Altered maternal hypothalamic-pituitary-adrenal axis activity in obese pregnancy is associated with macrosomia and prolonged pregnancy.

BACKGROUND: The hypothalamic-pituitary-adrenal (HPA) axis is important for fetal growth and timing of parturition. Maternal obesity is associated with macrosomia (birthweight ⩾4000g) and prolonged pregnancy (⩾41weeks). We aimed to characterise HPA axis hormones in obese pregnancy and to test associations with these pregnancy outcomes. METHOD: Fasting cortisol was measured by radioimmunoassay in venous blood at 16, 28 and 36 weeks of gestation in 286 obese (BMI 44.05±3.98kg/m(2)) and 137 lean (BMI 22.71±1.66kg/m(2)) pregnant women. In subsets (n=20 obese, 20 lean) we measured corticosteroid binding globulin (CBG) and CRH by radioimmunoassay; progesterone, estradiol (E2), estriol (E3) and sex-hormone-binding-globulin (SHBG) by ELISA; and albumin by bromocresol green binding. Free cortisol levels were calculated using Coolen's equation. RESULTS: Cortisol, CBG, calculated free cortisol, CRH, E2, E3, progesterone and SHBG levels rose similarly during pregnancy in obese and lean, but were significantly lower in obese (p<0.05). In obese, lower free cortisol at 16 weeks was associated with higher birthweight (r=-0.46, p<0.05). Cortisol was not associated with labour onset. CRH was significantly lower at 36 weeks in women who delivered at ⩾41weeks and in women with macrosomic babies (p<0.05); and correlated negatively with gestation at delivery in obese (r=-0.557, p<0.05). CONCLUSION: Our findings suggest that decreased HPA axis activity in obese pregnancy may be a mechanism underlying macrosomia and prolonged pregnancy.

The effect of omega-3 polyunsaturated fatty acids on the inflammatory response of the amnion.

OBJECTIVE: To investigate the effect of omega-3 PUFAs, eicosapentanoic acid (EPA) and docosohexanoic acid (DHA) on inflammatory cytokine production in the amnion. STUDY DESIGN: Amnion explants were obtained at elective caesarean sections and cultured in vitro with EPA and DHA. IL-8 and IL-6 secretion was determined by ELISA, the role of PPARγ was investigated using specific agonists and antagonists and activity of MMP assessed by gelatin zymography. RESULTS: A combination of EPA and DHA significantly reduced the concentration of IL-8 and IL-6 released into the supernatant compared to untreated controls (p<0.001). Stimulation of PPARγ with troglitazone reduced IL-8 production, and the PPARγ antagonist GW9662 partially reversed this effect. The activity of MMP-9 was also significantly reduced by treatment with EPA and DHA in combination compared to untreated control (p<0.05). CONCLUSION: The omega-3 PUFAs EPA and DHA decrease the inflammatory response of the amnion, and this may be partially mediated through PPARγ.

Effects of N-acetylcysteine on matrix metalloproteinase-9 secretion and cell migration of human corneal epithelial cells.

BACKGROUND: Matrix metalloproteinase-9 (MMP-9) secreted by corneal epithelial cells has a role in the remodelling of extracellular matrix and migration of epithelial cells. Elevated levels of MMP-9 activity in the ocular surface may be involved in the pathogenesis of corneal diseases. N-acetylcysteine (NAC) has been used to treat corneal diseases, including recurrent epithelial erosions. In this study, its effects on the MMP-9 secretion and human corneal epithelial (HCE) cell migration were evaluated in vitro. METHODS: Confluent HCE cell cultures were treated with 0-20 mM NAC, and tested for MMP-9 secretion and epithelial cell migration by gelatin zymography and scratch wound assay, respectively. Comparisons between different treatment groups were made using analysis of variance, followed by multiple pairwise comparisons. RESULTS: Twenty mM NAC inhibited the secretion of MMP-9 significantly. Cell migration, assessed after 24 h of wounding, showed a highly significant dose-dependent inhibitory effect. CONCLUSIONS: This study shows that NAC reduces MMP-9 production by HCE cells and inhibits cell migration in vitro. This information helps to elucidate the mechanisms by which NAC may be beneficial therapeutically and suggests that NAC may be useful for managing corneal erosions and related conditions.

