Rodney W. Kelly

PROSTAGLANDIN SYNTHESIS IN THE ENDOMETRIUM OF WOMEN WITH OVULAR DYSFUNCTIONAL UTERINE BLEEDING

The endogenous concentrations of prostaglandins F2α (PGF2α) and E (PGE) were measured during the luteal phase of the menstrual cycle in the endometrium from 14 women with unexplained menorrhagia (measured menstrual blood loss in excess of 50 ml) and 15 women with normal menses (blood loss 50 ml or less). Although there was no significant difference in the PGF 2α/PGE ratio between the two groups, this ratio was significantly lower in the endometrium from eight of the women whose blood loss exceeded 90 ml (p <0.05). There was a significant inverse correlation between the PGF2α/PGE ratio and blood loss (r = 0.36, p <0.025). The synthetic capacity of the endometrium was assessed by incubation of the tissue with 14C arachidonic acid. Endometria from nine women with unexplained menorrhagia synthesized more PGE2 than PGF2α, whereas the converse was true with 11 control endometria. Consequently the PGF2α/PGE2 ratio was significantly reduced in the former group (p <0.025). Oestradiol‐17β (200 μM) and to a greater extent 2 hydroxy oestradiol (200 μM) increased the total prostaglandin synthesis by the endometria, but did not significantly alter the PGF2α/PGE2 ratio. These results suggest that excessive blood loss may be associated with a shift in the endometrial conversion of prostaglandin endoperoxide from PGF2α to PGE2.

Innate immune defences in the human endometrium

The human endometrium is an important site of innate immune defence, giving protection against uterine infection. Such protection is critical to successful implantation and pregnancy. Infection is a major cause of preterm birth and can also cause infertility and ectopic pregnancy. Natural anti-microbial peptides are key mediators of the innate immune system. These peptides, between them, have anti-bacterial, anti-fungal and anti-viral activity and are expressed at epithelial surfaces throughout the female genital tract. Two families of natural anti-microbials, the defensins and the whey acidic protein (WAP) motif proteins, appear to be prominent in endometrium. The human endometrial epithelium expresses beta-defensins 1–4 and the WAP motif protein, secretory leukocyte protease inhibitor. Each beta-defensin has a different expression profile in relation to the stage of the menstrual cycle, providing potential protection throughout the cycle. Secretory leukocyte protease inhibitor is expressed during the secretory phase of the cycle and has a range of possible roles including anti-protease and anti-microbial activity as well as having effects on epithelial cell growth. The leukocyte populations in the endometrium are also a source of anti-microbial production. Neutrophils are a particularly rich source of alpha-defensins, lactoferrin, lysozyme and the WAP motif protein, elafin. The presence of neutrophils during menstruation will enhance anti-microbial protection at a time when the epithelial barrier is disrupted. Several other anti-microbials including the natural killer cell product, granulysin, are likely to have a role in endometrium. The sequential production of natural anti-microbial peptides by the endometrium throughout the menstrual cycle and at other sites in the female genital tract will offer protection from many pathogens, including those that are sexually transmitted.

The Pseudomonas Autoinducer N-(3-Oxododecanoyl) Homoserine Lactone Induces Cyclooxygenase-2 and Prostaglandin E2 Production in Human Lung Fibroblasts: Implications for Inflammation

Pseudomonas aeruginosa causes lethal lung infections in immunocompromised individuals such as those with cystic fibrosis. The lethality of these infections is directly associated with inflammation and lung tissue destruction. P. aeruginosa produces several acylated homoserine lactones (AHL) that are important in the regulation of bacterial virulence factors. Little is known about the effects of AHLs on human cells. In this work we report that the AHL N-(3-oxododecanoyl) homoserine lactone (3O-C12-HSL) from P. aeruginosa induces cyclooxygenase (Cox)-2, a seminal proinflammatory enzyme. When primary normal human lung fibroblasts were exposed to 3O-C12-HSL, an 8-fold induction in mRNA and a 35-fold increase in protein for Cox-2 were observed. In contrast, there was no substantial change in the expression of Cox-1. We also demonstrated that the induction of Cox-2 was regulated by 3O-C12-HSL activation of the transcription factor NF-κB. 3O-C12-HSL also stimulated an increase in the newly discovered inducible membrane-associated PGE synthase but had no effect on the expression of the cytosolic PGE synthase. We also demonstrate that 3O-C12-HSL stimulated the production of PGE2. PGE2 is known to induce mucus secretion, vasodilation, and edema, and acts as an immunomodulatory lipid mediator. We propose that 3O-C12-HSL induction of Cox-2, membrane-associated PGE synthase, and PGE2 likely contributes to the inflammation and lung pathology induced by P. aeruginosa infections in the lung. These studies further reinforce the concept that bacterial AHLs not only regulate bacterial virulence but also stimulate the activities of eukaryotic cells important for inflammation and immune defenses.

