We investigated the effect of n-6 polyunsaturated fatty acids (PUFAs) on prostaglandin (PG) production by the uterus. A mixed population of endometrial cells (epthelium and stroma) from late-gestation ewes were cultured in defined medium containing linoleic acid (LA, 18:2, n-6), gamma-linolenic acid (GLA, 18:3, n-6) or arachidonic acid (AA, 20:4, n-6) in concentrations of 0 (control), 20 or 100 microM. After 45 h in test medium with or without added PUFAs, cells were challenged with control medium (CM), oxytocin (OT, 250 nM), lipopolysaccharide (LPS, 0.1 micro g/ml) or dexamethasone (DEX, 5 microM) for 22 h in the continued presence of the same concentration of PUFA and the medium was collected for measurement of PGF(2alpha) and PGE(2). Supplementation with LA inhibited the production of PGF(2alpha) but did not alter PGE(2), whereas GLA and AA increased production of both PGs. All PUFA supplements thus increased the ratio of PGE(2) to PGF(2alpha) (E:F ratio) two- to threefold. In control cells, OT and LPS challenges stimulated the production of PGF(2alpha) and PGE(2). In all challenge groups, the concentrations of PGF(2alpha) in response to PUFAs followed the same pattern - LA<control<;GLA<AA - but there were significant alterations in responsiveness as a result of PUFA treatment. In the cells supplemented with 100 microM AA, there was no further increase in PGF(2alpha) output in the presence of OT or LPS and when 100 microM GLA was present neither LPS nor OT stimulated PGE(2) significantly. When LPS was given to AA-supplemented cells, the E:F ratio was increased. DEX did not change PGE(2) production in control or LA-treated cells, but the cells produced significantly less PGF(2alpha), so the E:F ratio was increased. In contrast, in GLA- and AA-treated cells, DEX reduced the production of both PGF(2alpha) and PGE(2), so the E:F ratio was unaltered. In summary, the study showed altered production of PGs in the presence of different PUFAs according to their position in the n-6 metabolic pathway. The type of PUFA present affected responsiveness to OT, LPS and DEX and also changed the ratio of PGE(2) to PGF(2alpha) produced. The possible implications of this work are discussed in relation to the effect of diet on term and pre-term labour, which both require upregulation of the endometrial PG synthetic pathway.

The present study examined the extent to which the late gestation rise in fetal plasma cortisol influenced adipose tIssue development in the fetus. The effect of cortisol on the abundance of adipose tIssue mitochondrial proteins on both the inner (i.e. uncoupling protein (UCP)1) and outer (i.e. voltage-dependent anion channel (VDAC)) mitochondrial membrane, together with the long and short forms of the prolactin receptor (PRLR) protein and leptin mRNA was determined. Perirenal adipose tIssue was sampled from ovine fetuses to which (i) cortisol (2-3 mg/day for 5 days) or saline was infused up to 127-130 days of gestation, and (ii) adrenalectomised and intact controls at between 142 and 145 days of gestation (term=148 days). UCP1 protein abundance was significantly lower in adrenalectomised fetuses compared with age-matched controls, and UCP1 was increased by cortisol infusion and with gestational age. Adrenalectomy reduced the concentration of the long form of PRLR, although this effect was only significant for the highest molecular weight isoform. In contrast, neither the short form of PRLR, VDAC protein abundance or leptin mRNA expression was significantly affected by gestational age or cortisol status. Fetal plasma triiodothyronine concentrations were increased by cortisol and with gestational age, an affect abolished by adrenalectomy. When all treatment groups were combined, both plasma cortisol and triiodothyronine concentrations were positively correlated with UCP1 protein abundance. In conclusion, an intact adrenal is necessary for the late gestation rise in UCP1 protein abundance but cortisol does not appear to have a major stimulatory role in promoting leptin expression in fetal adipose tIssue. It remains to be established whether effects on UCP1 protein are directly regulated by cortisol alone or mediated by other anabolic fetal hormones such as triiodothyronine.

Polyunsaturated fatty acids derived from the diet are incorporated into cell membranes where they act as precursors for prostaglandin (PG) synthesis. Linoleic acid (LA; 18:2 n-6) is a major constituent of plant oils and its consumption in Westernized populations is increasing. This study investigated the influence of LA on PG production by the uterus and placenta. Pregnant ewes were fed a control or an LA-enriched diet. Oxytocin (OT) was injected on day 45 (early) or day 133 (late) of gestation to measure the release of 13,14-dihydro-15-keto PGF_2α (PGFM). Ewes were killed on day 46 or day 138 for collection of uterine intercaruncular endometrium and fetal allantochorion. Basal and stimulated PG release from explant cultures was assessed before and after in vitro treatment with OT, lipopolysaccharide (LPS), dexamethasone (DEX) or calcium ionophore (CaI). Expression of cyclooxygenase (COX)-1 and COX-2 was determined by Western blot in endometrium of late-gestation ewes. Circulating PGFM levels in vivo did not differ according to diet but there were highly significant differences in the release of PGs in vitro. Basal production of PGF_2αand PGE₂ by the endometrium and of PGE₂ by the allantochorion were all higher in tissues from LA-supplemented ewes. Endometrial tissues produced more PG following OT and CaI treatment, whereas DEX inhibited production of both PGs at both stages of gestation. In allantochorion collected at day 46 LPS did not significantly alter PGE₂ release and DEX increased output, whereas at day 138 LPS was stimulatory but DEX was inhibitory. These data show that a high-LA diet can significantly increase the ability of both endometrium and placental tissues to produce PGs in vitro. This effect of diet may only become apparent after a sustained period of PG release, so was not seen following the brief pulse caused by OT treatment in vivo. As COX protein levels were unaltered, the main influence was likely to be via conversion of LA to arachidonic acid, providing an increased supply of precursor. These results support previous studies which suggest that alterations in dietary polyunsaturated fatty acids may influence the time of labour.

Rather than being a constitutive enzyme as was first suggested, endothelial nitric oxide synthase (eNOS) is dynamically regulated at the transcriptional, posttranscriptional, and posttranslational levels. This review will focus on how changes in eNOS function are conferred by various posttranslational modifications. The latest knowledge regarding eNOS targeting to the plasma membrane will be discussed as the role of protein phosphorylation as a modulator of catalytic activity. Furthermore, new data are presented that provide novel insights into how disruption of the eNOS dimer prevents eNOS uncoupling and the production of superoxide under conditions of elevated oxidative stress and identifies a novel regulatory region we have termed the ‘flexible arm’.