Secretion of plasminogen activator by cultured rat endometrial stromal cells from uteri differentially sensitized for the decidual cell reaction

Endometrial stromal cells from rat uteri differentially sensitized for the decidual cell reaction in vivo and which undergo differing degrees of decidualization in vitro were cultured and plasminogen activator (PA) in the medium determined. The cells were obtained by enzymatic dispersion from the uteri of ovariectomized, steroid-treated rats at the equivalent of day 4, 5, or 6 of pseudopregnancy or on day 5 from rats treated on day 4 with 0, 0.3, or 1.0 μg estradiol (low, intermediate, or high dose of estradiol, respectively) and cultured for 24, 48, or 72 hr. For cells from day 4, 5, and 6 uteri cultured under control conditions, PA activity in the medium was greatest for day 5 cells, which were from uteri maximally sensitized for decidualization both in vivo and in vitro. By contrast, for cells from low-, intermediate-, and high-estradiol uteri, PA activity in the medium was greatest for the high-estradiol cells; these cells do not undergo decidualization in vivo or in vitro to the same extent as intermediate-estradiol cells. Indomethacin, an inhibitor of prostaglandin (PG) synthesis, reduced PGE₂ accumulation to nondetectable amounts and for most cultures decreased PA activity in the medium, suggesting that endogenous PG production regulated in part PA secretion under control conditions. The addition of PGE₂ with indomethacin increased PA activities above those under control conditions, but activities were still lower for day 4 and 6 cells compared with day 5 cells, and for low- and intermediate-estradiol cells compared with high-estradiol cells. This indicates that the differences in PA secretion are not explainable by differences in PGE₂ production. Northern blot analysis of RNA from cells cultured for 72 hr under control conditions did not reveal significant differences in steady-state concentrations of mRNA for urokinase-type PA or plasminogen activator inhibitor 1, but those for tissue-type PA were lower in day 6 cells compared with day 4 and 5 cells. It is concluded that PA activity secreted by the cultured endometrial stromal cells, although controlled in part by the endocrine milieu to which they were exposed prior to culture, does not simulate decidualization in vitro and, therefore, that PA activity is not a marker for decidualization in vitro. Mol. Reprod. Dev. 49:268–276, 1998. © 1998 Wiley-Liss, Inc.     

Regulation of cytosolic phospholipase A2 in rat endometrial stromal cells: the role of epidermal growth factor

The effect of epidermal growth factor on the levels of cytosolic phospholipase A2 mRNA and protein in cultured rat endometrial stromal cells isolated from uteri sensitized for the decidual cell reaction was examined. Treatment with epidermal growth factor increased the steady-state cytosolic phospholipase A2 mRNA and protein levels as demonstrated by Northern and Western blot analyses, respectively. Immunocytochemical analysis demonstrated an increase of cytosolic phospholipase A2 protein in most cells, as opposed to a small subpopulation of cells in culture. These results show that epidermal growth factor causes an increase in steady-state cytosolic phospholipase A2 mRNA and protein levels in rat endometrial stromal cells from uteri sensitized for the decidual cell reaction. Epidermal growth factor receptor ligands may regulate cytosolic phospholipase A2 and thus prostaglandin production in the endometrial stromal cells during implantation.     

Role of interleukin 1 in the regulation of cyclooxygenase gene expression in rat endometrial stromal cells.

Interleukin 1 alpha (IL-1 alpha) stimulates prostaglandin production and cyclooxygenase activity in endometrial stromal cells isolated from the uteri of ovariectomized rats that have been sensitized for the decidual cell reaction. The aim of the present study was to examine the effect of IL-1 alpha on the amount of cyclooxygenase mRNA and protein in these cells. Treatment with IL-1 alpha (20 ng ml-1) for 24 h significantly increased steady-state concentrations of cyclooxygenase 2 (COX-2) mRNA and protein in the cells, as determined by northern and western blot analyses, respectively. Cyclooxygenase 1 (COX-1) mRNA and protein were not detected. Dexamethasone (5 mumol l-1) prevented the IL-1 alpha-induced increase in COX-2 steady-state mRNA. Immunocytochemical staining of COX-2 in the treated cells indicated that IL-1 alpha increased staining, while dexamethasone inhibited this increase. Furthermore, the changes in staining were generalized and not confined to a small subpopulation of cells. These data demonstrate that IL-1 alpha increases steady-state concentrations of COX-2 mRNA and protein in endometrial stromal cells isolated from the uteri of rats that have been sensitized for decidualization.     

