Ligand-induced autoregulation of IL-2 receptor {alpha} chain expression in murine T cell lines

The IL-2 receptor (IL-2R) is composed of three chains a, ßand . In mice, contrary to the human system, we have previouslydemonstrated that the IL-2Rß complex does not bindIL-2. Therefore, mouse IL-2 response is completely dependenton the expression of the IL-2R gene product. T cell clones expressingmouse IL-2Rß and the human IL-2R transgene have beenstudied. When cells are grown in IL-4, mouse IL-2R is not expressed.However, exposure to IL-2 leads to the expression of the endogenousmurine IL-2R subunit. The T cell line expressing mouse IL-2Rand human IL-2Rß can grow in IL-2 but does not expressendogenous murine IL-2 R. Transfection of these cells with thehuman IL-2R gene restores the capacity to induce murine IL-2R.This result demonstrates that IL-2-IL-2R interactions are requiredfor induction of IL-2R. The kinetics of induction and deinductionof murine IL-2R have been studied using clone 18.III. From negativecells, expression of murine IL-2R is a very slow phenomenon.From cells fully expressing IL-2R, deinduction is a two-stepprocess: after a rapid decrease of IL-2R the cells continueto express, for a long period of time, basal levels of murineIL-2R. When cells expressing basal levels of IL-2R are exposedto IL-2, induction of IL-2R is a very rapid phenomenon. Theautoregulatory loop formed by IL-2-IL-2R therefore displaysdifferent levels of functioning.     

Stimulation of prostaglandin (PG) F2{alpha} and PGE2 release by tumour necrosis factor-{alpha} and interleukin-1{alpha} in cultured human luteal phase endometrial cells

Tumour necrosis factor- (TNF-) and interleukin-1 (IL-1) areimportant mediators of cell signalling in the uterus. Prostaglandins(PG) have been implicated in the increase of endometrial vascularpermeability which occurs during the implantation process. Thisstudy evaluates the effect of these two pleiotropic cytokineson PGF₂ and PGE₂ release from human luteal phase endometrialglandular epithelial cells (GEC) and stromal cells (STC) inculture. Basal PGF and PGE release did not differ significantlyfrom each other or among cell types, and declined significantlywith increasing number of days in culture. On day 3, basal PGrelease had decreased to half of that on day 1 of culture. However,both cell types were still able to respond to the addition ofexogenous arachidonic acid (5 µM) on day 3 of culture,with PG release by GEC being elevated 7- to 10-fold and by STCmoderately, but still significantly, on day 4. The permissiveeffect of arachidonic acid on the stimulation of PG releasemay indicate the down-regulation of phospholipase A₂ with continuedtime in culture. However, the addition of arachidonic acid (5µM) on day 0 of culture, while able to cause significantlyincreased PG release from GEC, had no effect on STC. In contrast,the addition of a combination of arachidonic acid (5 µM),and either recombinant human TNF- (10 µg rhTNF-/I) or10 µg rhlL-1/I, had a synergistic action and caused thesignificantly increased release of PGF and PGE from both celltypes, compared with that achieved with either arachidonic acidor the cytokine alone (although GEC responded more than STC).During the first 24 h after the addition of rhTNF- or rhlL-1,both cytokines stimulated PG release from both cell types ina dose- and time-dependent fashion. Neither cycloheximide (10µM) nor actinomycin D (10 µM) affected basal PGrelease, but both blocked cytokine-induced PG release from bothcell types. These results suggest that there is a differentialcontrol of human endometrial cell PG biosynthesis, and thatPG release may be regulated through gene activation.     

Sharing of the IL-2 receptor {gamma} chain with the functional IL-9 receptor complex

The third subunit, the so-called common (_c) chain, of the IL-2receptor is shared among the receptors for IL-2, IL-4, IL-7and IL-15, and dysfunction of the _c chain is thought to causeX-linked severe combined immunodeficiency (XSCID) ascribed toimpairment of early T cell development. However, cytokines linkedto XSCID are as yet unidentified. A mAb specific for the _c chain,TUGm2, profoundly inhibited cell proliferation in response toIL-9. Another mAb, TUGm3, immunoprecipltated [¹²⁵I]IL-9 cross-linkedwith either the IL-9 receptor or the _c chain. These resultsdemonstrate that the _c chain is included in the functional receptorcomplex for IL-9, which was initially characterized as a T cellgrowth factor and is essential for IL-9-dependent growth signaltransductlon.     