G-1-activated membrane estrogen receptors mediate increased contractility of the human myometrium.

Estrogens are key mediators of increased uterine contractility at labor. We sought to determine whether membrane-associated estrogen receptors, such as the recently described seven-transmembrane receptor G protein-coupled receptor 30 (GPR30), mediated some of this effect. Using human myometrium obtained at term cesarean section before or after the onset of labor, we demonstrated the presence of GPR30 mRNA and protein using quantitative RT-PCR and Western blotting. GPR30 receptor was localized to the cell membrane and often colocalized with calveolin-1. Using the specific estrogen membrane receptor agonist G-1 and myometrial explants, we showed that membrane receptor activation led to phosphorylation of MAPK and the actin-modifying small heat shock protein 27. Using myometrial strips incubated with G-1 or vehicle we demonstrated that estrogen membrane receptor activation increased the myometrial contractile response to oxytocin. These data suggest that activation of the plasma membrane estrogen receptor GPR30 likely participates in the physiology of the human myometrium during pregnancy and identifies it as a potential target to modify uterine activity.

Placental structure and inflammation in pregnancies associated with obesity.

BACKGROUND: Pre-gravid obesity is associated with increased morbidity and mortality for both mother and offspring. Recent studies have demonstrated a heightened inflammatory response both systemically and locally within the adipose and placental tissue in women with pre-gravid obesity, which may play a role in mediating the adverse pregnancy outcomes. The aim of this study was to characterise the maternal and placental inflammatory status and investigate associated changes in placental structure in obese women. METHODS: The pro-inflammatory status of a cohort of 47 non-obese (BMI 20-25 kg/m(2)) and 33 obese (≥30 kg/m(2)) women was characterised by measuring maternal circulating levels and placental gene expression of pro-inflammatory cytokines, and quantifying immune cell populations within the placenta. The effect of pre-gravid obesity on placental structure was investigated by examining placental maturity, vessel density, the formation of syncytial knots and sprouts, and the degree of fibrin deposition, chorangiosis and muscularisation of vessel walls. RESULTS: Maternal obesity was associated with significantly greater IL-1ß (p < 0.05), IL-8 (p < 0.05), MCP-1 (p < 0.001) and CXCR2 (p < 0.05) mRNA expression within the placenta and higher circulating maternal levels of IL-6 (3.30 ± 0.38 vs. 1.77 ± 0.15 pg/ml) (p < 0.001) compared with non-obese women. There were no differences in the number of CD14(+), CD68(+) cells or neutrophils within the placental villi of non-obese and obese women. However there were significantly higher numbers of neutrophils within the interstitial space (p < 0.05). Greater muscularity of placental vessel walls was associated with maternal obesity (p = 0.03), however no other associated structural changes were observed. CONCLUSIONS: Our findings show that although pre-gravid obesity was associated with greater expression of placental pro-inflammatory cytokines and higher circulating IL-6 in pregnancy, there were no major differences in immune cell populations within the placental villi and only a greater degree of muscularity in the vessel walls.

An investigation of the subjective experiences of sex after alcohol or drug intoxication.

Despite long-standing concern over the sexual health of the population there has been little work undertaken in the UK investigating sexual risk taking and sexual behaviours in the context of substance use. To investigate this further, 270 non-drug treatment seeking members of the public aged between 18 and 66 were administered a questionnaire containing the Alcohol Use Disorders Identification Test (AUDIT), the Drug Abuse Screening Test (DAST), the Severity of Dependence Scale (SDS), the Sexual Risks Scale and Attitudes toward condom use (SRSA), the Sexual Sensation Seeking Scale (SSSS); the Hospital Anxiety and Depression Scale (HADS), and questions pertaining to sexual episodes proximal to substance use. The population reported a varied history of substances and despite there not being self-awareness of problematic drug use, 39.4% reported above the cut-off mark of six on the DAST. An even greater percentage (57.8%) reported a score above eight on the AUDIT indicating hazardous or harmful drinking behaviour. The substance most often associated with sexual episodes was alcohol, followed by cannabis and ecstasy, and all were most frequently consumed in private houses. Sexual activity after drug use was most frequently circumstantial (i.e. the individual hadn't taken the substance for the specific purposes of sex), and was significantly associated with use of cannabis and ecstasy. The second most frequently reported association between drug use and sex was facilitation of a sexual encounter (i.e. to lower sexual inhibitions, increase self esteem and confidence), which was associated with use of alcohol, cannabis, cocaine and ecstasy. Although it was not possible to identify differences in subjective sexual changes after use of particular drugs, subjects reported that compared to sex after alcohol, sex on other drugs was more pleasurable and satisfying, with a greater perception of interpersonal contact with the partner and a greater willingness to sexually experiment. However, this latter change was not associated with changes in the type of sexual activity engaged in. Regression analysis revealed that the greatest subjective changes in sexual experiences were reported by younger participants who had ingested either ecstasy or cannabis prior to the sexual episode. These results are discussed in the context of sexual risk taking and suggest areas of intervention focus which may address substance use and sexual risk taking together.

