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Experimental Studies |
4, and 1 in Human Endometrial Epithelial Cells in Vitro1
Instituto Valenciano de Infertilidad and Department of Pediatrics, Obstetrics and Gynecology, (C.S., A.M., M.J.G., J.R., A.P.), and Department of Biochemistry and Molecular Biology (J.E.O.), Valencia University, Valencia, Spain; Obstetrics and Gynecology (M.L.P.), Stanford University Medical Center, Stanford, California
Address all correspondence and requests for reprints to: Carlos Simón, Instituto Valenciano de Infertilidad, Guardia Civil 23, 46020 Valencia, Spain. E-mail: ivi{at}futurnet.es
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Abstract |
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1 and 4 integrins was negative in EEC
monolayers studied, regardless of the presence or absence of embryos,
and these findings were confirmed by flow cytometry. The possibility
that the embryonic IL-1 system and leukemia inhibitory factor were
involved in the endometrial ß3 up-regulation was
investigated by neutralizing experiments demonstrating a significant
inhibition of ß3-stained cells when EEC monolayers were
cultured in the presence of EEC/blastocyst-conditioned media with
(n = 4) vs. without (n = 8) antihuman
interleukin (IL)-1 + IL-1ß (1.65% vs. 14.6%;
P < 0.05). Dose-response experiments further
demonstrated an up-regulation of ß3 positive cells when
IL-1 + IL-1ß were added to the medium at a concentration of 10
pg/mL compared with control medium without added cytokines (40%
vs. 20%, n = 4). The functional relevance of the
EEC ß3 up-regulation was tested using a mouse blastocyst
adhesion assay. More mouse blastocysts attached to EEC previously in
contact with human blastocyst (72.7%) compared with those EEC
previously in contact with arrested embryos (40%). Our results
demonstrate the selective effect of a developing human embryo on EEC
expression of ß3, which is maximal when a human
blastocyst instead of an arrested embryo is considered. Furthermore,
the embryonic IL-1 system seems to be involved in the EEC
ß3 up-regulation, reinforcing the concept of precise
paracrine cross-talk between blastocyst and endometrial epithelium
during embryonic implantation. |
Introduction |
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TopAbstractIntroductionMaterial and MethodsResultsDiscussionReferences |
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and
ß) forming homologous groups. Their primary function is to mediate
cell-to-cell and cell-to-extracellular matrix binding by specialized
cell attachment sites, such as the tripeptide sequence Arg-Gly-Asp
(RGD), which is a target sequence for integrin binding (4). This
property provides cells with a number of possibilities to recognize
different adhesive substrates.
Embryonic implantation occurs in humans from cycle days 20–24 in the
so-called implantation window, which is the period of optimal
endometrial receptivity (5). During this time, ß3,
4, and 1 integrins are considered
potential markers of uterine receptivity (6, 7, 8, 9). The 1
subunit is present only during the luteal phase (days 15–28) (7, 8, 9).
The 4 integrin is expressed from days 14–24 (7, 8, 9)
whereas the ß3 subunit appears only on day 20 of the
menstrual cycle and continues in the midluteal phase (7, 9). Therefore
coexpression of ß3, 4, and
1 integrins occur on the glandular epithelium during the
implantation window.
It is clear that these endometrial integrins are hormonally regulated
(10). Integrins 4 and 1 are progesterone
driven; they appear when progesterone production starts and endometrial
progesterone receptors are highest (10, 11). In contrast,
ß3 appears when progesterone production is maximal, and
endometrial progesterone receptors are lowest (10, 11). However, there
is a lack of information regarding the role of the human embryo on the
regulation of these endometrial integrins.
It is also evident that paracrine-autocrine cytokine systems such as the interleukin-1 (IL-1) system (12) and leukemia inhibitory factor (LIF) (13) seem to control at least in part the adhesion phase of implantation in mice. The IL-1 system is present in the human endometrium (14, 15), in the human embryo (16), and at the maternal-embryonic interface (17). Moreover, embryonic IL-1 release occurs only when embryos are cocultured with human endometrial epithelial cells (EEC) or EEC-conditioned media (16). This led us to search for the possibility that the embryonic IL-1 system and LIF could possibly mediate the embryonic regulation of integrins.
The purpose of our study was to investigate the effect of single human
embryos on the regulation of ß3, 4, and
1 integrins in cultured human endometrial epithelial
cells and to search for the IL-1 system and LIF as possible embryonic
factors implicated in this regulation.
