Molecular Human Reproduction, Vol. 9, No. 2, 91-95,
February 2003
© 2003 European Society of Human Reproduction and Embryology
Article |
Transforming growth factor ß1 exerts an autocrine regulatory effect on human endometrial stromal cell apoptosis, involving the FasL and Bcl-2 apoptotic pathways
Submitted on April 20, 2002; resubmitted on November 3, 2002. accepted on November 13, 2002
Departments of 1 Pharmacology and 2 Clinical Chemistry, School of Medicine, University of Crete, Heraklion GR-711 10, Crete, Greece
3 To whom correspondence should be addressed. e-mail: gravanis{at}med.uoc.gr
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ABSTRACT |
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Transforming growth factor ß1 (TGFß1) is expressed in human endometrium. It regulates epithelial cell proliferation and apoptosis. The aim of the present work was to examine the role of TGFß1 on human endometrial stromal cell apoptosis. Primary cultures of isolated stromal cells were obtained from biopsies of late secretory phase endometrium. We have found the following: (i) TGFß1 induced apoptosis of stromal cells in a time- and dose-dependent manner; (ii) blockade of TGFß1’s autocrine/paracrine effect by TGFß1-neutralizing antibodies diminished the basal rate of stromal cell apoptosis; (iii) semi-quantitative Western blot analysis showed that TGFß1 caused a rapid but transient elevation of the pro-apoptotic FasL protein, without affecting the levels of Fas receptor; (iv) TGFß1 increased the levels of the anti-apoptotic Bcl-2 and Bcl-xL proteins, while having no significant effects on the pro-apoptotic proteins Bax and Bak, suggesting the activation of a transient survival mechanism activated in stromal cells as a parallel rescue response to the apoptosis-inducing FasL protein. In conclusion, our data provide evidence that TGFß1 exerts an autocrine pro-apoptotic effect on human endometrial stroma, via the FasL/Fas system.
Key words: apoptosis/Bcl-2/cytokines/endometrium/Fas/FasL/TGFß1
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Introduction |
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Human endometrium undergoes cyclic phases of proliferation, differentiation and degeneration. Endometrial tissue remodelling is the result of alternating cell proliferation and apoptosis. Ovarian steroids and locally produced factors, including cytokines, neuropeptides or growth factors, orchestrate endometrial cell growth (Tanaka et al., 1998; Gravanis et al., 2001). Apoptosis is of crucial importance in human endometrium. Indeed, stromal differentiation to decidua and reorganization of endometrial tissue at the implantation chamber of the blastocyst requires the apoptotic death of large numbers of endometrial cells (Gu et al., 1994). However, the exact regulatory mechanisms controlling these apoptotic events remain unclear.
Transforming growth factor ß1 (TGFß1) is an important growth regulator of human endometrium. Both TGFß1 and its receptors are constantly expressed in human endometrium, suggesting that endometrial TGFß1 exerts local autocrine and/or paracrine effects that occur throughout the menstrual cycle (Kauma et al., 1990; Chegini et al., 1994; Tang et al., 1994; Dumont et al., 1995). Recent experimental findings support the hypothesis that endometrial TGFß1 exerts its main effect during the secretory phase of the menstrual cycle. Indeed, TGFß1 expression is elevated in human endometrium following ovulation, while progestins have been shown to stimulate its production in human endometrial stromal cells in culture (Chegini et al., 1994; Kanzaki et al., 1995; Casslen et al., 1998). It should be noted that TGFß1 exerts growth inhibitory effects in both epithelial and stromal cells of human endometrium; it is thus considered to be one of the most significant factors mediating endometrial cell dynamics (Marshburn et al., 1994; Tang et al., 1994). TGFß1 regulates endometrial cell dynamics by also regulating cell apoptosis. Indeed, TGFß1 is known to be involved in the apoptosis of epithelial cells (Tanaka et al., 1998; Garcia-Velasco et al., 1999).