人子宫内膜纤蛋白溶酶元激活因子及其抑制因子的分布与调控

人子宫内膜中存在组织型(tPA)及尿激酶型(uPA)两类纤蛋白溶酶元激活因子,其含量在增殖期高于分泌期。本文应用免疫组织化学定位证实uPA及tPA两类抗原存在于子宫内膜的腺体细胞和间质细胞中。应用SDS-PAGE分高蛋白质,继而应用纤蛋白-琼脂糖铺盖技术测得离体培养下间质细胞仅释放tPA,腺体细胞仅释放uPA,但两种细胞均分泌PA的抑制因子(PAI)。培液中加入孕酮,明显抑制PA和刺激PAI生成。雌二醇作用与孕酮相反。某些肽类激素hCG、PRL、GnRH及cAMP作用基本与雌二醇相同。但福司克林(FK)则刺激间质、腺体两种细胞产生tPA及少量uPA,抑制PAI生成。本工作表明人子宫内膜中存在PA及PAI作用相反的酶,受激素调控,其生理意义尚待进一步探讨。     

Expression of plasminogen activator and plasminogen activator inhibitor mRNA in human fibroblasts grown on different substrates

mRNA levels for urokinase type plasminogen activator (uPA), tissue type plasminogen activator (tPA), plasminogen activator inhibitor-1 (PAI-1) and plasminogen activator inhibitor-2 (PAI-2) were examined in human diploid (neonatal foreskin) fibroblasts grown in 200-ml microcarrier suspension culture. Four different substrates were used. These included gelatin-coated polystyrene plastic, DEAE-dextran, glass-coated polystyrene plastic and uncoated polystyrene plastic. Our previous studies have shown that culture fluids from diploid fibroblasts grown on DEAE-dextran contained higher levels of plasminogen-dependent fibrinolytic activity than culture fluids from the same cells grown on other substrates. The increased plasminogen activator activity was due largely to elevated amounts of tPA (In Vitro Cell. Develop. Biol. 22: 575–582, 1986). The present study shows that there is a corresponding elevation of tPA mRNA in diploid fibroblasts cultured on DEAE-dextran relative to the other substrates. There does not appear to be any difference in uPA mRNA or in mRNA for PAI-1 or PAI-2 produced by the same cells on the four substrates. These data suggest that the influence of the substrate on plasminogen activator production is mediated at the genetic level.     

Regulation of tissue-type plasminogen activator and plasminogen activator inhibitor type-1 in cultured rat Sertoli and Leydig cells

New data are provided to show that (i) rat Sertoli cells produce two types of plasminogen activators, tissue type (tPA) and urokinase type (uPA), and a plasminogen activator inhibitor type-1 (PAI-1); (ii) both tPA (but not uPA) and PAI-1 secretion in the culture are modified by FSH, forskolin, dbcAMP, GnRH, PMA and growth factors (EGF and FGF), but not by hCG and androstenedione (△4); (iii) in vitro secretion of tPA and PA-PAI-1 complexes of Sertoli cells are greatly enhanced by presence of Leydig cells which produce negligible tPA but measurable PAI-1 activity;(iv) combination culture of Sertoli and Leydig cells remarkably increases FSH-induced PAI-1 activity and decreases hCG- and forskolin-induced inhibitor activity as compared with that of two cell types cultured alone. These data suggest that rat Sertoli cells, similar to ovarian granulosa cells, are capable of secreting both tPA and uPA, as well as PAI-1. The interaction of Sertoli cells and Leydig cells is essential for the cells to response to     

The THP-1 cell line is a urokinase-secreting mononuclear phagocyte with a novel defect in the production of plasminogen activator inhibitor-2