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Material and Methods |
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TopAbstractIntroductionMaterial and MethodsResultsDiscussionReferences |
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This project was approved by the institutional review board on the use of human subjects in research at the Instituto Valenciano de Infertilidad, complies with the Spanish Law of Assisted Reproductive Technologies (35/1988) and conforms to guidelines established by the Ethics Committee of the American Society for Reproductive Medicine on human embryo research. Endometrial samples were also obtained after written consent from fertile patients. The clinical and laboratory work was performed at the Instituto Valenciano de Infertilidad in Spain.
Experimental design
Based on our previous work (16), we developed a clinical program in which embryos are cocultured with EEC until blastocyst stage and transferred back to the mother. Embryos were obtained after ovarian superovulation and insemination employing routine in vitro fertilization (IVF) procedures. EEC were isolated from endometrium of fertile patients and cultured until confluence. Endometrium donors were screened as negative for human immunodeficiency virus, hepatitis C and B, VDRL (syphilis), and mycoplasma. Individual human embryos were cocultured with EEC for 5 days (from day 2 until day 6 of embryonic development). After embryo transfer, EEC monolayers were used for immunocytochemistry and flow cytometry (FC), and conditioned media were removed and stored at -20 C for neutralizing experiments. EEC wells were divided according to the embryonic status: EEC with embryos that reached the blastocyst stage, EEC with arrested embryos, and EEC without embryos.
To investigate the effect of single human embryos in regulating
ß3, 4, and 1 integrins on
human endometrial epithelial cells, several approaches were followed.
First, to localize morphologically these molecules, immunocytochemistry
was performed using indirect immunofluorescence. Second, to quantify
the embryonic regulatory effect on EEC monolayers, FC of
ß3, 4, and 1 integrins was
performed. Third, the possibility that the embryonic IL-1 system and
LIF were involved in the endometrial ß3 up-regulation was
investigated by neutralizing experiments and further confirmed by
scanning electron microscopy (SEM) and dose-response experiments.
Finally, the functional significance of the endometrial
ß3 up-regulation induced by the embryo was studied using
a mouse embryonic adhesion assay.
Clinical IVF protocol
The ovarian stimulation protocol using GnRH-a and gonadotropins has been described elsewhere (18). Briefly, a long protocol was used for pituitary desensitization with administration of leuprolide acetate (Procrin, Abbot S.A., Madrid, Spain), 1 mg/day sc, starting in the luteal phase of the previous cycle. After ovarian quiescence, human menopausal gonadotropins were administered (Pergonal and Neo-Fertinorm, Serono, Madrid, Spain) for ovarian stimulation and monitored by serum estradiol (E2) levels and transvaginal ovarian ultrasound scans. Oocyte retrieval was performed 36–38 h after human CG administration (10,000 IU, Profasi, Serono). The standard IVF procedure has been described elsewhere (18). Oocyte-cumulus complexes were evaluated under the dissecting microscope and classified according to Laufer et al. (19). Oocyte-cumulus complexes were incubated at 37 C under 5% CO2 in atmospheric air.
Embryo coculture
Samples of endometria were obtained in the luteal phase from fertile patients undergoing endometrial biopsy (ages 23–39 yr). A portion of each specimen was stained with hematoxylin-eosin for dating according to the method of Noyes et al. (20). Endometrial samples were minced into small pieces <1 mm, and then subjected to mild collagenase digestion. EEC were grown from isolated endometrial glands purified as previously described (21). This cell type was cultured and grown to confluence in steroid depleted medium: 75% DMEM (GIBCO, Grand Island, NY) and 25% MCDB-105 (Sigma, St. Louis, MO) containing antibiotics, supplemented with 10% charcoal-Dextran treated FBS (Hyclone, Logan, UT) and 5 µg/mL insulin (Sigma) as described (21). The homogeneity of cultures was determined by morphological characteristics and verified by immunocytochemical localization of cytokeratin, vimentin, and CD68 antigen as described (21). Functionality of EEC monolayers was demonstrated by the production of PGE2 in response to IL-1 (21) and morphological features displayed by SEM described below. After confluence, growth media were replaced by Hatch 50 Medium (Scandinavian IVF Science AB, Gothenburg, Sweden), and the endometrial cells cocultured with single human embryos.
For embryo coculture, individual human embryos were cocultured with experimental EEC for 5 days in 600-µL drops of Hatch 50 starting at day 2 after insemination when they were at the two- to four-cell stage, with conditioned media being removed every 24 h as described (16). In each experiment, cultured EEC in the same volume (600 µL) of Hatch 50 without any human embryos were used as a control. Embryos achieving the blastocyst stage were transferred back to the mother.