Concentrations of inhibin, progesterone and oestradiol in fluid from dominant and subordinate follicles from mares during spring transition and the breeding season.

Dominant and subordinate follicles were collected from mares on the day after the dominant follicle reached 30 mm in diameter, to investigate regulation of folliculogenesis during spring transition and the breeding season. Concentrations of oestradiol-17beta, progesterone and inhibin A, but not inhibin isoforms with pro- and alpha C-immunoreactivity, were significantly higher in preovulatory follicles than in dominant anovulatory transitional follicles. Steroidogenic activity was regained gradually in the dominant follicles of successive anovulatory waves through spring transition. The dominant follicles, during both spring transition and cyclicity, contained higher concentrations of oestradiol, progesterone and inhibin A, but not inhibin pro- and alpha C-isoforms, than subordinate follicles. The results indicate that high follicular levels of oestradiol, progesterone and inhibin A are associated with continued follicle growth and ovulation. The low concentrations of oestradiol and progesterone in transitional follicles indicate that the deficiency in steroidogenesis exists early in the steroidogenic pathway. The similarity in patterns of follicular hormones in spring transition and during cyclicity strongly suggests that the mechanism of dominance is the same in both types of follicle.

Matrix metalloproteinases-2 and -9 and their endogenous tissue inhibitors in tissue remodeling after sealing of the fetal membranes in a sheep model of fetoscopic surgery.

OBJECTIVES: We studied collagen plugging of the fetoscopic access site in an in vivo fetal lamb model for fetoscopic surgery and possible role for matrix metalloproteinase (MMP)-2 and -9 and tissue inhibitors (TIMPs). METHODS: Eight ewes had fetoscopic balloon occlusion of the trachea as an experimental treatment for congenital diaphragmatic hernia between days 88 and 99 of gestation (term 145 days) with sampling of amniotic, allantoic, and tracheal fluid. Nonoperated cotwins were used as controls. The fetoscopy port was closed using a collagen plug. Ten days (range 9-12) later, fluids were sampled and plug sites collected for histologic analysis. Activity of MMP-2 (72 kDa, gelatinase A) and MMP-9 (92 kDa, gelatinase B) was determined in the fluids by zymography and secretion of TIMPs (27-30 kDa; TIMP-1, glycosylated TIMP-3 and TIMP-4, 24 kDa; unglycosylated TIMP-3, 21 kDa; TIMP-2) by reverse zymography and quantified by densitometric analysis. RESULTS: No pregnancy was complicated by amniorhexis or preterm labor. At cesarean, normal volumes of amniotic and allantoic fluid were present in all cases. Histology of the plug sites revealed good integration of the collagen plug without complete restoration of membrane integrity. MMP-2, MMP-9, and TIMPs were detected in all fluids. In the operated animals, significantly (P <.05) higher activity of MMP-9 was found in amniotic fluid, with lower concentrations of TIMPs in allantoic fluid (P <.01). Tracheal occlusion was associated with a significant (P <.02) increase in both MMP-2 and -9 in tracheal fluid. CONCLUSION: Collagen plugging of the fetoscopic access port sites in sheep resulted in functionally effective sealing of the fetal membranes. Changes in MMP-2, MMP-9, and TIMPs suggest an active remodeling of both the fetal lung and the fetal membranes.

Plasma FSH, inhibin A and inhibin isoforms containing pro- and -alphaC during winter anoestrus, spring transition and the breeding season in mares.