Mononuclear phagocytes regulate the generation of plasmin by secreting urokinase-type plasminogen activator (uPA) and plasminogen activator inhibitor-2 (PAI-2). We investigated the production of plasminogen activator (PA) and PA inhibitor by the human monocytic leukemia cell line, THP-1. Similar to U937 monoblast-like cells and peripheral blood monocytes (PBM), THP-1 cells produce a PA that is specifically neutralized by anti-uPA antibody and comigrates with human high molecular mass uPA (54 kDa) on casein-plasminogen zymogaphy. PA activity could be dissociated from intact THP-1 cells by brief treatment with a weak acid-glycine buffer, indicating that the uPA is secreted and bound to receptors on the plasma membrane. Regulation of uPA proceeds normally in THP-1 cells, with cell-associated PA activity increasing from 77 +/- 20 to 163 +/- 26 and 325 +/- 30 mPU/10(6) cells in response to PMA and LPS, respectively; parallel increases in steady state levels of uPA mRNA were observed. In contrast to normal expression of uPA activity, functional PAI-2 could not be demonstrated in either the conditioned media or cell lysates of THP-1 under basal or stimulated conditions. Both U937 and PBM secrete low levels of PA inhibitor activity that increase substantially in response to stimulation with PMA and LPS. Immunoreactive PAI-2, measured by ELISA, was undetectable in THP-1 lysates or conditioned medium, but was consistently present in U937 and PBM, paralleling the presence of PA inhibitor activity. THP-1 cells express low levels of an abnormally sized mRNA for PAI-2 and demonstrate a regulatory defect whereby steady state levels of PAI-2 mRNA are markedly reduced upon stimulation with PMA or LPS. By contrast, U937 and PBM respond to identical stimulation with increases in PAI-2 mRNA. We conclude that THP-1 cells express a structurally abnormal species of PAI-2 mRNA, with complete loss of inhibitory activity as well as altered function of PMA- and LPS-responsive regulatory elements.     

Induction of urokinase-type plasminogen activator by UV light in human fetal fibroblasts is mediated through a UV-induced secreted protein.

Plasminogen activator was previously shown to be induced by UV light in human cells with low capacity to repair UV-induced DNA lesions. We now show that in human fetal fibroblasts UV light enhanced the two mRNA species coding for the urokinase-type plasminogen activator (uPA) and the tissue-type plasminogen activator, but immunological analysis revealed exclusively uPA activity. Several independent and complementary experiments indicated that induction of uPA was mediated, apparently entirely, through a UV-induced, secreted protein (UVIS) in the growth medium of irradiated cells. First, elevation of uPA mRNA after irradiation was severely blocked by cycloheximide. Second, replacement of conditioned medium in irradiated cells while the rate of plasminogen activator induction was maximal rapidly and completely stopped any further increase in uPA activity. Third, addition of the same removed conditioned medium to nonirradiated cells mimicked UV light in enhancing the level of uPA activity as well as that of uPA mRNA. Fourth, UVIS activity was completely lost by treating the conditioned medium with trypsin but not with nucleases. Kinetic measurements indicated that the accumulation of UVIS rather than the induction of uPA by UVIS conferred the rate-limiting step in the overall process of uPA induction. Both UV light and UVIS acted synergistically with inhibitors of DNA repair for uPA induction. Based on these results, a model is proposed implicating relaxation of DNA torsional stress of an as yet undefined DNA sequence(s) in the induction of UVIS, which is then responsible for activation of the uPA gene.     