The aim of the present study was to investigate the role of TGFß1 in the apoptosis of human endometrial stromal cells, and to determine if this effect involves the apoptotic Fas/FasL and Bcl-2-related proteins, which are expressed in endometrial stroma (Gompel et al., 1994; Otsuki et al., 1994; Yamashita et al., 1999; Chatzaki et al., 2001; Selam et al., 2001). For this purpose, purified primary cultures of stromal cells, isolated from biopsies of human secretory endometrium, were exposed to exogenous TGFß1, or to TGFß1-neutralizing antibodies to test the potential autocrine/paracrine effect of endogenous TGFß1 on stromal cell apoptosis (Chatzaki et al., 2000). Apoptosis was quantified by measuring nucleosomal DNA fragmentation and levels of Fas/FasL and Bcl-2-related proteins by semi-quantitative Western blot analysis.
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Materials and methods |
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Primary culture of stromal cells isolated from human endometrium
Specimens of endometrium in the secretory phase of the menstrual cycle (day 22 to day 26) were obtained from women of reproductive age undergoing diagnostic curettage for fertility or hysterectomy evaluation or for benign disease. Ethics committee approval was granted for this study. All patients gave informed consent. The histology and pathology of each biopsy was established. The state of glandular, stromal and vascular development served to date the endometrium, by the criteria of Noyes et al. (1950), as confirmed by Dallenbach-Hellweg (1975). Purified endometrial stromal cell cultures were obtained as described previously (Satyaswaroop et al., 1979). Briefly, tissue was collected in Dulbecco’s modified Eagle’s medium (Gibco BRL, MD, USA) supplemented with 10% fetal calf serum and 1% antibiotic–antimycotic solution, trimmed and minced mechanically and digested for 90 min at 37°C with 0.25% of Type I collagenase (Sigma Chemicals, St Louis, MO, USA). Separation of stromal and epithelial cells was carried out by filtration through a 45 µm stainless steel sieve permitting passage of stromal cells but not glandular structures. Isolation of stromal cells from contaminating epithelial and blood cells in the filtrate was performed by the more rapid adhesion of stromal cells than contaminants to tissue culture plastic. The collected filtrate was plated for 30 min in 24-well plates or 25 cm2 flasks (Corning Inc., NY, USA). Unattached cells were discarded and the remaining attached stromal cells were supplemented with 1 ml (24-well plates) or 5 ml (252 cm flasks) of culture medium and incubated in a humidified atmosphere of 5% CO2 at 37°C to reach confluence. This method of endometrial cell separation (Satyaswaroop et al., 1979; Makrigiannakis et al., 1995) allows the isolation of highly purified stromal cell cultures containing <5% epithelial cells or macrophages, as judged by immunohistochemistry (using cytokeratin as a marker for epithelial cells and 3C10 as a marker for CD14 positive macrophages) (Van Voorhis et al., 1983; Lachapelle et al., 1996).
Stromal cell cultures at 90% confluence were exposed to: (i) a range of concentrations of recombinant human TGFß1 (0.01–5 ng/ml) (R&D Systems, Inc., Minneapolis, USA), (ii) anti-human TGFß1 neutralizing monoclonal antibody (5 µg/ml; R&D Systems, Inc.), and (iii) non-immune mouse IgG (5 µg/ml) diluted in phenol red-free and serum-free Roswell Park Memorial Institute 1640 medium (Biochrom Co., Berlin, Germany) supplemented with 0.25% bovine serum albumin fraction V (Sigma), insulin from bovine pancreas (5 mg/l), transferrin (5 mg/l) and sodium selenite (5 ng/l) (ITS cell culture supplement; Sigma), 2 mmol/l L-glutamine and 1% antibiotic–antimycotic solution. The dose of TGFß1 was replaced daily.