Ten mares were studied from February (winter anoestrus) to their second ovulation in the breeding season to investigate the relationship between resumption of ovarian cyclicity in the spring and circulating concentrations of FSH, inhibin A and inhibin isoforms containing pro- and -alphaC immunoreactivity. An additional four mares were studied during one oestrous cycle. Growth and regression of ovarian follicles were monitored by transrectal ultrasonography. The frequency of blood sampling varied from three times a week to once a day, depending on the follicular activity present. Concentrations of FSH, oestradiol, inhibin A and pro- and -alphaC isoforms were low during deep winter anoestrus when minimal follicular activity was present in the ovaries. During spring transition, an increase in FSH concentration preceded the emergence of each follicular wave. Concentrations of inhibins were significantly higher (P < 0.05) during growth of anovulatory follicles in spring transition than during winter anoestrus. Plasma concentrations of oestradiol and inhibin A were significantly higher (P < 0.001, P < 0.05, respectively) during the growth of preovulatory follicles than during the growth of transitional anovulatory follicles, but concentrations of inhibin pro-alphaC isoforms did not differ between the two types of follicle. During the oestrous cycle, there was a significant inverse relationship (P < 0.001) between concentrations of FSH and the inhibins. Plasma inhibin pro-alphaC isoforms, but not inhibin A, reached a peak on the day of ovulation. The results strongly indicate that FSH regulates growth of spring anovulatory and preovulatory follicles. Inhibins are likely to contribute to negative feedback on the release of FSH from the pituitary gland both during the transitional period and the breeding season in mares.

Effect of ascorbic acid on health and morphology of bovine preantral follicles during long-term culture.

During ovarian folliculogenesis, ascorbic acid may be involved in collagen biosynthesis, steroidogenesis and apoptosis. The aims of this study were to determine the effects of ascorbic acid on bovine follicle development in vitro. Preantral follicles were cultured for 12 days in serum-free medium containing ascorbic acid (50 microg ml(-1)). Half of the medium was replaced every 2 days, and conditioned medium was analysed for oestradiol and matrix metalloproteinase 2 (MMP-2) and MMP-9 secretion. On day 12, cell death was assessed by TdT-mediated dUTP-biotin nick end labelling (TUNEL). In the absence of serum, there was significant (P < 0.05) follicle growth and oestradiol secretion over the 12 day culture period. Ascorbic acid had no effect on these parameters. The addition of serum from day 0 stimulated follicle growth (P < 0.05), but compromised follicle integrity. By day 12 of culture, a higher proportion of follicles remained intact in the presence of ascorbic acid in serum-free conditions (P < 0.05), and significantly (P < 0.01) less granulosa and theca cell death was observed in these follicles than in control follicles. Moreover, ascorbic acid significantly (P < 0.05) increased production of MMP-9, an enzyme involved in basement membrane remodelling. In conclusion, this culture system was capable of supporting follicle differentiation over the 12 day culture period. Furthermore, ascorbic acid maintains bovine follicle health and basement membrane remodelling in vitro.

Patterns of recreational drug use at dance events in Edinburgh, Scotland.

AIMS: To describe the patterns of drug use at dance (rave) events in terms of prevalence, frequency, type of drugs used, patterns of use, access and risk-associated behaviours. DESIGN: Self-selecting participant-completed survey. SETTING: Three dance events in Edinburgh, Scotland, UK. PARTICIPANTS: One hundred and twenty-two drug users (57% males, 43% females), 90% of whom were in employment or education, with an age range of 16-47, 80% between 18 and 23 years. MEASUREMENTS: Participants who answered 'yes' to the question 'Have you used drugs for dance events in the past year' reported (i) the prevalence, types and frequency of drugs used; (ii) prevalence and contents of mixing drugs; (iii) accessing drugs; and (iv) engagement with drug-associated risk behaviours. FINDINGS: Over 80% of the participants had used ecstasy and amphetamine, over 30% cocaine and LSD; over 10% nitrites, psilocybin and ketamine and less than 5% had used crack or tranquillizers. Participants reported regular consumption of ecstasy and amphetamine (e.g. 35% used ecstasy and 25% amphetamine on a weekly basis) often taken in combination, with the occasional use of cocaine, LSD, ketamine and psilocybin. Poly- and mixing-drug behaviours were significantly more likely than monodrug usage. Drugs were accessed through friends than from any other source. Eighty-five per cent reported mixing drugs and/or alcohol, 35% driving on drugs, 36% having a bad experience on drugs; 30% unprotected sex; and 0.9% injecting drugs. Women in the sample reported higher consumption than men. CONCLUSIONS: Dance-drug use has a characteristic pattern that has implications for health promotion and criminal policy.