TNF-{alpha} regulates IL-4-induced Fc{varepsilon}RII/CD23 gene expression and soluble Fc{varepsilon}RII release by human monocytes

We examined the regulatory effects of TNF- on IL-4-induced geneexpression of the low-affinity receptor for IgE (FcRII/CD23)in human monocytes and IL-4-induced soluble FcRII (sFcRII) releasefrom monocytes. IL-4-induced FcRII expression on the surfaceof monocytes was reduced by TNF- as early as 1 day after cultureand the effect of TNF- increased with prolonged culture. Thepresent analysis was designed to examine whether or not TNF-could suppress IL-4-induced FcRII mRNA expression and enhancedIL-4-induced sFcRII release. The addition of TNF- to monocytecultures with IL-4 significantly reduced FcRII expression onthe surface of monocytes and significantly increased sFcRIIrelease from monocytes. Over time, there was an inverse relationshipbetween the disappearance of cell surface FcRII and the appearanceof sFcRII in culture supernatants. FcRII mRNA expression inmonocytes cultured with IL-4 was not affected by TNF- when examinedat 6 h after cultivation. When the cells were cultured withTNF- for more than 24 h, however, TNF- down-regulated IL-4-inducedFcRII mRNA levels. This correlated with the kinetics of down-regulationof IL-4-induced FCRII expression on the surface of monocytesby TNF-. These results suggest that TNF-dependent reductionof IL-4-;induced FcRII expression on the surface of monocytesresulted, at least in part, from the suppression of FcRII mRNAexpression and the enhancement of sFcRII release.     

Involvement of {alpha}1 and {alpha}4 integrins in gut mucosal injury of graft-versus-host disease

We have investigated the involvement of adhesion molecules inthe lymphocyte infiltration associated with acute intestinalgraft-versus-host disease (GVHD) induced by injection of C3Hlymph node cells into irradiated (C3H x DBA/2)F₁ mice. Firstwe analyzed the expression profile of adhesion molecules including1, 2, 4, 5, 6, L and ß7 integrins, CD44 and L-selectinof lymphocytes from lymph nodes and gut mucosa in normal mice.In normal mice, intraepithelial lymphocytes (IEL) and laminapropria lymphocytes (LPL) uniquely showed increased expressionof 1, 2 and ß7 integrins, and decreased expressionof L-selectin compared with that of lymphocytes of the lymphnodes and Peyer's patches. In mice with GVHD, IEL and LPL ofdonor lymph node cells origin underwent phenotyplc changes characterizedby the increased expression of 1, L and ß7 integrins,and the loss of L-selectin. The expression profile of adhesionmolecules on IEL and LPL of GVHD mice resembled that of normalmice except for the lack of 2 integrin. Treatment of GVHD micewith anti-1,-4 or-ß7 integrin antibody alone partiallyprevented the mucosal pathology of intestinal GVHD, whereasonly mice treated with anti-1 showed reduced donor lymphocyticinfiltration into the intestinal mucosa. In contrast, treatmentwith anti-L or anti-CD44 antibody did not affect the intestinalGVHD. Furthermore, dual blockade of both 1 and 4 integrins completelyinhibited the mucosal pathology and donor lymphocyte infiltrationof intestinal GVHD. These results indicate that 1 and 4 integrinsplay an important role in the pathology of intestinal GVHD.     

IL-7 receptor alpha+ CD3(-) cells in the embryonic intestine induces the organizing center of Peyer's patches.

Peyer's patch (PP) organogenesis proceeds through three histologically distinct steps: formation of organizing centers expressing VCAM-1 and ICAM-1 in segregated regions of the intestine at 15.5 days post-coitus (d.p.c.) (step I), accumulation of blood cells expressing different sets of surface markers to this region at 16.5-17.0 d.p.c. (step II), and entry of CD3+ and B220+ lymphocytes just before birth (step III). PP formation of both Il7ra-/- and Lta-/- mice is impaired from step I, suggesting involvement of the two molecules at the same timing in PP organogenesis. Expression of lymphotoxin (LT) alpha and LTbeta in IL-7 receptor (IL-7R) alpha+ cells in the intestine indicates that defects of Il7ra-/- and Lta-/- mice are due to functional inability of IL-7Ralpha+ cells in the induction of PP anlage. Blocking of IL-7Ralpha function by a single injection of the antagonistic mAb in 15.5 d.p.c. embryos suppressed appearance of VCAM-1(+) spots and expression of LTalpha and LTbeta in the intestine, which eventually resulted in mice without PP but are otherwise normal. Intestinal IL-7Ralpha+ cells are lymphoid in morphology but CD3(-) and functional in both nu/nu and Rag2-/- mice. These results implicate IL-7Ralpha+ CD3(-) cells as the direct inducer of the organizing center of PP.     