Quantitative measurement of apoptosis
Apoptosis was quantified by direct determination of nucleosomal DNA fragmentation with the ‘cell death detection’ enzyme-linked immunosorbent assay (ELISA plus) kit (Boehringer Mannheim GmbH, Mannheim, Germany). The assay uses specific monoclonal antibodies directed against histones of fragmented DNA, allowing the determination of mono- and oligonucleosomes in the cytoplasmic fraction of cell lysates. Briefly, after treatment, stromal cells in 24-well plates were centrifuged (200 g) and lysed according to the manufacturer’s manual. Cell lysates were centrifuged again (200 g). The concentrations of mono- and oligonucleosomes contained in the supernatants were determined by anti-histone–biotin antibody staining. The concentration of nucleosome–antibody complexes was determined photometrically using 2,2'-azino-di(3-ethylbenzthiazolin-sulphonate) as substrate. The optical density was read on a Dynatech MicroELISA reader (Chantilly, VA, USA) at a wavelength of 405 nm. The data are expressed in photometric units. Each unit corresponds to 
12 500 apoptotic cells. The detection limit of the assay is 625 apoptotic cells.
Semi-quantitative Western blot analysis
At the end of each experiment, cells in the flasks were washed twice with phosphate-buffered saline, removed by scraping and centrifuged at 800 g. Cell lysis was performed at 4°C by shaking the pellet vigorously for 30 min and reconstituting in a lysis buffer composed of 50 mmol/l Tris–HCl pH 8, 150 mmol/l NaCl, 0.1% sodium dodecyl sulphate (SDS), 0.5% sodium deoxycholate, 1% detergent NP-40 and freshly added protein inhibitors 10 µg/ml phenylmethylsulphonyl fluoride and 1 µg/ml aprotinin. Solid cellular debris was removed by centrifugation at 12 000 g for 15 min. The cytoplasmic fractions were collected and stored at –80°C. Protein concentration was measured by the Bio-Rad Protein Assay Kit II (Bio-Rad Laboratories, Hercules, CA, USA) following the instructions of the manufacturer. Samples of cytoplasmic protein fractions, containing 20 µg of protein, were solubilized with SDS–polyacrylamide gel electrophoresis sample buffer and electrophoresed through a 12% SDS gel. The resulting protein bands were transferred to nitrocellulose membranes, using an LKB electroblot apparatus (LKB, Bromma, Sweden). Standard Western blotting procedures were employed, as previously described (Dermitzaki et al., 2000). Band intensities were quantified by PC-based Image Analysis (Image Analysis Inc., Ontario, Canada). The following antibodies were used: as primary antibody, anti-human Bcl-2 monoclonal antibody (clone 124, 1:200; Dako A/S, Glostrup, Denmark), rabbit polyclonal anti-sera against Bax, Bak and Bcl-xs/l (1:100; Santa Cruz Biotechnology, Inc., CA, USA), anti-Fas (1:2500) and anti-FasL (1:1000; Transduction Laboratories, Lexington, KY, USA) and, as secondary antibody, goat peroxidase-conjugated anti-mouse IgG (1:10 000; Chemicon International Inc., Temecula, CA, USA) or anti-rabbit IgG (1:4000; Immunotech, Marseille, France). For purposes of normalization, the blots were also stained with a monoclonal anti-actin antibody in a dilution of 1:400 (Amersham, Amersham, UK).
Statistical analysis
Results are presented as the means ± SE and as percentage changes compared with non-treated controls. All apoptosis quantification experiments were performed in triplicate, in at least four stromal cell preparations from different biopsies (n = 4). In semi-quantitative Western blot experiments, the concentration of the antigen was normalized per actin control. Each treatment was performed in triplicate, and in at least four stromal cell preparations from different biopsies (n = 4). For statistical evaluation of data normalized to actin, we used analysis of variance, post-hoc comparison of means. Additionally, we have performed the non-parametric Kruskal–Wallis test for the evaluation of percentage changes to control for n independent experiments. P 
0.05 was considered significant.