Co-localization of matrix metalloproteinase-1 and mast cell tryptase in the human uterus.

The endometrium displays characteristic cyclical changes involving proliferation and differentiation. The differentiation that takes place requires major tissue remodelling involving the matrix metalloproteinase (MMP) family as key enzymes in this process. Mast cells, containing the tryptase and chymase enzymes that are capable of stimulating the MMP cascade, have been identified in the endometrium, but their role is still unclear. In this study, we observed that the majority of mast cells in the uterus reside in the myometrium and that they co-express mast cell tryptase and MMP-1 in the same intracellular granules. In endometrium exposed to synthetic progestogen via an intrauterine levonorgestrel system a significant increase in mast cells numbers was observed in women experiencing breakthrough bleeding compared to those in women with no reported bleeding. We conclude that mast cells contain MMP-1 and we postulate a potential role for mast cells in breakthrough bleeding.

Matrix metalloproteinases and tissue inhibitors of metalloproteinases in human fetal testis and ovary.

Matrix metalloproteinases (MMP) and tissue inhibitors of metalloproteinases (TIMP) are major regulators of tissue remodelling of the extracellular matrix (ECM) and may also be involved in the control of growth factor availability. We have investigated their production and localization in the developing human gonad during mid-gestation using zymographic techniques and immunohistochemistry. The secretion of MMP-2, MMP-9 and all four TIMP was demonstrated from both testis and ovary, with the predominant gelatinase produced by both being MMP-2. In the testis, MMP-1, MMP-2, MMP-9 and all TIMP family members were localized to the interstitium and to varying degrees within the tubules. MMP-9 and TIMP-4 were abundant in both Sertoli cells and gonocytes and MMP-1 and TIMP-1 were localized in particular to Sertoli cells. In the ovary, all TIMP and MMP-1, MMP-2 and MMP-9 were localized to the oogonium/oocyte cytoplasm with varying intensities and MMP-1, TIMP-2 and TIMP-3 were also detected in the ovarian stroma. This study demonstrates that MMP-1, MMP-2, MMP-9 and all TIMP family members are secreted by the developing ovary and testis and are localized to specific cell and tissue sites. MMP and TIMP are likely to play a role in ECM remodelling during gonadal development and also in the cell and matrix interactions that control a range of cellular functions.

Secretion of matrix metalloproteinases 2 and 9 and tissue inhibitor of metalloproteinases into follicular fluid during follicle development in equine ovaries.

Extensive tissue remodelling is required in equine ovaries for follicle growth and development and also migration of the follicle to the ovulatory fossa, where ovulation occurs. The mechanisms for these processes are largely unexplored. Matrix metalloproteinases (MMPs) and their endogenous tissue inhibitors (TIMPs) are important for control of breakdown of extracellular matrix during tissue remodelling. The aims of this study were to determine the pattern and sites of secretion of the gelatinases MMP-2 and -9 and TIMPs into follicular fluid during follicle development in mare ovaries. The predominant gelatinase detected in follicular fluid was MMP-2, which was present in similar amounts throughout follicular development, as demonstrated by zymography. MMP-9 was also present in follicular fluid and secretion increased significantly (P < 0.05) with development of follicles from < 10 mm to 11-20 mm in diameter. Follicular fluid also contained TIMP-1, TIMP-2, unglycosylated and glycosylated TIMP-3, and TIMP-4, as shown by reverse zymography. The abundance of TIMPs remained largely unchanged during follicle development. MMP-2 and -9 were localized by immunohistochemistry to stromal cells and granulosa and theca cells. TIMP-1, -2, -3 and -4 were present in granulosa and theca cells of the follicle and in stromal cells and also associated with extracellular matrix of the ovarian stromal tissue. The MMPs and TIMPs are likely to be involved in the regulation of the breakdown of extracellular matrix during tissue remodelling for follicle development and migration to the ovulation fossa in mares.