Decidualization of the human endometrial stromal cell: an enigmatic transformation.

Changes in human endometrium are essential to allow the establishment of pregnancy. These changes are induced in vivo by progesterone, and include appearance within the tissue of a specific uterine natural killer cell, characterized by an abundant expression of CD56. Changes also occur in the stromal cells, which undergo a characteristic decidualization reaction. Decidualized stromal cells are derived from the fibroblast-like cells within the endometrium, which maintain their progesterone receptors in the presence of progesterone. Prolonged exposure to progesterone induces a rounded cell characterized by release of prolactin and insulin-like growth factor binding protein-1 (IGFBP-1), and expression of tissue factor. Additional changes include the secretion of interleukin (IL)-15, vascular endothelial growth factor, and surface expression of zinc dependent metalloproteinases such as CD10 and CD13. In vitro, elevated intracellular cAMP as well as progesterone is necessary for decidualization. In vivo, these conditions may be provided by progesterone from the corpus luteum, by prostaglandin E, a stimulator of adenyl cyclase, and relaxin, which has recently been shown to be a phosphodiesterase inhibitor. Given the co-distribution of uterine natural killer cells and decidualized stromal cells, a mutual interaction might provide the correct regulatory environment for successful implantation, and penetration of the maternal blood vessels by trophoblastic cells.

The endocrinology of menstruation – a role for the immune system

The human endometrium displays characteristic features, both structural and functional, across the menstrual cycle. It is the sex steroid hormones, oestrogen and progesterone, that drive the endometrium through the different phases of the cycle. Oestrogen and progesterone act sequentially to regulate cellular concentrations of their respective receptors, this interaction initiates gene transcription. Thereafter a cascade of local events prepares the endometrium for implantation, but in the absence of pregnancy, progesterone withdrawal leads to menstruation and cyclic repair. Withdrawal of progesterone from an oestrogen-progesterone primed endometrium is the initiating event for the cascade of molecular and cellular interactions that result in menstruation. Progesterone withdrawal first affects cells with progesterone receptors. Early events in the menstrual process are vasoconstriction and cytokine up-regulation. The activation of lytic mechanisms is a later event and involves cells that may lack progesterone receptors, for example, uterine leucocytes and epithelial cells. Hence progesterone withdrawal results in a local increase of inflammatory mediators and the enzymes responsible for tissue breakdown. The total complex of local factors implicated in normal menstrual and aberrant menstrual bleeding are yet to be fully defined.

Inflammatory mediators and cervical ripening

The mechanisms which soften the cervix and allow it to dilate at birth are not well known. This is a crucial element in labour and current pharmacological approaches, largely the use of prostaglandins (PG), are only semi-selective for the cervix and can cause inappropriate myometrial contractions. Cervical ripening is accompanied by the influx of neutrophils, the neutrophil is a ready source of collagenase, and the cervix is dependent on collagen for its rigidity. Thus it is important to study factors controlling neutrophil influx into the cervix at term. PGE and interleukin-8 (IL-8, or neutrophil chemotactic factor) work synergistically in inducing neutrophil influx into tissue. Activating this type of synergy, between a vasoactive and a chemotactic agent is likely to be the physiological mechanism for inducing cervical ripening. Future approaches to control the cervix are likely to exploit these pathways and lead to more effective and acceptable methods for inducing labour.