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Results |
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TGFß1 induces apoptosis of endometrial stromal cells in vitro
Primary cultures of purified human endometrial stromal cells were exposed to TGFß1 (0.5 ng/ml) for 4 days and the rate of apoptosis was quantified as described in Materials and methods. The dose of TGFß1 was replaced daily. Results are expressed as percentage of basal apoptosis on day 1, as apoptosis at day 0 (time of plating the cells in the presence of various agents) was found to be minimal. The apoptotic process became measurable at the completion of day 1. During the period of exposure the rate of basal apoptosis increased, reaching a maximum on day 3 (156.4 ± 18.2 of non-treated control day 1, n = 4) (Figure 1A). TGFß1 induced stromal cell apoptosis almost 2-fold compared with basal levels on day 1 (180 ± 43% of non-treated controls n = 4, P < 0.05). Similarly, apoptosis levels of TGFß1-treated cells were significantly higher compared with controls on each of the following 3 days (P < 0.05). The effect of TGFß1 on stromal cell apoptosis was dose dependent (Figure 1B). Maximal effects were observed at 0.5 ng/ml, corresponding to a final concentration of 0.1 nmol/l, which is approximately equal to the Kd of human TGFß1 receptors (Massague et al., 1987; Chegini et al., 1994).
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Purified primary cultures of human endometrial stromal cells produce TGFß1 in their supernatant (Chatzaki et al., 2000). Thus, in order to examine autocrine or paracrine effects of the endogenously produced TGFß1, apoptosis was measured in the presence of a specific human TGFß1-neutralizing anti-serum. Blockage of TGFß1 by its antibodies during day 1 resulted in a statistically significant decrease in apoptosis by 21.9 ± 1.8% compared with controls (Figure 1C, n = 7, P < 0.01). This effect was not detectable in control experiments using non-immune mouse IgG (Figure 1C).
TGFß1 induces the Fas/FasL apoptotic system
The effect of TGFß1 on the Fas/FasL system of endometrial stromal cells was examined using semi-quantitative Western blot analysis. Stromal cells were exposed to TGFß1 (0.5 ng/ml) for 3 and 24 h and the levels of Fas and FasL were determined in cell homogenates. Time-points were selected based on previous observations (Garcia-Velasco et al., 1999; Chatzaki et al., 2001). Results are presented in Figure 2. Blotting with anti-Fas and anti-FasL antibodies revealed bands at 45 and at 37 kDa corresponding to the Fas and FasL antigens respectively. Band density was normalized as per corresponding actin bands. Our results show that TGFß1 caused a statistically significant acute increase in the FasL content at 3 h of treatment (216.4 ± 18.3% compared with controls, n = 4, P < 0.05). The stimulatory effect of TGFß1 was transient, since it was not measurable 24 h after treatment (64 ± 13% of non-treated controls, n = 4). In contrast, no effect of TGFß1 was observed on expression of Fas receptor (117 ±10 and 96 ± 8% of non-treated controls for 3 and 24 h respectively, n = 5).
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TGFß1 induces the anti-apoptotic members of the Bcl-2 family of proteins
We have previously reported that endometrial stromal cells in primary culture express the Bcl-2-related proteins Bcl-2, Bcl- xs/l, Bak, and Bax (Chatzaki et al., 2001). The effect of TGFß1 on these proteins was tested in stromal cell primary cultures. Cells were lysed after 3 and 24 h of treatment with TGFß1 (0.5 ng/ml). Cell homogenates were subjected to semi-quantitative Western blot analysis, using antibodies against Bcl-2, Bcl-xl, Bax and Bak (Figures 3 and 4). Data were corrected for protein content by staining with an anti-actin antibody. The band revealed by each antibody was of the predicted size, thus confirming previously published observations (Chatzaki et al., 2001). Exposure to TGFß1 resulted in an increase in the anti-apoptotic proteins Bcl-2 (169 ± 18 and 144 ± 21% of non-treated controls, n = 4, P < 0.05) and Bcl-xl (208 ± 25 and 162 ± 14% of non-treated controls, n = 4, P < 0.05) at 3 and 24 h respectively (Figure 3). In contrast, there was no statistically significant effect on the apoptosis-inducing homologues of the Bcl-2 family, Bax and Bak (Figure 4), at any time point.