Expression of the IL-2 receptor {gamma} chain on various populations in human peripheral blood

We have established two rat mAbs, TUGH4 and TUGh5, specificfor the human chain of the IL-2 receptor (IL-2R), which isknown to be shared among receptors for IL-2, IL-4 and IL-7.The antibodies bound to cell lines transfected with the human chain gene but not to their parental cell lines, and precipitated65–70 and 80–90 kDa cell surface molecules fromlysates of human T cells surface-labeled with Na¹²⁵I and chemicallycross-linked with ^[125]IL-2 respectively. Flow cytometry withTUGh4 and TUGh5 detected the chain in a wide variety of peripheralblood cell populations including CD4⁺ T cells, CD20⁺ T cells,CD20⁺ B cells, CD56⁺ natural killer cells, CD4⁺ monocytes andgranulocytes, contrasting with expression of the and ßchains of IL-2R.     

Rapid acceleration of neutrophil apoptosis by tumor necrosis factor-{alpha}

We demonstrate here that human necrosis factor-, a potent neutrophilactivator, induces rapid (within 3 h) apoptosis of these cells,i.e. neutrophils treated with this cytokine exhibit (I) lightand electron microscopic changes characteristic to apoptoticcells, (II) reduced propidium iodide binding to DNA, and (III)the ladder form of DNA, as shown by agarose gel electrophoresis.These results suggest that apoptosis acceleration may be involvedin processes by which neutrophils are prevented from damagingtissues.     

In vivo administration of TNF-{alpha} prevents EMC-M virus induced viral encephalitis

EMC-M virus causes a monophasic paralytic syndrome characterizedby encephalltic lesions in the brain and patchy demyellnatinglesions in the spinal cord and nerve roots of BALB/c mice. Sincethe replication of EMC virus in vitro is inhibited by tumornecrosis factor (TNF)- we have studied the effect of in vivoadministration of this cytokine on the acute disease. Our studiesshow that periodic administration of TNF- to animals infectedwith EMC-M reduces viral titers in the brain, and decreasesthe degree of clinical paralysis and the severityof the inflammatorylesions in the brain.     

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.     

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.     

Critical proline residues of the cytoplasmic domain of the IL-5 receptor {alpha} chain and its function in IL-5-mediated activation of JAK kinase and STAT5

The high-affinity receptor (R) for IL-5 consists of a uniquea chain (IL-5Rc) and a ß chain (ßc) thatisshared with the receptors for IL-3 and granulocyte macrophagecolony stimulating factor (GM-CSF). We defined two regions ofIL-5R for the IL-5-induced proliferative response, the expressionof nuclear proto-oncogenes, and the tyrosine phosphorylationof cellular proteins including ßc, SH2/SH3-containingproteins and JAK2 kinase. In the studies described here, wedemonstrate that IL-5, IL-3 or GM-CSF stimulation induces thetyrosine phosphorylation of JAK2, and to a lesser extent JAK1,and of STAT5. Mutational analysis revealed that one of the prolineresidues, particularly Pro352and Pro355, in the membrane-proximalproline-rich sequence (Pro352-Pro353-X-Pro355) of the cytoplasmicdomain of IL-5R is required for cell proliferation, and forboth JAK1 and JAK2 activation. In addition, transfectants expressingchimeric receptors which consist of the extracellular domainof IL-5R and the cytoplasmic domain of ßc respondedtoIL-5 for proliferation and tyrosine phosphorylation of JAK1.Intriguingly, electrophoretic mobility shift assay analysisrevealed that STAT5 was activated in cells showing either JAK1or JAK2 tyrosine phosphorylation. These results indicate thatactivation of JAK1, JAK2 and STAT5 is critical to coupling IL-5-inducedtyrosine phosphorylation and ultimately mitogenesis, and thatPro352 and Pro355 in the proline-rich sequence appear to playmore essential roles in cell growth andin both JAK1/STAT5 andJAK2/STAT5 activation than Pro353 does.     

