Integrins and adhesion mlecules: Cell adhesion molecules on the oocyte and preimplantation human embryo

The presence of cell adhesion molecules on human oocytes, earlyembryos, and pre-hatched blastocysts was examined by indirectimmunofluorescence and compared to the distribution found onfirst trimester villous placenta with the same antibodies. Sixintegrin subunits (3, V, 1, 3, 4, 5) were observed consistentlythroughout preimplantation development. Evidence was also obtainedfor the presence of integrin subunits 2, 4, L, 2, and 7 on asmall number of oocytes. A more restricted developmental analysisof E-cadherin, ICAM-1, NCAM, and VCAM-1 demonstrated that thesecell adhesion molecules are also present on oocytes and earlyembryos. L-selectin was detected on oocytes but was not foundon 8-cell embryos. The oocyte and early blastomeres have complexsurfaces in which the integrin and CAM families are represented.     

Interleukin-1{alpha} and -{beta} modulation of luteinized human granulosa cell oestrogen and progesterone biosynthesis

This study was designed to test the hypothesis that inter-leukin-1(IL-1) and directly affect progesterone, and oestradiol productionin cultures of purified human granulosa cells. Luteinized granulosacells were obtained from women during in-vitro fertilizationcycles. Granulosa cells with and without associated white bloodcells were cultured in the presence of IL-1 and IL-1 (0.5–50ng/ml) for 48 h. Media were changed at 24 h intervals and assayedfor progesterone and oestradiol. In separate experiments, granulosacell viability was assessed with the tetrazolium salt reductionassay, haemocytometer cell counts, and Trypan blue dye exclusion.Our results indicate that progesterone synthesis by basal andhuman chorionic gonadotrophin (HCG)-stimulated granulosa cellsco-cultured with white blood cells was inhibited by 5.0 ng/mlof IL-1 and IL-1 at 48 h of culture. In the presence of whiteblood cells, granulosa cell oestradiol synthesis was inhibitedby IL-1 but not IL-1. Oestradiol was inhibited after both 24and 48 h of culture and was maximally affected by 5.0 ng/mlof IL-1. In contrast, basal and HCG-stimulated oestradiol productionby granulosa cells cultured free of white blood cells was inhibitedonly by IL-1. IL-1 at 5.0 ng/ml produced maximal inhibitionof basal oestradiol (57%) and HCG-stimulated oestradiol (41%)production at 48 h of culture. Gonadal steroid inhibition byIL-1 and IL-1 was not mediated through cytotoxic or antiproliferativeeffects on granulosa cells. Specificity of the granulosa cellresponse to IL-1 and IL-1 was demonstrated by abrogation ofsteroid inhibition with anti-IL-1 and IL-1 neutralizing antibodies.In conclusion, IL-1 directly inhibited the production of oestradiolby human ovarian granulosa cells. IL-1 and IL-1 also exertedindirect effects on steroid production via white blood cellsthat are usually present in granulosa cell cultures if stepsare not taken to remove them. These data support the hypothesisthat cytokines play an important role in intra-ovarian regulationof steroid biosynthesis.     

Integrins are not involved in the process of human sperm-oolemmal fusion

BACKGROUND: We investigated whether integrins are required forthe human sperm–oocyte binding and fusion processes. METHODS:The expression of several integrin subunits at the human oocyteplasma membrane was investigated using immunofluorescence microscopy,and the functional role of integrins expressed at the humanoocyte surface in sperm–oocyte interaction was studiedusing a zona-free human oocyte binding and fusion assay. A totalof 144 unfertilized oocytes were stained with anti-integrinantibodies and 147 zona-free unfertilized oocytes were inseminatedin the presence of various anti-integrin antibodies that wereexpressed in oocyte plasma membrane. RESULTS: The antibodiesof six integrin subunits (₂, ₃, ₅, ₆, _V, _M) and six integrinsubunits (₁, ₂, ₃, ₄, ₅, ₆) were bound to the surface of fixedunfertilized oocytes. In contrast, the presence of ₁ and ₄ subunitscould not be verified. The human sperm–oocyte bindingwas only partially inhibited by blocking antibodies of ₂, ₃,₅, ₆, _V, _M, ₁, ₂ and ₃ with a maximum of 55% inhibition, butantibodies of ₄, ₅ and ₆ showed no effect on sperm–oolemmalbinding. A similar reduction of the number of fused sperm wasobserved. However, the ratio of fused sperm to total sperm (boundand fused) was not impaired by all integrin antibodies, suggestingthat integrins had no role in the sperm–oolemmal fusionprocess. CONCLUSIONS: These results suggest that one of thebinding mechanisms can be inhibited by integrin antibodies butthat this mechanism does not play an essential role in the humansperm–oolemmal binding and fusion processes. The othermechanisms, insensitive to integrins, may involve both bindingand fusion processes in human oocytes.     