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Discussion |
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Cell apoptosis is crucial in the physiology of endometrial stroma. Indeed, apoptotic cells are found scattered in endometrial stroma during the proliferative phase, their population increasing progressively during the secretory phase, peaking in the menstrual phase. We have previously shown that TGFß1, a protein known to induce apoptosis of endometrial epithelial cells, is produced by human endometrial stromal cells in culture (Chatzaki et al., 2000). Our current data suggest that endometrial stromal TGFß1 exerts a significant autocrine and/or paracrine pro-apoptotic effect on human endometrial stromal cells in culture. More specifically, we have found that TGFß1 increased the apoptotic rate of endometrial stromal cells in primary culture over a period of 4 days. Blockage of endogenously secreted TGFß1 with neutralizing antibodies decreased basal apoptosis by 22%, suggesting that endogenous TGFß1 exerts a tonic pro-apoptotic effect on stromal cells. The significant but rather narrow decrease of basal apoptosis, shown in the presence of TGFß1-neutralizing antibodies, could be attributed to the low levels of active TGFß1 produced in stromal cell culture. TGFß1 is secreted by stromal cells mainly in its latent form which does not bind TGFß receptors. Thus, the autocrine/paracrine action of TGFß1 might become more evident by blocking the TGFß receptor type II with a specific antagonist (currently not available), followed by treatment with TGFß1. Other autocrine/paracrine factors, not inhibited by TGFß1 antibodies, could also contribute to the regulation of basal apoptosis. In particular, locally produced platelet-derived growth factor (PDGF) and
-opioids have recently been shown to exert pro-apoptotic effects on human endometrial stromal cells in culture, inducing the pro-apoptotic FasL protein (Chegini et al., 1992; Garcia-Velasco et al., 1999; Chatzaki et al., 2001).
TGFß1 has been previously linked to apoptotic phenomena in endometrial stroma of other species. Moulton has shown that various TGFß isoforms stimulate apoptosis of rat endometrial stromal cells, decidualized in vitro (Moulton, 1994). Our results showed that neutralization of the endogenously secreted TGFß1 with a specific antiserum for this isoform, caused a significant reduction in basal apoptosis of stromal cells. Stromal cells in Moulton’s study were previously exposed to steroid treatment for the experimental induction of decidualization, representing an early pregnancy rather than a normally cycling secretory endometrium. It appears from previous reports (Lea et al., 1992; Manova et al., 1992) that the formation of the decidual membranes in the rat uterus is followed by a switch from TGFß1 expression to the ß2 isoform, suggesting that TGFß2 may be more important in inducing apoptosis in the rat decidua. It should be stressed that TGFß1 was not detectable in the in-vitro decidualized rat stromal cells in Moulton’s study, in contrast to data from our laboratory showing that stromal cell cultures from normally cycling secretory human endometrium produce high amounts of TGFß1 (Chatzaki et al., 2000). These previously published data taken together with our current study suggest the existence of species specificity in the pro-apoptotic effects of TGFß on endometrial stroma. Indeed, we have previously shown an interaction between TGFß1 with another paracrine regulator of human endometrial stromal cell apoptosis, endometrial -opioids (Chatzaki et al., 2000). -opioids exert an inhibitory effect on TGFß1 production by endometrial stromal cells, and induce apoptosis of the same cells (Chatzaki et al., 2001). It is thus possible that endometrial TGFß1 and -opioids form a local regulatory loop within endometrium, protecting stromal cells from extensive cell death. Counteracting molecular mechanisms of this type are known to account for the flexible balance of major cellular events that lead to cell survival or death.