Implication of the common {gamma} chain of the IL-7 receptor in intrathymic development of pro-T cells

The effects of IL-7 on the growth and differentiation of thymocyteswere analyzed using murine fetal thymua organ cultures (FTOC)in the presence of mAbs specific for the conventional IL-7 receptor(1L-7R) and for the common (c) chain. In FTOC, the developmentof CD4^–CD8^– double-negative thymocytes to CD4+CD8+double-positive (DP) and CD4+ or CD8+ single-positive (SP) cellswas not completely blocked by adding these mAbs, although cellgrowth was reduced by the treatment. To define a developingstage sensitive to the mAbs, most immature thymocytes, Pgp-1⁺c-kit cells, were cultured in the 2-deoxyguartosine treatedfetal thymus. In the presence of both mAbs in the culture, neitherDP nor SP thymocytes developed whereas either of the mAbs partiallyblocked their development. These results indicate that the Cchain is involved in early T cell development as an indispensablesubunlt of the functional IL-7 receptor complex.     

Polymorphism of the human T cell receptor alpha chain variable genes: identification of a highly polymorphic V gene probe

In this study, we report the RFLP of the human T cell receptor (TCR) alpha chain variable gene segments. Using DNA samples from 20 individuals and three restriction endonucleases (BamHI, EcoRI and HindIII), the degree of RFLP of a number of different V gene segments was defined. Half of the V alpha subfamilies (6/12) were characterized by a predominant hybridization pattern, with only a few individuals displaying a second pattern. However, one particular V gene family, V alpha 6, has at least five allelic forms that segregated consistently in familial studies. The V alpha polymorphisms revealed in this study, together with those exhibited by V beta gene subfamilies, should prove useful in studying possible associations between TCR gene usage and disorders of the immune system.     

CD38 expression on mouse T cells: CD38 defines functionally distinct subsets of {alpha}{beta} TCR+CD4 CD8 thymocytes

We have examined CD38 expression on mouse lymphocytes usingthe rat mAb NIM-R5 and demonstrate that CD38 expression is restrictedto {small tilde}8% of thymocytes. Although CD38 is absent fromthe majority of CD4+ CD8^– and CD4^–CD8⁺ T cells,we detected a strong correlation between CD36 expression andß⁺CD4^–CD8^– T cells in the thymus, withnearly 80% of ß TCR⁺CD4^–CD8^– thymocytesbeing CD38⁺. Using heat stable antigen (HSA) and CD38, we dividedß⁺CD4⁺CD8^– thymocytes into four subsets: HSA⁺CD38^–,HSA^– CD38^hi, HSA^–CD3810^low and HSA^– CD38^–.Two established characteristics of ß TCR⁺CD4CD8^–cells, bias towards Vß 8.2 TCR expression and highlevels of IL-4 production, were used to establish a possiblerelationship between the above thymocyte subsets. Our presentdata show that the HSA⁺CD38^– subset is not biased towardsVß8.2 TCR expression whereas the HSA^– CD38^–subset does show this bias (–47%). Neither of these subsetsmake IL-4 upon CD3 mediated stimulation. In contrast, the CD38⁺subsets are heavily biased toward Vß8.2 expressionand produce large amounts of IL-4 upon stimulation, particularlythe CD38^low cells. Taken together, these data suggest that thesefour subsets represent various stages of a possible differentiationpathway for ß TCR⁺ CD4^–CD8^– cells, withthe HSA⁺CD38^– subset being the most Immature while theHSA^–CD38^low subset is the most functionally mature. Thesecharacteristics support the view that ap TCR⁺CD4^–CD8^–T cells represent an independent lineage with a distinct, butas yet obscure, role in immunity     