Ovary: Immunolocalization of transforming growth factor-{beta}1 and transforming growth factor-{beta}2 in the mouse ovary during gonadotrophin-induced follicular maturation

An immunohistochemical approach was utilized to evaluate thecellular distribution of transforming growth factor-₁ (TGF₁)and transforming growth factor ₂ (TGF₂) at different stagesof follicle development in the prepubertal mouse ovary underthe following conditions: (i) after pregnant mare's serum gonadotrophin(PMSG) treatment; (ii) after PMSG and human chorionic gonadotrophin(HCG) treatment; (iii) after PMSG and HCG treatment plus mating.In the immature ovary, TGFF₁ and TGF₂ immunoreactivities arelocalized in theca and granulosa cells and in oocytes. AfterPMSG treatment, TGF₁ and TGF₂ immunoreactivities are localizedin granulosa cells; in addition, TGF₂ staining is noted in thematrix surrounding antral cells. Staining for both TGF₁ andTGF₂ drops in the theca but persists in the oocyte. PMSG plusHCG treatment results in a significant increase in TGF₁ andTGF₂ immunoreactivity in the theca and in the maintenance ofTGF₁ staining in both basal granulosa cells and cumulus cellswhereas TGF₂ immunoreactivity is essentially localized in thematrix surrounding cumulus cells. Staining for TGF₁ and TGF₂persists in the oocyte. Following PMSG plus HCG treatment andmating, TGF₁ immunoreactivity is localized in the luteal cellsof corpora lutea and TGF₂ shows a similar localization pattern.This study provides evidence that TGF₁ and TGF₂ peptides areexpressed in specific cell types during induced follicular maturationin the mouse ovary.     

Molecular aspects of implantation: Human trophoblast adhesion to matrix proteins: inhibition and signal transduction

At the time of implantation, the extracellular matrix proteinslaminin and fibronectin are abundant in the decidua and aredistributed pericellularly around each individual stromal cell.First trimester human trophoblast expresses both laminin andfibronectin receptors, specifically the ₁₁, ₅₁, ₆₁ and ₆₄ integrinheterodimers. In this study we have demonstrated that in-vitroadhesion of first trimester human trophoblast to purified extracellularmatrix proteins and to purified decidual stromal cell monolayerscan be inhibited by monoclonal antibodies directed against appropriateintegrin subunits and by synthetic peptides containing an arginine-glycine-asparticacid sequence. Monoclonal antibodies (mAbs) to the ₅ and ₁ integrinsubunits and a synthetic peptide significantly inhibited adhesionto fibronectin. Binding of trophoblast to laminin was blockedwith mAbs to the ₆ and ₁ but not ₁ and ₄ integrin subunits.Similarly, integrin-mediated adhesion to monolayers of decidualstromal cells could be blocked with mAbs to the ₅, ₆, ₁ and₄ integrin subunits. Integrin-mediated signal transduction innormal and malignant trophoblast was investigated by Westernblotting. A 115 kDa protein was the major tyrosine phosphorylatedprotein detected in trophoblast after binding to laminin orfibronectin. The profile of tyrosine phosphorylated proteinsdiffered for malignant trophoblast.     

Endocrinology: Substrate dependency of C19 conjugates in hirsute hyperandrogenic women and the influence of adrenal androgen

Serum C₁₉ conjugates, specifically 3-androstanediol glucuronide(3G), reflect peripheral androgen action through the actionof 5-reductase activity. The origin of 5-reduced C₁₉ conjugateshas been controversial and it has been suggested that they arederived primarily from adrenal androgens. We examined concentrationsof 3G, 3-androstanediol sulphate (3S), androsterone glucuronide(AoG) and androsterone sulphate (AoS) in 40 hirsute hyperandrogenicwomen. These patients were divided into four groups based uponindividual, combined or normal concentrations of the adrenalandrogens dehydroepiandrosterone (DHEAS) and 11-hydroxy-androstenedione.Testosterone, unbound testosterone and androstenedione weresimilar in these groups. Serum 3G was equally high in all groupsand was correlated significantly with hirsutism, while the otherconjugates were not. Androsterone glucuronide was raised inall groups but was higher in patients with raised DHEAS. Serum3S was raised in all groups and was higher where both adrenalandrogens were raised. Serum AoS was highly correlated withDHEAS. Serum 3G was correlated with unbound testosterone andandrostenedione but not with the adrenal androgens. The glucuronideconjugates were correlated with one another as were the sulphateconjugates but glucuronides and sulphates were not correlated.These data confirm ovarian and adrenal dependency of C₁₉ conjugates.Serum 3G appears to reflect hirsutism most accurately and isleast dependent on adrenal androgens in patients with mixedhyperandrogenism.     