Prostaglandin E2 regulates production of plasminogen activator isoenzymes,urokinase receptor,and plasminogen activator inhibitor-1 in primary cultures of rat calvarial osteoblasts

The bone resorbing agent, prostaglandin E₂ (PGE₂), was found to alter several components of the plasminogen activator (PA)/plasmin pathway in primary cultures of rat neonatal osteoblast-like cells. The mRNA and activities of both urokinase-type PA (uPA) and tissue-type PA (tPA) were enhanced by PGE₂ treatment. The presence of mRNA for the uPA receptor (uPAR) has been demonstrated in these cells and steady-state levels shown to be greatly enhanced, the response being rapid and sustained for at least 24 hours. mRNA for plasminogen activator inhibitor 1 (PAI-1) was modulated in a biphasic manner, with inhibition of the constitutive level apparent at 4 hours of treatment and stimulation apparent at 12 hours and longer, while PAI-1 protein, measured by an ELISA assay for rat PAI-1, was diminished over this period. Neither PAI-2 mRNA nor mRNA for the broad spectrum protease inhibitor, protease nexin-1 (PN-1), was found to be modulated by PGE₂. Therefore, PGE₂ is likely to stimulate cell surface proteolytic activity, since uPA mRNA and cell-associated activity were elevated, as was mRNA for the cellular receptor for uPA. Although it was not possible to measure uPAR number and affinity it seems likely that elevated uPAR mRNA would translate into increased uPARs which would localize the increased uPA activity to the pericellular region. tPA mRNA and activity were also increased transiently with the activity inhibited with prolonged incubations, apparently by PAI-1. Elevation of tPA mRNA and activity may result in elevated activity within the extracellular matrix as tPA has been reported to associate with several matrix proteins. Thus the early effect of PGE₂ would be to promote proteolysis, both pericellularly and in the extracellular matrix. The inhibition of PAI-1 mRNA and protein, which would contribute to the elevation of activity, is due to PGE₂, but the later stimulatory effect on PAI-1 mRNA may be due to feedback regulation by transforming growth factor beta (TGFβ), secreted by osteoblasts and activated by elevated levels of PA. © 1995 Wiley-Liss Inc.     

Cell-type specificity of interleukins 1alpha and 1beta on prostaglandin and plasminogen activator production in bovine endometrial cells

Interleukin (IL)-1alpha is a potent stimulator of prostaglandin production in bovine endometrium, and IL-1 affects plasminogen activator (PA) activity in several types of cells. In this study, we determined the effects of IL-1alpha and IL-1beta on production of the prostaglandins PGF(2alpha) and PGE(2) and on PA activity in cultured bovine endometrial epithelial and stromal cells. We also determined the effects of PGE(2) and PGF(2alpha) on PA activity in these cells. Finally, we used RT-PCR to examine the expression of IL-1alpha, IL-1beta, and IL-1 receptor type 1 (IL-1R) mRNA in cultured bovine endometrial cells. This analysis revealed that IL-1alpha mRNA was present only in the stromal cells, whereas IL-1beta and IL-1R mRNAs were present in both cell types. When cultured cells were exposed to IL-1alpha and IL-1beta at concentrations ranging from 0.006 to 3 nM for 24h, IL-1alpha and IL-1beta were found to dose-dependently stimulate PGE(2) and PGF(2alpha) production in stromal cells (P<0.05) but not in epithelial cells. On the other hand, exposure to IL-1alpha and IL-1beta dose-dependently increased PA activity in the epithelial cells, whereas neither stimulated PA production in the stromal cells. When cells were exposed to IL-1alpha and IL-1beta at concentrations ranging from 0.06 to 3 nM for 24h, the two IL-1s differed in their effects on both PGE(2) and PGF(2alpha) production in stromal cells and had significantly differed in their effects on PA activity in epithelial cells. Exposure to PGE(2) and PGF(2alpha) did not affect PA activity in either stromal or epithelial cells (P>0.05). Taken together, these results suggest the possibility that both IL-1alpha and IL-1beta are produced by the stromal cells, that IL-1beta is produced by the epithelial cells, and that IL-1alpha is a far more potent stimulator than IL-1beta of prostaglandin and PA production in cultured bovine endometrial epithelial and stromal cells.     