Over-expression of CD3{varepsilon} transgenes blocks T lymphocyte development

We have reported previously that mice carrying >30 copiesof the human CD3 transgene completely lose their T lymphocytesand NK cells (36). Here we demonstrate by immunohistology thatin the most severely immunodeficient mouse, tg26, the thymusis very small, has sizeable vacuoles and does not contain recognizableT lymphocytes except for a small percentage of Thy- 1⁺ cellsand B cells. Cell surface phenotyping and TCR and -ßrearrangement studies confirm that the arrest in T lymphocytedevelopment precedes the arrest in rag-1^null, rag-2^null andTCRß^null mice. Since the T cell progenitors in whichthe arrest occurred were absent in the transgenic mice, indirectapproaches were taken to examine the causes of the block inT cell development. Analyses of 12 independently establishedmutant mouse lines, generated with five different transgenicconstructs, revealed that the severity of the abrogation inT cell development was dependent on the number of copies oftransgenes. Since the number of transgene copies generally correlatedwith the levels of expression of the transgenic CD3 proteins,we concluded that over-expression of the CD3 protein was thelikely cause of the block in T lymphocyte development. The Tcell immunodeficiency was caused by either the human or themurine CD3 protein. Since transgene coded mRNAs were found insignificantly higher quantities than endogenous CD3 mRNAs infetal thymi on days 13 and 14 of gestation, over-expressiontook place very early in development, probably prematurely.Over-expression of the CD3 transgene in thymocyte precursorsmay therefore affect T lymphocyte development in the absenceof TCR and possibly in the absence of the other CD3 proteins.More importantly, over-expression of the CD3 protein in thymocytesof mice with a low copy number of transgenes had a significanteffect on late thymic development Over-expression of the CD3protein in immature thymocytes mimicked the effects caused byexposure of CD4^–; CD8^– thymocytes to anti-CD3 treatment:apoptosis and lack of TCRß expression. We thereforespeculate that in the homozygous tg26 animals the arrest inT cell development was caused by excessive signal transductionevents rather than by a toxic effect of the transgenic protein.     

IL-7 decreases IL-7 receptor alpha (CD127) expression and induces the shedding of CD127 by human CD8+ T cells

IL-7 receptor alpha (CD127) signaling is essential for T-cell development and regulation of naive and memory T-cell homeostasis. Fewer CD8(+) T cells from HIV-infected patients express CD127 compared with healthy individuals, suggesting that specific host and/or viral factors regulate IL-7 receptor expression. Factors relevant to HIV infection that could potentially decrease CD127 expression on human CD8(+) T cells and the mechanisms by which this occurs were therefore evaluated. IL-7, but not HIV gp120, IL-1-beta, IL-6, IL-10, IL-13, transforming growth factor-beta or tumor necrosis factor-alpha, reduced CD127-surface expression and did so without altering CD127 mRNA expression. Furthermore, IL-7 did not increase the amount of cytoplasmic CD127 in CD8(+) T cells. Interestingly, IL-7 induced the shedding of CD127 from CD8(+) T cells, suggesting a mechanism that may contribute to the increased concentration of CD127 in the plasma of HIV(+) individuals, a novel finding reported here. Naive CD8(+) T cells are more sensitive to IL-7 that mediated the down-regulation of CD127, suggesting that these effects may have particular significance early in T-cell life cycle. Since CD127 down-regulation may be an important contributor to HIV-associated T-cell dysfunction, determining the mechanism thereof may prove to be of considerable significance.     

Estrogen receptors alpha and beta (ERalpha and ERbeta) and androgen receptor (AR) in human sperm: localization of ERbeta and AR in mitochondria of the midpiece

BACKGROUND: The central role of estrogens and androgens in the male reproductive system has focused attention on the presence and distribution of their cognate receptors [estrogen receptor (ER) alpha, ERbeta and androgen receptor (AR)] in male reproductive tissues and cells. Since the presence of steroid hormone receptors in mitochondria of mammalian cells has been well documented, we investigated the possibility of mitochondrial localization of sex steroid hormone receptors in sperm. METHODS AND RESULTS: Applying immunofluorescence labelling and confocal laser scanning microscopy we show that the estrogen receptor beta and the AR of human sperm are specifically enriched in the midpiece, at the site of the mitochondria, which were visualized by labelling with the vital dye CMX. Nuclear and mitochondrial localization of AR was also detected in LnCap human prostate cancer cells. Differentially, most of the ERalpha immunostaining is in the form of a compact zone at a region corresponding to the equatorial segment of the upper post-acrosomal region of the sperm head. Immunoblotting experiments using sperm extracts revealed the presence of a 66 and a 45 kDa protein reacting with the ERalpha antibody, one 64 kDa protein reacting with the ERbeta antibody and a 110 and a 90 kDa protein reacting with the antibody against AR. CONCLUSIONS: Our findings suggest that the differential localization of AR and ER isoforms in human sperm reveals distinct roles of these receptors in the physiology of sperm cells and, perhaps, also in the process of fertilization.     

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.