Concentration of interleukin-1{beta} correlates with prostaglandin E2 and F2{alpha} in human pre-ovulatory follicular fluid

To investigate the role(s) of interleukin-1 (IL-1) in humanovarian function, we measured the concentrations of IL–1,prostaglandins (PGs) and steroids in follicular fluid of 90stimulated ovaries, with reference to oocyte maturation. Concentrationsof IL-1 were significantly higher in the follicles from whichmature oocytes were recovered than in follicles from which oocytescould not be recovered (P < 0.05). IL-1 concentrations alsoincreased in association with oocyte maturation. Positive significantcorrelations were seen between IL-1 and prostaglandin E2 (PGE2)(r = 0.47, P < 0.001), and between IL-1 and prostaglandinF₂ (PGF₂) (r = 0.22, P < 0.05) in pre-ovulatory follicularfluid, but not between IL–1 and oestradiol, or betweenIL-1 and oestradiol, or between IL-1 and progesterone. 0Follicularfluid IL-1 might contribute to prostaglandin-induced oocytematuration and ovulation.     

Embryonic resistance to tumour necrosis factor-{alpha} mediated cytotoxicity: novel mechanism underlying maternal immunological tolerance to the fetal allograft

The cytokine tumour necrosis factor- (TNF) has been postulatedto play an essential role in the cytotoxic activity of cell-mediatedimmunity against allogenic or tumour cells invading the host.Several tumour cell lines, however, are resistant to TNF mediatedcytotoxicity and respond paradoxically by cellular proliferationand by autocrine secretion of TNF. In view of the metastaticcharacter of the mammalian embryo, the aim of this study wasto assess the potential of murine embryos to secrete TNF invitro, to express TNF receptors and to resist TNF mediated cytotoxicityduring their in-vitro development to the blastocyst stage. Thepotential of human embryos to secrete TNF in vitro until theblastocyst stage was also investigated. From a total of 11 humanembryos, which were allowed to proceed to blastocyst formation,seven secreted TNF in the range of 2–117 pg/ml/24 h. Atotal of 123 C57BL/6J mouse embryos were studied of which 55%secreted TNF in the range of 1.25–3.95 mg/ml/24 h. Thepresence of high levels of exogenous TNF (10–300 IU) wasnot detrimental to the in-vitro development of murine embryos.Using immunohistochemical techniques, we were not able to detectthe presence of type I or II TNF receptors on the surface ofmurine embryos. Our findings suggest that human and C57BL/6Jmurine embryos have the potential to secrete TNF in vitro duringthe developmental stages leading to blastocyst formation. Inboth species, the presence of TNF in the culture medium didnot cause subsequent necrosis of the conceptus, suggesting thatmammalian embryos may be TNF resistant cell lines. The observedembryonic resistance to TNF may be explained by the absenceof TNF receptors by which the cytotoxic effect is usually mediated.It is suggested that embryonic resistance to physiological concentrationsof TNF released by effectors of the host's immune system, couldbe via a mechanism underlying the maternal immunological toleranceto the fetal allograft.     

Immunology: {beta}2-Glycoprotein I-dependent and -independent anticardiolipin antibodies in healthy pregnant women

The purpose of this study was to determine the association between₂-glycoprotein I (₂GPI)-dependent anticardiolipin antibodies(aCL) and ₂GPI-independent aCL and their respective relevanceto adverse pregnancy outcomes. Therefore, we prospectively studied210 normal pregnant women, utilizing a modified enzyme-linkedimmunosorbent assay method for ₂GPI-dependent and -independentaCL. Seven of the 210 pregnant women (3.3%) demonstrated evidencefor ₂GPI-independent immunoglobulin G (IgG)-aCL. Two patients,who also appeared positive for ₂GPI-dependent IgG-aCL, wereproven to be false positives. Amongst the 210 patients, notone was thus positive for ₂GPI-dependent aCL. Women with ₂GPI-independentaCL demonstrated no adverse pregnancy outcomes. These resultssuggest that the presence of ₂GPI-independent aCL is not associatedwith the presence of 2GPI-dependent aCL, though it may giverise to false positive results. Since the presence of 2GPI-independentaCL does not appear to be associated with adverse pregnancyoutcomes, ₂GPI-dependent assays may represent better markersof miscarriage risk.     