The Role of Tissue-type Plasminogen Activator in Rat Corpus Luteum

It is here reported for the first time that luteal cells are capable of secreting plasminogen activators(PA),(both tissue-type,tPA,and urokinase-type,uPA),and plasminogen activator inhibitor type-1(PAl-1).Using organ culture model,we have demonstrated that tPA,but not uPA,showed markedchange during luteolytic period in rat corpus luteum.A great amount oftPA was secreted in corpusluteum on D 14 and D 17 while very low level of tPA activity was detected before D 12.Correspondingly,the progesterone production in the corpus luteum increased gradually in a time-dependent manner from D 1 to D 12 but dropped abruptly to a very low level on D 14.Additionof exogenous tPA to the CL culture caused considerable decrease in progesterone secretion whileinclusion of purified monoclone tPA antibodies in the culture augmented progesterone productionof CL.It is therefore suggested that tPA may play an important role in luteolytic process.     

Collagen modulates gene activation of plasminogen activator system molecules

Skin extracellular matrix (ECM) molecules regulate a variety of cellular activities, including cell movement, which are central to wound healing and metastasis. Regulated cell movement is modulated by proteases and their associated molecules, including the serine proteases urinary-type plasminogen activator (uPA) and tissue-type plasminogen activator (tPA) and their inhibitors (PAIs). As a result of wounding and loss of basement membrane structure, epidermal keratinocytes can become exposed to collagen. To test the hypothesis that during wounding, exposed collagen, the most abundant ECM molecule in the skin, regulates keratinocyte PA and PAI gene expression, we utilized an in vitro model in which activated keratinocytes were cultured in dishes coated with collagen or other ECM substrates. tPA, uPA, and PAI-1 mRNA and enzymatic activity were detected when activated keratinocytes attached to fibronectin, vitronectin, collagen IV, and RGD peptide. In contrast, adhesion to collagen I and collagen III completely suppressed expression of PAI-1 mRNA and protein and further increased tPA expression and activity. Similarly, keratinocyte adhesion to laminin-1 suppressed PAI-1 mRNA and protein expression and increased tPA activity. The suppressive effect of collagen I on PAI-1 gene induction was dependent on the maintenance of its native fibrillar structure. Thus, it would appear that collagen- and laminin-regulated gene expression of molecules associated with plasminogen activation provides an additional dimension in the regulation of cell movement and matrix remodeling in skin wound healing.     

Characterization of morphogenetic and invasive abilities of human mammary epithelial cells: Correlation with variations of urokinase-type plasminogen activator activity and type-1 plasminogen activator inhibitor level

Urokinase-type plasminogen activator (uPA) and one of its inhibitors, the PAI-1, are involved in the proteolytic cascade of matrix degradation during in vivo morphogenesis or metastasis. In the present study, we have characterized the in vitro morphological behavior of human normal and malignant mammary epithelial cells and determined the levels of uPA activity and PAI-1 during these events. Two-dimensional cultures in the presence of inductive fibroblast-conditioned medium (CM) allowed migration of HBL-100 cells and MDA-MB-231 cells. Normal human mammary epithelial cells (HMEC) and MCF-7 cells failed to migrate under these conditions. The epithelial cell migration correlated with an increase in the uPA activity whereas their immobility correlated with both increases in uPA activity and PAI-1 level. In three-dimensional cultures in collagen gel, fibroblasts or fibroblast CM induced branching tubular morphogenesis to HMEC, cord-like extensions to HBL-100 cells and a greater invasiveness ability to MDA-MB-231 cells. These events correlated with an increased uPA activity. In contrast, no morphological rearrangement was observed in MCF-7 cells and this correlated with both increases in uPA activity and PAI-1 level. Altogether, these results show that the in vitro mammary epithelial behavior is under the influence of mesenchymal inductive signals and is in agreement with modifications of uPA activity and PAI-1 levels. Our culture system gives a suitable model to study the mechanisms of mammary development and metastasis and to highlight the involvement of proteases and their inhibitors in cell-cell positioning and cell-matrix reorganization.     