Induction of stromal cell apoptosis by TGFß1 was accompanied by an increase in the levels of the pro-apoptotic FasL protein. The Fas/FasL system represents a major pathway for the induction of apoptosis. Upon binding to its receptor, FasL initiates apoptotic signals that induce cell death (Nagata, 1994). In our hands, the effect of TGFß1 on FasL was acute at 3 h of treatment, and transient since it was not measurable after 24 h of treatment. It should be noted that the effect of TGFß1 on stromal cell apoptosis persisted for up to 4 days. Thus, it is possible that the Fas/FasL system mediates an early response mechanism for cells to the apoptotic inducer that may no longer be necessary for long-term actions, requiring the transcription of other apoptosis-related genes. We did not detect any significant effect of TGFß1 on Fas receptor expression by stromal cells. These findings suggest that TGFß1 exerts a pro-apoptotic action, inducing the apoptotic signal, but does not affect the sensitivity of stromal cells to it. This phenomenon has already been described in the effect of TGFß1 on epithelial cells of human endometrium. Indeed, TGFß1 enhances susceptibility to FasL-induced apoptosis without affecting Fas expression in the human endometrial epithelial cell line HHUA (Tanaka et al., 1998). This mode of pro-apoptotic actions of TGFß1 is a direct contrast with that of endometrial -opioids. -opioid-induced apoptosis is mediated through up-regulation of Fas receptor (Chatzaki et al., 2001).
In an effort to uncover other pathways of apoptosis involved in the pro-apoptotic effect of TGFß1 on endometrial stromal cells, we have also tested its effects on members of the major family of apoptosis-related factors, the Bcl-2 proteins. TGFß1 did not have any significant effect on the levels of pro-apoptotic proteins Bax and Bak. However, TGFß1 elevated the protein content of the apoptosis-preventing Bcl-2 and Bcl-xL proteins. The effect of TGFß1 on anti-apoptotic Bcl-2 proteins remained elevated for up to 24 h, in contrast to its acute action on pro-apoptotic FasL, which lasted for only 3 h. It is possible that a Bcl-2-related mechanism is activated that counteracts the stress signals generated by the apoptosis-inducing factor FasL in order to rescue the cells from programmed cell death. Similar phenomena have been recently described in other cell types, mainly of tumoral origin (Jarpe et al., 1998). In pheochromocytoma PC12 cells, prevention of apoptosis caused by -opioids is accompanied by decreased anti-apoptotic Bcl-2 protein levels (Dermitzaki et al., 2000). Additionally, in human medullary thyroid carcinoma cells, rapamycin up-regulates anti-apoptotic Bcl-2 proteins while inducing apoptosis (Pfragner et al., 2000). The induction of apoptosis and the parallel increase of anti-apoptotic Bcl-2 protein by TGFß1 needs further investigation. It would be of interest to clarify if these effects of TGFß1 involve direct transcriptional events on Bcl-2 and Bcl-xL gene expression or if they represent the cellular response to induction of apoptosis initiated by FasL. Furthermore, our data, showing that TGFß1 drives stromal cell apoptosis for up to 4 days, in spite of the parallel induction of Bcl-2 apoptosis-preventing mechanisms, suggest that in human endometrial stroma, FasL is the predominant regulator of cell apoptosis which is not circumvented by the longer-lasting increase of the anti-apoptotic bcl-2 proteins.
In conclusion, our results suggest that TGFß1 plays an important role in human endometrial stromal cells, inducing their apoptosis via the FasL/Fas system.
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Acknowledgement |
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This work was supported by a grant from Menarini Hellas.
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REFERENCES |
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Casslen, B., Sandberg, T., Gustavsson, B., Willen, R. and Nilbert, M. (1998) Transforming growth factor beta1 in the human endometrium. Cyclic variation, increased expression by estradiol and progesterone, and regulation of plasminogen activators and plasminogen activator inhibitor-1. Biol. Reprod., 58, 1343–1350.
Chatzaki, E., Margioris, A.N., Makrigiannakis, A., Castanas, E., Georgoulias, V. and Gravanis, A. (2000) Kappa opioids and TGFbeta1 interact in human endometrial cells. Mol. Hum. Reprod., 6, 602–609.
Chatzaki, E., Makrigiannakis, A., Margioris, A.N., Kouimtzoglou, E. and Gravanis, A. (2001) The Fas/FasL apoptotic pathway is involved in kappa-opioid-induced apoptosis of human endometrial stromal cells. Mol. Hum. Reprod., 7, 867–874.