Implantation: Tumour necrosis factor {alpha} inhibits in-vitro decidualization of human endometrial stromal cells

Interleukin-1 (IL-1) has been reported previously to inhibitthe in-vitro decidualization of human endometrial stromal cellsas assessed by progesterone-induced prolactin production andmorphological transformation. In this study we examined whetherother cytokines, such as tumour necrosis factor-(TNF), interferon-(IFN), IFN or granulocyte-macrophage colony-stimulating factor(GM-CSF), could affect the decidualization of human endometrialstromal cells in vitro. Of these cytokines, TNF significantlysuppressed prolactin production in a dose-dependent manner,with no apparent effect on cell number. The morphological transformationof endometrial stromal cells was also inhibited by TNF. TNFand IL-1 significantly suppressed cAMP-stimulated prolactinproduction by endometrial stromal cells. Neither the progesteroneconcentration in the supernatant of the endometrial stromalcell culture system nor intracellular calcium concentrationof the endometrial stromal cells were affected by the additionof TNF or IL-1. These results indicated that TNF and IL-1 suppressboth progesterone-induced and cAMP-mediated prolactin productionin endometrial stromal cells, and that this inhibition was notattributable to direct effects on progesterone metabolism orrelated to Ca2+-mediated signal transduction. These experimentssuggested that a local increase of TNF and IL-1 under certainpathological conditions in vivo may disturb blastocyst implantationand/or the maintenance of pregnancy by inhibiting the decidualizationof endometrial stromal cells.     

Immunology: Autoimmunity to spermatozoa, asymptomatic Chlamydia trachomatis genital tract infection and {gamma}{delta} T lymphocytes in seminal fluid from the male partners of couples with unexplained infertility

The relationship between an undetected, asymptomatic Chlamydiatrachomatis genital tract infection, the concentration of andb T cells in semen and sperm autoimmunity was examined in 48male partners of couples with unexplained infertility. ImmunoglobulinA (IgA) antibodies to C.trachomatis were detected in seminalfluids from 14 (29.2%) of the men. Only four of these were positivefor circulating anti-chlamydial IgA, suggesting that the stimulusfor antibody production was within the genital tract. In contrast,four men were positive for anti-chlamydial IgG in their semen;all were also seropositive for anti-chlamydial IgG. T lymphocytesbearing the and antigen receptors were present in every semensample. Men with seminal anti-chlamydial IgA, however, had significantly(P = 0.035) elevated semen T cell concentrations (median 3100cells/ml) than did men lacking this antibody (median 1400 cells/ml);concentrations of T cells were comparable in both groups. Genitaltract sperm autoimmunity, as shown by antibodies bound to motileejaculated spermatozoa, was detected in 13 (27.1%) men. Thepresence of these antibodies was associated with elevated concentrationsof both (median 4200 versus 700 cells/ml) and (median 5000versus 850 cells/ml) T cells (P = 0.0002 and 0.0001 respectively).Men with antisperm antibodies only in their serum had seminalT cell concentrations comparable with men testing negative forantisperm antibodies. Anti-chlamydial IgA was identified insemen from four of 10 men with IgA bound to their spermatozoaand in none of the men with only spermatozoabound IgG. Therewas no relationship between sperm quality and the occurrenceof seminal IgA antibodies to either C.trachomatis or spermatozoa.An asymptomatic C.trachomatis infection activates T cells withinthe male genital tract, which may lead to antisperm antibodyformation and immune-mediated infertility.     

Alterations in mitochondrial membrane potential during preimplantation stages of mouse and human embryo development

Mitochondria are cellular organelles regulating metabolism andcell death pathways. This study examined changes in mitochondrialmembrane potential (m) throughout the stages of preimplantationdevelopment in mouse embryos conceived either in vivo or invitro and human embryos donated to research from IVF. Embryosstained with the m-sensitive dye (JC-1) were quantified forthe ratio of high- to low-polarized mitochondria using a deconvolutionmicroscope. Overall, mouse zygotes and early embryos containa subset of high-polarized mitochondria with a progressive increasein the ratio of m observed with increasing cleavage. A transientincrease in the ratio of high to low m was observed in in vivofertilized 2-cell stage embryos, coincident with embryonic genomeactivation in the mouse, but not in 2-cell embryos obtainedthrough IVF. We further observed that arrested mouse 2-cellembryos possessed an increased ratio of m compared with non-arrestedembryos. In human 8-cell embryos we observed an increased ratioof high- to low-polarized mitochondria with increasing degreesof embryo fragmentation. We concluded that the pattern of mitochondrialmembrane potential progressively changes throughout preimplantationdevelopment, and that an aberrant shift in m could contributeto, or is associated with, decreased developmental potential.     