Disruption of the cytoskeleton-extracellular matrix linkage promotes the accumulation of plasminogen activators in F9 derived parietal endoderm

When F9 teratocarcinoma cells are treated with retinoic acid plus cyclic AMP (RACF9) they differentiate into parietal endoderm. Differentiation is accompanied by the acquisition of substrate adhesion sites and a change in the pattern of gene expression, including the synthesis of tissue-type plasminogen activator (tPA). We demonstrate here that dihydrocytochalasin B (DHCB) treatment of differentiating F9 cells prevents the assembly of a structured actin cytoskeleton and generates a more rounded and stellate cell morphology. This morphological change is accompanied by the accumulation of the usually visceral endoderm-specific marker urokinase-type plasminogen activator (uPA) and an increase in tPA levels in comparison to untreated RACF9 controls. The increase in tPA accumulation is preceded by an increase in tPA mRNA levels. These effects are reversible, with a lag, when DHCB is removed, and PA accumulation can be stimulated within 24 h when differentiated cells are exposed to DHCB. Exposure to the microtubule disrupting agent colchicine has no effect on uPA or tPA accumulation. In addition, antibody directed against the beta 1 integrin subunit can also specifically elicit increased PA production. Thus disturbing the cytoskeleton and cytoskeleton associated substrate adhesions promotes PA accumulation.     

Evidence for a role for a uterine renin-angiotensin system in decidualization in rats.

Experiments were performed in vivo and in vitro to determine the effects of enalaprilat, a specific inhibitor of angiotensin-converting enzyme, on various aspects of the decidual cell reaction in rats. Ovariectomized, adult female rats were sensitized for the decidual cell reaction with steroid treatments. For in vivo experiments, intrauterine infusions of enalaprilat alone, and in combination with angiotensin II and prostaglandin E2 (PGE2), were initiated on the day of uterine sensitivity. Enalaprilat inhibited the increases in uterine PG concentrations, endometrial vascular permeability, alkaline phosphatase activity and uterine weight that occurred sequentially following infusion of vehicle. Concurrent infusion of angiotensin II did not reverse any of these inhibitory effects; PGE2 infusion partially, but not completely, reversed the inhibition of increase in uterine weight, although it did not alter the inhibition of endometrial vascular permeability. For in vitro experiments, endometrial stromal cells were obtained from uteri on the day of sensitivity and cultured for up to 3 days in the presence of enalaprilat and angiotensin II. Enalaprilat inhibited in a dose-dependent manner the increases in stromal cell alkaline phosphatase activity and media PGE concentration that occurred in the control cultures; these effects were fully reversed by concurrent treatment with angiotensin II. The inhibition of stromal alkaline phosphatase activity was also reversed by PGE2; conversely, the ability of angiotensin II to reverse the effect of enalaprilat was lost in the presence of indomethacin. These studies provide evidence of a requirement for angiotensin II during the decidual cell reaction in rats and suggest that it acts, at least in part, through a PG-dependent mechanism.     

Synthesis of Urokinase-Type Plasminogen Activator and of Type- 1 Plasminogen Activator Inhibitor in Neuronal Cultures of Human Fetal Brain: Stimulation by Phorbol Ester

Human neuronal brain cultures established from 12- and 14-week-old fetuses synthesize and secrete urokinase-type plasminogen activator (uPA) and limited amounts of tissue-type plasminogen activator (tPA). These cells also produce and secrete the endothelial cell-type PA inhibitor (PAI-1), which forms sodium dodecyl sulfate-stable tPA/PAI-1 complexes in the culture medium. Immunocytochemistry shows a predominant localization of uPA, tPA, and PAI-1 in neuronal cells, with only a very weak positivity detectable in the few glial cells present in these cultures. The protein kinase C (PKC) activator 12-O-tetradecanoylphorbol 13-acetate (TPA) stimulates the synthesis of both uPA and PAI-1, resulting in a final increase in the plasmin-generating capacity of neuronal cell cultures. No significant effect is observed, however, when cells are treated with the TPA analogue 4 alpha-phorbol 12,13-didecanoate, which is inactive as a PKC inducer, or with the neurotrophic polypeptide basic fibroblast growth factor. These data represent the first characterization of the plasmin-generating system in human fetal brain neurons and suggest a role for PKC in the modulation of uPA and PAI-1 synthesis.     