Chegini, N., Rossi, M.J. and Masterson, B.J. (1992) Platelet-derived growth factor (PDGF), epidermal growth factor (EGF), and EGF and PDGF beta-receptors in human endometrial tissue: localization and in vitro action. Endocrinology, 130, 2373–2385.[Abstract]
Chegini, N., Zhao, Y., Williams, R.S. and Flanders, K.C. (1994) Human uterine tissue throughout the menstrual cycle expresses transforming growth factor-beta 1 (TGF beta 1), TGF beta 2, TGF beta 3, and TGF beta type II receptor messenger ribonucleic acid and protein and contains [125I]TGF beta 1-binding sites. Endocrinology, 135, 439–449.[Abstract]
Dallenbach-Hellweg, G. (1975) Histopathology of the Endometrium. Springer-Verlag, Berlin, pp. 325–346.
Dermitzaki, E., Chatzaki, E., Gravanis, A. and Margioris, A.N. (2000) Opioids transiently prevent activation of apoptotic mechanisms following short periods of serum withdrawal. J. Neurochem., 74, 960–969.[CrossRef][ISI][Medline]
Dumont, N., O’Connor-McCourt, M.D. and Philip, A. (1995) Transforming growth factor-beta receptors on human endometrial cells: identification of the type I, II, and III receptors and glycosyl-phosphatidylinositol anchored TGF-beta binding proteins. Mol. Cell Endocrinol., 28, 57–66.
Garcia-Velasco, J.A., Arici, A., Zreik, T., Naftolin, F. and Mor, G. (1999) Macrophage derived growth factors modulate Fas ligand expression in cultured endometrial stromal cells: a role in endometriosis. Mol. Hum. Reprod., 5, 642–650.
Gompel, A., Sabourin, J.C., Martin, A., Yaneva, H., Audouin, J., Decroix, Y. and Poitout, P. (1994) Bcl-2 expression in normal endometrium during the menstrual cycle. Am. J. Pathol., 144, 1195–1202.[Abstract]
Gravanis, A., Makrigiannakis, A., Zoumakis, E. and Margioris, A. (2001) Endometrial and myometrial corticotropin releasing hormone (CRH): its regulation and possible roles. Peptides, 22, 785–793.[CrossRef][ISI][Medline]
Gu, Y., Jow, G.M., Moulton, B.C., Lee, C., Sensibar, J.A., Park-Sarge, O.K., Chen, T.J. and Gibori, G. (1994) Apoptosis in decidual tissue regression and reorganization. Endocrinology, 135, 1272–1259.[Abstract]
Jarpe, M.B., Widmann, C., Knall, C., Schlesinger, T.K., Gibson, S., Yujiri, T., Fanger, G.R., Gelfand, E.W. and Johnson, G.L. (1998) Anti-apoptotic versus pro-apoptotic signal transduction: checkpoints and stop signs along the road to death. Oncogene, 17, 1475–1482.[CrossRef][ISI][Medline]
Kanzaki, H., Hatayama, H., Narukawa, S., Kariya, M., Fujita, J. and Mori, T. (1995) Hormonal regulation in the production of macrophage colony-stimulating factor and transforming growth factor-beta by human endometrial stromal cells in culture. Horm. Res., 44, 30–35.
Kauma, S., Matt, D., Strom, S., Eierman, D. and Turner, T. (1990) Interleukin-1 beta, human leukocyte antigen HLA-DR alpha, and transforming growth factor-beta expression in endometrium, placenta, and placental membranes. Am. J. Obstet. Gynecol., 163, 1430–1471.[ISI][Medline]
Lachapelle, M., Miron, P., Hemmings, R., Baron, C. and Roy, D.C. (1996) Flow-cytometric characterization of hematopoietic cells in non-pregnant human endometrium. Am. J. Reprod. Immunol., 35, 5–13.
Lea, R.G., Flanders, K.C., Harley, C.B., Manuel, J., Banwatt, D. and Clark, D.A. (1992) Release of a transforming growth factor (TGF)-beta 2-related suppressor factor from post-implantation murine decidual tissue can be correlated with the detection of a subpopulation of cells containing RNA for TGF-beta 2. J. Immunol., 148, 778–787.[Abstract]
Makrigiannakis, A., Zoumakis, E., Margioris, A.N., Theodoropoulos, P., Stournaras, C. and Gravanis, A. (1995) The corticotropin-releasing hormone (CRH) in normal and tumoral epithelial cells of human endometrium. J.Clin. Endocrinol. Metab., 80, 185–189.