Integrins and adhesion molecules: Tumour necrosis factor-{alpha}-mediated dyscohesion of epithelial cells is associated with disordered expression of cadherin/{beta}-catenin and disassembly of actin filaments

Tumour necrosis factor (TNF)- induced, in a time- and dose-dependentfashion, dyscohesion (cell-cell dissociation) of the endometrialepithelial cells. TNF- impaired the ability of cells to aggregateand to attain compaction. The cell-cell adherent junction isa specialized region of the plasma membrane where cadherin moleculesact as adhesion molecules and actin filaments are densely associatedwith the plasma membrane through a well-developed plasmalemmalundercoat. Dyscohesion induced by TNF- was associated with thedisordered expression of cadherin\-catenin at the sites of cell-cellcontact. In addition, within the time-frame that dyscohesionwas induced, TNF- down-regulated the expression of actin mRNAonly at 100 ng/ml without modulating the overall amount of actinprotein, its -isoform or the amount of ribosylated actin. However,TNF--mediated dyscohesion of epithelial cells was associatedwith loss of plasmalemmal undercoat as well as intracytoplasmicaggregates of F-actin and a simultaneous increase in G-actin.The effect of cytochalasin-B, which disrupts actin filamentson cell-cell binding, was less pronounced than the effect ofTNF-, suggesting that the effect of this cytokine on dyscohesionis not solely dependent on the disassembly of actin filaments.These findings show that the induction of disordered expressionof adhesion molecules, as well as disassembly of actin filaments,are implicated in the dyscohesion induced by TNF-.     

Integrins and adhesion molecules: A positive correlation between expression of {beta}1-integrin cell adhesion molecules and fertilizing ability of human spermatozoa in vitro

The purpose of this study was to investigate firstly whether(1-integrin cell adhesion molecules are expressed by human spermatozoa,and secondly whether there is any relationship between the expressionof 1-integrin cell adhesion molecules and the fertilizing abilityof human spermatozoa in vitro. A total of 50 semen samples wereexamined. The samples were obtained from the male partners ofcouples undergoing in-vitro fertilization (IVF) for either unexplained,tubal or male factor infertility. A panel of six monoclonalantibodies against 1-integrin cell adhesion molecules and immunohistochemicaltechniques were used to identify the presence of these moleculeson the spermatozoa. The percentage of spermatozoa showing strongimmunolabelling with each monoclonal antibody was assessed ineach sample. The relationship between these results and theaetiology of infertility and incidence of fertilization wasexamined. 1-Integrins, and primarily the ones with 4-, 5- and6-chains, were expressed by human spermatozoa. Compared withsemen samples from unexplained or male factor infertility patients,samples from tubal infertility patients had a significantlyhigher (P < 0.05) percentage of spermatozoa expressing adhesionmolecules. There was a positive correlation between the expressionof 4, 5 and a6 adhesion molecules and the fertilizing abilityof spermatozoa. The positive correlation between the presenceof certain (1-integrin cell adhesion molecules and the fertilizingability of human spermatozoa suggests that integrins may beputative determinants in egg-sperm recognition and interaction.     

Pregnancy: Cervical ripening with the cytokines interleukin 8, interleukin 1{beta} and tumour necrosis factor {alpha} in guinea-pigs