Serum inhibin, activin and follistatin in postmenopausal women with epithelial ovarian carcinoma.

OBJECTIVE: To investigate the role of serum inhibin A, inhibin pro-alphaC immunoreactivity, activin A, and follistatin in postmenopausal women with epithelial ovarian cancer. DESIGN: Case-control study. SAMPLE: e Serum samples from 27 postmenopausal women with epithelial ovarian cancer and 54 controls from the general population participating in an ovarian cancer screening trial. RESULTS: Women with epithelial ovarian cancer had significantly higher serum levels of pro-alphaC immunoreactivity (P = 0.03), activin A (P = 0.004) and follistatin (P = 0.04), but not inhibin A (P = 0.13). Using the 90th centile in the control group as the cut off, pro-alphaC levels were elevated in 41% of women with epithelial ovarian cancer, while inhibin A was elevated in only 15%. Using the 95th centile as the cut off, serum pro-alphaC was elevated in only 11% of women with epithelial ovarian cancer (3/27), while activin A was elevated in 48% (11/23). Follicle stimulating hormone levels were significantly lower in women with epithelial ovarian cancer (P = 0.01). Although, inhibin-related peptides can modulate follicle stimulating hormone levels, there was no correlation between inhibin A, pro-alphaC immunoreactivity, activin A or follistatin and follicle stimulating hormone. CONCLUSION: These data demonstrate that though there is preferential secretion of precursor forms of the alpha subunit rather than dimeric inhibin A by epithelial ovarian cancer, pro-alphaC is unlikely to be a useful tumour marker. Activin A is more commonly elevated in postmenopausal women with epithelial ovarian cancer and its role as a tumour marker in the diagnosis and screening of epithelial ovarian cancer warrants further evaluation.

Culture of bovine preantral follicles in a serum-free system: markers for assessment of growth and development.

Satisfactory development of bovine follicles in vitro remains elusive. This study used a serum-free system to evaluate the effects of insulin-like growth factor-1 (IGF-1) on bovine preantral follicles in culture and to identify the activity of gelatinase matrix metalloproteinases (MMPs) and their endogenous inhibitors (TIMPs) in vitro to assess their potential as markers of development. Preantral follicles were cultured for 6 days in serum-free medium containing insulin and IGF-1 (10 ng/ml). No difference was observed in follicular growth, health, or antrum formation between IGF-1-treated follicles and controls. However, IGF-1 had a negative effect (P < 0.01) on oocyte size and granulosa cell proliferation. When MMP-9 was secreted, the probability of follicles having healthy granulosa or theca cells at the end of the culture period was 0.85 and 0.60, respectively. If TIMP-1 was released, the probability of follicles having healthy somatic cells was 0.79. When TIMP-2 was detected, the probability of granulosa and theca cell health was 0. 78 and 0.67, respectively. These results demonstrate no positive effects of IGF-1 on bovine follicles in this system. Furthermore, MMP-9 and TIMPs are related to follicular health and, therefore, can be used as markers of follicular development.

Increase in 15-hydroxyprostaglandin dehydrogenase activity in the ovine placentome at parturition and effect of oestrogen.

Type 1 NAD(+)-dependent 15-hydroxyprostaglandin dehydrogenase (PGDH) is the key enzyme for metabolism of active primary prostaglandins to inactive forms in gestational tissues. The present study examined the activity and immunolocalization of PGDH in the ovine placenta, fetal membranes and uterus over the latter half of pregnancy, and its potential regulation by oestradiol. Placenta, fetal membranes and myometrium were collected from sheep with known single insemination dates on days 70, 100 and 135 of gestation and in active labour demonstrated by electromyographic activity. In addition, chronically catheterized fetuses were infused with oestradiol (100 microgram kg(-1) per 24 h) (n = 5) or saline vehicle into the fetus from day 120 to day 125. PGDH activity measured in placental extracts remained constant from day 70 to day 135 of gestation, and then significantly (P < 0.05) increased by 300% in active labour. Immunoreactive PGDH was localized in the placentome at all stages and was present predominantly in the fetal component of the placentome in uninucleate, but not in binucleate, trophoblast cells. Similarly, in the fetal membranes PGDH immuno-reactivity was present in the uninucleate trophoblast but not in the binucleate cells of the chorion. PGDH immunostaining was also present in the endometrial luminal epithelium, in the smooth muscle of the myometrium, and the glandular epithelium of the cervix. Infusion of oestradiol into the fetal circulation from day 120 to day 125 of gestation had no effect on placental PGDH activity. Immunohistochemistry was used to localize oestrogen receptor alpha in intrauterine tissues to investigate further the failure of oestradiol to increase PGDH activity. Immunoreactive oestrogen receptor alpha was not present in the fetal component of the placenta, although it was expressed in adjacent maternal-derived cells. It is concluded that (1) PGDH activity increases in late gestation; (2) PGDH is expressed in uninucleate trophoblast cells in the ovine placenta and fetal membranes, and also in the maternal endometrial epithelium and stroma, myometrium and cervix; (3) oestrogen receptor alpha is not expressed in fetal cells in the placenta or fetal membranes; and (4) the increase in PGDH activity is not regulated by oestradiol administered to the fetus.