Manova, K., Paynton, B.V. and Bachvarova, R.F. (1992) Expression of activins and TGF beta 1 and beta 2 RNAs in early post-implantation mouse embryos and uterine decidua. Mech. Dev., 36, 141–152.[CrossRef][ISI][Medline]
Marshburn, P.B., Arici, A.M. and Casey, M.L. (1994) Expression of transforming growth factor-beta 1 messenger ribonucleic acid and the modulation of deoxyribonucleic acid synthesis by transforming growth factor-beta 1 in human endometrial cells. Am. J. Obstet. Gynecol., 170, 1152–1158.[ISI][Medline]
Massague, J., Cheifetz, S., Ignotz, R.A. and Boyd, F.T. (1987) Multiple type-beta transforming growth factors and their receptors. J. Cell. Physiol., 5, 43–47.
Moulton, B. (1994) Transforming growth factor ß stimulates endometrial stromal apoptosis in vitro. Endocrinology, 134, 1055–1060.[Abstract]
Nagata, S. (1994) Apoptosis regulated by a death factor and its receptor: Fas ligand and Fas, Phil. Trans. R. Soc. Lond. B Biol. Sci., 345, 281–287.[ISI][Medline]
Noyes, R.W., Hertig, A.T. and Rock, J. (1950) Dating the endometrial biopsy. Fertil. Steril., 1, 3–9.
Otsuki, Y., Misaki, O., Sugimoto, O., Ito, Y., Tsujimoto, Y. and Akao, Y. (1994) Cyclic bcl-2 gene expression in human uterine endometrium during menstrual cycle. Lancet, 344, 28–29.[ISI][Medline]
Pfragner, R., Girsch, T., Siegl, V., Tschernatsch, M., Wacheck, V., Jansen, B., Purstner, P., Behmel, A. and Niederle, B. (2000) Bcl-2 and the regulation of apoptosis in human medullary thyroid carcinoma cell lines. Int. J. Mol. Med., 6 (Suppl. 1), S59.
Satyaswaroop, P.G., Bressler, R.S., de la Pena, M.M. and Gurpide, E. (1979) Isolation and culture of human endometrial glands. J. Clin. Endocrinol. Metab., 48, 639–641.
Selam, B., Kayisli, U.A., Mulayim, N. and Arici, A. (2001) Regulation of Fas ligand expression by estradiol and progesterone in human endometrium. Biol. Reprod., 65, 979–985.
Tanaka, T., Umesaki, N., Mizuno, K., Chang, L., Miyama, M., Ohtaki, S. and Ogita, S. (1998) Enhancement of apoptotic susceptibility in human endometrial epithelial cell line HHUA by transforming growth factor-beta 1. Horm. Metab. Res., 30, 61–65.[ISI][Medline]
Tang, X.M., Zhao, Y., Rossi, M.J., Abu-Rustum, R.S., Ksander, G.A. and Chegini, N. (1994) Expression of transforming growth factor-beta (TGF beta) isoforms and TGF beta type II receptor messenger ribonucleic acid and protein, and the effect of TGF betas on endometrial stromal cell growth and protein degradation in vitro. Endocrinology, 135, 450–459.[Abstract]
Van Voorhis, W.C., Stainman, R.M., Hair, L.S., Luban, J., Witmer, M.D., Koide, S. and Cohn, Z.A. (1983) Specific antimononuclear phagocyte monoclonal antibodies. Application to the purification of dendritic cells and the tissue localization of macrophages. J. Exp. Med., 158, 126–145.
Yamashita, H., Otsuki, Y., Matsumoto, K., Ueki, K. and Ueki, M. (1999) Fas ligand, Fas antigen and Bcl-2 expression in human endometrium during the menstrual cycle. Mol. Hum. Reprod., 5, 358–364.
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