COX-2(−/−) females and none of them was fertilized. These eggs were developmentally abnormal, since extrusion of the first polar body was rarely observed. The results establish that COX-2(−/−) females have defective ovulation and complete failure of fertilization. We surmised that these defects could be due to deficiency of pituitary gonadotropins and/or ovarian responsiveness to gonadotropins. Thus, COX-2(−/−) females were subjected to superovulation with gonadotropins. As shown in Figure 1A, an average of 36 eggs/mouse were recovered from all of the wild-type mice (n = 9) examined, while an average of only 9 eggs/mouse were recovered from 8 of 10 COX-2(−/−) mice. Although the number of ovulation was slightly improved in gonadotropin-stimulated COX-2(−/−) mice, the presence of numerous developing follicles suggested that the ovaries responded to gonadotropins (data not shown). Again, fertilization was severely compromised in COX-2(−/−) mice [1.4% in COX-2(−/−) versus 55% in wild-type]. These results establish that the reduced rate of ovulation was not the result of deficiency of pituitary gonadotropins. Further, the presence of numerous sperm in the oviduct suggested that the fertilization failure was not due to the absence of sperm at the site of fertilization (data not shown). Collectively, these results establish that COX-2 deficiency is the major cause of defective ovulation and fertilization. The induction of COX-2 in ovarian follicles by gonadotropins ([44 and 43]) is also consistent with this conclusion. The cause of fertilization failure in COX-2(−/−) females could be due to impaired oocyte maturation.
Species: mouse
Mutation name: None
type: null mutation fertility: infertile - ovarian defect Comment: Anovulation in cyclooxygenase-2-deficient mice is restored by prostaglandin E2 and interleukin-1beta. Davis BJ et al. Mice carrying a null mutation for either of the two cyclooxygenase (COX) isoenzymes, necessary for prostanoid production, exhibit several isotype-specific reproductive abnormalities. Mice deficient in COX-1 are fertile but have decreased pup viability, whereas mice deficient in COX-2 fail to ovulate and have abnormal implantation and decidualization responses. The present study identifies the specific contribution of each COX isoenzyme in hypothalamic, pituitary, and ovarian function and establishes the pathology and rescue of the anovulatory syndrome in the COX-2-deficient mouse. In both COX-1- and COX-2-deficient mice, pituitary gonadotropins were selectively increased, whereas hypothalamic LHRH and serum gonadotropin levels were similar to those in wild-type animals (+/+). No significant differences in serum estrogen or progesterone were noted among the three genotypes. Exogenous gonadotropin stimulation with PMSG and hCG produced a comparable 4-fold increase in ovarian PGE2 levels in wild-type and COX-1(-/-) mice. COX-2(-/-) mice had no increase in PGE2 over PMSG-stimulated levels. Wild-type and COX-1(-/-) mice ovulated in response to PMSG/hCG; very few COX-2(-/-) animals responded to this regimen. The defect in ovulation in COX-2 mutants was attributed to both an abnormal cumulus oophorum expansion and subsequent stigmata formation. Gonadotropin stimulation and concurrent treatment with PGE2 or interleukin-1beta resulted in ovulation of COX-2(-/-) mice comparable to that in COX-2(+/+), whereas treatment with PGF2alpha was less effective. Collectively, these data demonstrate that COX-2, but not COX-1, is required for the gonadotropin induction of ovarian PG levels; that COX-2-related prostanoids are required for stabilization of the cumulus oophorum during ovulation; and that ovulation can be restored in the COX-2(-/-) animals by simultaneous treatment with gonadotropins and PGE2 or interleukin-1beta.
Species: None
Mutation name: None
type: None fertility: infertile - ovarian defect Comment: Maturation and Fertilization of nonhuman primate oocytes are compromised by oral administration of a cyclooxygenase-2 inhibitor. Duffy DM et al. OBJECTIVE: To determine if oral administration of a cyclooxygenase-2 (COX2) inhibitor affects oocyte nuclear maturation and fertilization in nonhuman primates. DESIGN: Laboratory research study. SETTING: Medical school. ANIMAL(S): Adult female cynomolgus monkeys (Macaca fascicularis). INTERVENTION(S): Monkeys received gonadotropins to stimulate multiple follicular development. An ovulatory dose of hCG was administered either alone or with oral celecoxib, a COX2 inhibitor. Oocytes were retrieved 36 hours later and exposed to sperm in vitro. MAIN OUTCOME MEASURE(S): Oocytes were assessed for nuclear status at retrieval, resumption of meiosis in vitro, and success of in vitro fertilization. RESULT(S): Treatment with hCG alone yielded oocytes that were primarily (72.9%) at the meiosis II (MII) stage of nuclear maturation; few oocytes were obtained at the germinal vesicle and germinal vesicle breakdown stages. Treatment with hCG and celecoxib yielded fewer mature (MII) oocytes (35.6%) and more oocytes at less mature stages compared with oocytes from monkeys treated with hCG alone. The majority (68.3 ? 15.9%) of MII oocytes from monkeys treated with hCG alone fertilized in vitro, compared with only 11.0 ? 5.9% of MII oocytes from monkeys treated with hCG and celecoxib. CONCLUSION(S): Oral administration of a COX2 inhibitor reduced the rate of oocyte nuclear maturation and the success of in vitro fertilization. Drugs of this class may block multiple essential steps in female reproduction and be effective contraceptives for women.