Response of bovine endothelial cells to FGF-2 and VEGF is dependent on their site of origin: Relevance to the regulation of angiogenesis

Angiogenesis, the formation of new capillary blood vessels, occurs almost exclusively in the microcirculation. This process is controlled by the interaction between factors with positive and negative regulatory activity. In this study, we have compared the effect of two well described positive regulators, vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (FGF-2) on bovine adrenal cortex-derived microvascular endothelial (BME) and bovine aortic endothelial (BAE) cells. The parameters we assessed included (a) cellular reorganization and lumen formation following exposure of the apical cell surface to a three-dimensional collagen gel; (b) organization of the actin cytoskeleton; (c) expression of thrombospondin-1 (TSP-1), an endogenous negative regulator of angiogenesis; and (d) extracellular proteolytic activity mediated by the plasminogen activator (PA)/plasmin system. We found that (a) collagen gel overlay induces rapid reorganization and lumen formation in BME but not BAE cells; (b) FGF-2 but not VEGF induced dramatic reorganization of actin microfilaments in BME cells, with neither cytokine affecting BAE cells; (c) FGF-2 decreased TSP-1 protein and mRNA expression in BME cells, an effect which was specific for FGF-2 and BME cells, since TSP-1 protein levels were unaffected by VEGF in BME cells, or by FGF-2 or VEGF in BAE cells; (d) FGF-2 induced urokinase-type PA (uPA) in BME and BAE cells, while VEGF induced uPA and tissue-type PA in BME cells with no effect on BAE cells. Taken together, these findings reveal endothelial cell-type specific responses to FGF-2 and VEGF, and point to the greater specificity of these cytokines for endothelial cells of the microvasculature than for large vessel (aortic) endothelial cells. Furthermore, when viewed in the context of our previous observation on the synergistic interaction between VEGF and FGF-2, our present findings provide evidence for complementary mechanisms which, when acting in concert, might account for the synergistic effect.     

Plasminogen activator activity in differentiating leukemia cells

Plasminogen activator (PA) activity of human promyelocytic leukemia cell line HL-60 was assayed by following the conversion of plasminogen to plasmin and the plasmin-mediated hydrolysis of 14C-labeled globin. When HL-60 cells were induced to differentiate into macrophages by 12-O-tetradecanoyl-phorbol-13-acetate (TPA), cell-associated PA activity and secretion of PA into the conditioned medium increased profoundly. PA activity increased earlier and as a result of lower concentrations of TPA than the ability of the cells to adhere. Exposure to 10(-6)M dexamethasone did not prevent TPA-induced adherence and produced a slight inhibition of cellular PA activity. These findings imply that TPA-induced differentiation of HL-60 cells to macrophage-like cells is associated with induction of PA activity.     

Metabolism of arachidonic acid and prostanoids in human endometrial stromal cells in monolayer culture

In the present investigation, we compared the metabolism of arachidonic acid in human endometrial stromal cells maintained in monolayer culture with that in human decidual tissues. By gas-chromatographic analysis, the distribution of arachidonic acid in glycerophospholipids and in the neutral lipids of decidual tissues and stromal cells in culture was similar. After the addition of [14C]arachidonic acid to the culture medium, steady-state conditions with respect to radioactive labeling of the lipids of the cells were attained after 24 h, except for phosphatidylethanolamine and neutral lipids. The percentage distribution of [14C]arachidonic acid in the lipids of the cells in culture was as follows: phosphatidylcholine, 41%; phosphatidylserine, 5%; phosphatidylinositol, 19%; phosphatidylethanolamine, 22%; neutral lipids, 11%. This distribution of arachidonic acid among the lipids is similar to that in decidual tissue, except for that in phosphatidylethanolamine. The amount of radioactivity in phosphatidylethanolamine continued to increase up to 72 h whereas that in neutral lipids declined after a maximum amount was present at 4 h. In the cells in monolayer culture, [14C]prostaglandin E2 and [14C]prostaglandin F2 alpha were produced from [14C]arachidonic acid, as is true in superfused decidual tissue. The similarities in arachidonic acid metabolism in these cells to that in decidual tissue are supportive of the proposition that endometrial stromal cells in monolayer culture are an appropriate model for the study of the regulation of arachidonic acid release and prostaglandin formation by endometrium and decidua vera.