It has been suggested that the collagenolytic enzymes releasedfrom white blood cells which infiltrate the pregnant human uterinecervix at term are responsible for connective tissue changeswhich take place during the ripening process. Similarly, aninfiltration of inflammatory cells occurs in pregnant guinea-pigseither spontaneously at term or at preterm after treatment withthe antiprogestin onapristone. The objective of this study wasto evaluate the effects of the inflammatory cytokines interleukin8 (IL-8), interleukin 1 (IL-1), tumour necrosis factor (TNF-)and a combination of IL-1 and TNF- on cervical ripening in guinea-pigsduring advanced pregnancy. The cytokines were applied locally(intracervically) in a gel for 2 days and the effects were assessedon the third day by both extensibility measurements and morphologicalevaluation. IL-8 treatment on days 42 and 43 post coitum (p.c)and on days 48 and 49 p.c. (term: day 67± 3 p.c.) significantly(P < 0.05) increased cervical extensibility at both stagesof pregnancy. Although IL-1 treatment (days 42 and 43 p.c.)led to a slight increase in cervical extensibility, this effectwas not statistically significant. An electron microscope studyperformed on days 48 and 49 p.c. revealed a pronounced cervicalripening accompanied by the dissolution of collagen fibres,stromal oedema and the infiltration of polymorphonuclear leukocytesin all cytokine-treated groups. The morphological effects ofIL-8 and IL-1 were indistinguishable from those observed duringnormal cervical ripening at term. In contrast, TNF-, both aloneand in combination with IL-1, brought about a severe inflammatoryreaction with a massive infiltration of lymphocytes, marcophagesand polymorphonuclear leukocytes at the investigated dose. Weconclude that the local application of the inflammatory cytokinesIL-8, IL-1 and TNF- produces cervical ripening without inducinglabour in pregnant guinea-pigs; the morphological effects ofIL-8 and IL-1 being similar to the physiological cervical ripening.Our data support the view that cytokines, particularly IL-8,may play an important role during physiological, pathologicaland induced cervical ripening and could be clinically usefulas an adjunct to labour and delivery.     

The production of tumour necrosis factor a (TNF-{alpha}) by human endometrial cells in culture

The production of tumour necrosis factor a (TNF-) by culturedhuman endometrial epithelial and stromal cells prepared fromendometrium obtained at different stages in the menstrual cyclehas been investigated. TNF- was not detectable in the supernatantsof stromal cell cultures prepared from endometrial tissue obtainedat any time in the menstrual cycle. TNF- production by endometrialepithelial cells in culture varied depending on the time inthe cycle at which the endometrial tissue was taken. Cells preparedfrom tissue obtained during the late proliferative phase ofthe cycle produced more TNF- than those prepared from tissueobtained at other times in the cycle. In addition, a small increasein TNF- production was seen by cells prepared from tissue obtainedduring the mid-secretory phase of the cycle. Interieukin 1 (IL-1)(1.4–140 pmol/1) caused a dose-dependent increase in TNF-production by cells prepared from both proliferative and secretoryendometrium. Maximum LL-1-stimulated increases in TNF- productionwere similar in cells from both proUferative and secretory endometriumand typically reached from four to 10 times basal values. Highdoses of progesterone, either alone or in the presence of oestradiol,also affected TNF-a production by epithelial cells. TNF- productionby cells prepared from proliferative endometrium was increasedby progesterone. In contrast, TNF- production by cells preparedfrom secretory endometrium was decreased in the presence ofprogesterone. The effects of steroids on TNF- production wereless marked than that of IL-1, with values increasing or decreasingto a maximum of three times the basal value. Placental protein14 (PP14) (0.18 and 1.8 nmol/1) also increased TNF- productionby cells prepared from proliferative tissue, but had no effecton its production by cells prepared from secretory endometrium.PP14-stimulated TNF- levels typically only reached a maximumof two times basal values.     

Reactive oxygen species stimulate prostaglandin F2 alpha production in human endometrial stromal cells in vitro

BACKGROUND: The present study was undertaken to investigatethe effect of reactive oxygen species on prostaglandin F2 (PGF2)production by human endometrial stromal cells (ESC). METHODSAND RESULTS: Isolated ESC were incubated with hydrogen peroxide,which induces lipid peroxidation. Hydrogen peroxide increasedboth intracellular and medium concentrations of PGF2 (P <0.01). A time course study showed that hydrogen peroxide significantlyincreased PGF2 concentrations in the medium after 6 h incubation(P < 0.01), after which no further increase was observed.To study whether the increase in PGF2 production caused by hydrogenperoxide was mediated by cyclooxygenase, ESC were incubatedwith indomethacin (0.5 µg/ml), an inhibitor of cyclooxygenase,in the presence of hydrogen peroxide. Indomethacin significantlyblocked the increases in PGF2 production caused by hydrogenperoxide (P < 0.01). Hydrogen peroxide also increased PGF2production by decidualized ESC (P < 0.01), induced by theincubation with medroxyprogesterone acetate (10^–6 mol/l)and oestradiol (10^–8 mol/l). CONCLUSIONS: Reactive oxygenspecies stimulate PGF2 production in ESC, suggesting that theymight influence endometrial function by regulating PGF2 production.