Choriodecidual production of interleukin-8 and mechanism of parturition

Both prostaglandins and antiprogestagens can induce labour and ripen the cervix, but the mechanisms are unclear. The collagenases that bring about cervical ripening are neutrophil derived. We examined the potential of uterine tissues to control neutrophil attraction by measuring interleukin-8 production. Choriodecidual cells in culture produced substantial amounts of interleukin-8; release was inhibited by progesterone and stimulated by the antiprogestagen mifepristone. Interleukin-8 production was similar in cells from spontaneously delivered placentas and from those obtained at caesarean section. Since prostaglandin E and interleukin-8 have synergistic effects, we suggest that interleukin-8 activity is the final common step of prostaglandin and antiprogestagen action in parturition.

Differential production of prostaglandins within the human uterus

The ability of broken cell preparations of human endometrium, myometrium and a mixture of endometrium and myometrium to convert 14C arachidonic acid to prostaglandins (PGs) was compared. Endometrium metabolished arachidonic acid predominantly to a mixture of PGF2 alpha and PGE2. A similar weight of myometrium showed relatively little activity, the major product identified was 6 oxo PGF1 alpha. However, a combination of endometrium and myometrium showed an enhanced conversion of arachidonic acid to 6 oxo PGF1 alpha associated with a decreased production of PGF2 alpha and PGE2. This suggests that human endometrium and myometrium differ in their ability to metabolize arachidonic acid and in their ability to convert the endoperoxides formed, to PGs.

Interleukin-8 production by the human cervix

OBJECTIVES Our purpose was (1) to determine whether the human cervix is capable of producing interleukin-8 in vitro and to examine the possibility of stimulating an increase in any such output and (2) to examine the concomitant production of prostaglandins. STUDY DESIGN Cervical tissue was obtained from 48 women, 29 pregnant women undergoing surgical termination of pregnancy (20 of whom were treated with the prostaglandin analog Cervagem), 14 nonpregnant, premenopausal women, and three postmenopausal women. Explants were cultured and the medium was assayed for interleukin-8 and prostaglandin E2. Analysis of variance and Newman-Keuls statistical tests were used. RESULTS Significant quantities of interleukin-8 were produced by the tissue, and the data indicate that cervical explants from pregnant and nonpregnant women behave in a similar way when challenged by phorbol myristate acetate but that the postmenopausal cervix loses its capacity for interleukin-8 production. CONCLUSIONS Human cervix is capable of producing large amounts of interleukin-8 in vitro, and it may be influenced by the steroid hormones. Thus interleukin-8 could be an excellent candidate for a prime role in neutrophil-mediated cervical ripening.

Extracellular organelles (prostasomes) are immunosuppressive components of human semen.

Numerous reports have ascribed immunosuppressive activity to human seminal plasma and there is growing agreement that much of this activity can be accounted for by the very high levels of E series prostaglandins present (up to 300 μm 19‐hydroxy prostaglandin E). However not all suppressive activity is due to prostaglandin since several reports have appeared of high molecular weight active substances and we have found that stripped seminal plasma is still effective in inhibiting the mitogen‐induced proliferation of lymphocytes. In this study such immunosuppressive activity has been separated by molecular size fractionation and the activity has been found to be particulate and corresponded to the previously reported prostasomes. These are trilaminar to multilaminar vesicles (150 nm diameter) which are secreted by the prostate. Pure preparations of prostasomes inhibited mitogen‐induced lymphoproliferation in a dose‐dependent manner with a concentration of prostasomes equivalent to 40% of that seen in seminal fluid giving 69% suppression of thymidine incorporation. The suppressive activity survived boiling and therefore was unlikely to be due to enzymatic action associated with these organdies. Interaction with the accessory cells, involved in full development of the lymphoproliferation induced by mitogen, was indicated and this possibility was supported by the demonstration of a direct effect of prostasomes on macrophage function using a mouse macrophage cell line. The prostasomes in semen may play a complementary role to the prostaglandins in neutralizing the immune defences of the female reproductive tract. This combination would allow the alloantigenic spermatozoa the best chance of achieving fertilization, but at the same time leave the recipient open to any infection present in the semen.