Dose-finding study of oral desogestrel with testosterone pellets for suppression of the pituitary-testicular axis in normal men.

Prototype hormonal male contraceptive regimens generally achieve only incomplete suppression to azoospermia with potentially adverse metabolic effects. We have carried out a short-term dose-finding study to investigate the potential of an oral gestogen, desogestrel, with testosterone pellets. Normal men received a single dose of 300 mg testosterone with 75 microg, 150 microg or 300 microg desogestrel daily for 8 weeks (n = 10 per group). LH and FSH were rapidly suppressed, with little difference between groups. Testosterone concentrations fell slightly during treatment with evidence of a linear dosage effect. Plasma inhibin B showed minor changes, but in seminal plasma it was suppressed, becoming undetectable in all men in the 300 microg desogestrel group. There were no significant changes in lipoproteins, fibrinogen or sexual behaviour during treatment, and minor falls in haematocrit and haemoglobin concentration. Sperm concentration fell in a dose-dependent manner, with three men, one man and seven men in the three groups respectively achieving severe oligozoospermia (<3 x 10(6)/ml), and three men achieving azoospermia in the 300 microg group despite the short duration of the study. The combination of oral desogestrel with depot testosterone thus results in profound suppression of gonadotrophin secretion without adverse metabolic or behavioural effects. Desogestrel with a long-acting testosterone preparation is a promising approach to hormonal male contraception.

Involvement of matrix metalloproteinases 2 and 9, tissue inhibitor of metalloproteinases and apoptosis in tissue remodelling in the sheep placenta.

Placental growth and development is crucial for successful pregnancy. The aim of this study was to characterize the activity and localization of the matrix metalloproteinase 2 (MMP-2) and MMP-9, which are capable of degrading basement membrane collagen (predominantly collagen type IV), and their endogenous tissue inhibitor of matrix metalloproteinases (TIMPs), in amniotic fluid and in the developing ovine placenta. Cell deletion by apoptosis during placental development was also examined. Zymography with gelatin as substrate indicated that MMP-2 (72 kDa gelatinase A; predominantly latent form) was present in increasing amounts in amniotic fluid from day 70 of gestation to labour (days 140-145), and MMP-9 (92 kDa gelatinase B; predominantly latent form) was detectable from day 125 to labour; there was no increase in MMP-2 or -9 in labour. A broad range of TIMPs was detected in amniotic fluid; the molecular masses corresponded to TIMP-1, -2 and -3. Immunohistochemical techniques localized MMP-2, MMP-9 and TIMP-3 in the sheep placenta, predominantly in the trophoblast layer in uninucleate, but not binucleate, cells. However, MMP-2 and -9 activated proteins in placental homogenates were low throughout pregnancy. Apoptosis was identified by morphological criteria and also by TdT-mediated dUTP nick end labelling. Apoptosis was present in discrete regions in the placenta, predominantly in trophoblast cells near the tips and the basal regions of the fetomaternal interdigitations. During pregnancy the sheep placenta becomes more complex and the area of the fetomaternal interface increases. MMP-2 and -9 are likely to be involved in breaking down basement membranes to allow cell migration during this process. It is suggested that digestion of supporting extracellular matrix may trigger apoptosis and in some way increase the branching pattern in the villi.