Immunocytochemical staining
After embryo transfer, EEC grown on eight-chamber tissue culture
slides (Lab Tek; Miles Scientific, Naperville, IL) were washed with
0.01 M PBS, 0.15 M NaCl, pH 7.4, and
fixed/permeabilized with methanol/acetone (1/1) for 20 min at -20 C.
An indirect immunofluorescence method was used on EEC monolayers as
previously described (21). To reduce the nonspecific binding, 1% BSA
in PBS was applied for 30 min at 37 C. Thereafter they were rinsed with
PBS, pH 7.4, with 0.05% Tween-20 (PBS-T) (Sigma) twice, and then
incubated with the primary antibodies: TS2/7 mouse antihuman
1 (1:10000 dilution), B-5410 mouse antihuman
4 (1:3000 dilution), AP3 mouse antihuman
ß3 (1:1500 dilution) (generously provided by Bruce
Lessey, University of North Carolina at Chapel Hill) each for 60 min at
room temperature. After rinsing with PBS-T, cells were incubated with a
secondary antibody: fluorescein isothiocyanate (FITC)-conjugated goat
antimouse IgG whole molecule (60 min, 7.8 µg/mL at room temperature)
(Sigma). EEC were visualized and photographed using an Olympus 35 mm
camera attached to an inverted Nikon Diaphot 200 microscope (Nikon,
Madrid, Spain). The specificity of these antibodies has been previously
tested (7, 9, 10, 31). The positive controls were mid-late secretory
human endometrium (data not shown). Control incubations included
deletion of the primary antibody.
FC
For FC experiments, EEC monolayers were detached by treatment
with HBS 1 mM EDTA/trypsin EDTA (1/1), washed in PBS, pH
7.2, centrifuged, and the cell pellet was blocked with 1% BSA in PBS
for 90 min at 4C. After washing, cells were incubated with TS2/7 mouse
antihuman 1 (1:10,000 dilution), B-5410 mouse antihuman
4 (1:3,000 dilution), AP3 mouse antihuman
ß3 (1:1, 500 dilution) (150 min, at 4 C each) according
to provider’s instructions. EEC suspensions were washed and mixed with
FITC-conjugated goat antimouse IgG whole molecule (150 min, 7.8 µg/mL
at 4 C) (Sigma). Cell suspensions were fixed with 1% paraformaldehyde
for 30 min at room temperature, resuspended in PBS, and analyzed in an
Epics Elite flow cytometer (Coulter Cytometry, Hialeah, FL) using an
argon-ion laser tuned at 488 nm and 15 mW. FITC-fluorescence was
collected by 575 DC+ 525BP filters. Data were collected in four-decade
logarithmic amplification. Debris was excluded by analysis of scatter
properties. At least 10,000 events per sample were stored in list-mode
files. Data were expressed as the percentage of stained cells.
Neutralizing experiments
Monolayers of EEC were cultured for 24 h in a pool of
conditioned media from cocultured blastocysts in the absence (n =
8) or presence of saturating concentrations of antihuman IL (hIL)-1ß
(5 µg/500 µL) + anti-hIL-1 (500 µg/500 µL) (n = 4),
with saturating doses of anti-hIL-1 receptor antagonist (ra) (1
µg/500 µL) (n = 4), with recombinant human IL-1ra (10 µg/500
µL) (n = 3) or with blocking doses of anti-LIF (250 µg/500
µL) (n = 4) (all from Genzyme Corp., Cambridge, MA). The
neutralizing concentrations were calculated according to the
manufacturer’s instructions, and the cytokine levels secreted by
single human embryos (16). As negative controls, monolayers of EEC were
incubated with growth media alone. EEC monolayers were either
trypsinized and analyzed for antihuman ß3 by FC or fixed
with glutaraldehide 1% for SEM.
Fixation and SEM
For fixation, 1 mL 1% gluteraldehide (Sigma) in PBS was added to the each EEC monolayer studied and stored in the fixative at 4 C for several days until processed. For SEM, the specimens were dehydrated in alcohol series and then dried according to the critical point method using CO2. After drying, EEC monolayers were mounted on the specimen holder, sputter-coated with gold (14 nm thickness), and observed under accelerated voltage of 10.0 kV at a short working distance in a Cambridge Stereoscan 360 scanning electron microscope. For measurements, the screen magnification was increased to 20,000, and three representative areas of 4 µm2 were examined for each specimen. The specimens were processed all at once.
Dose-response experiments
EEC monolayers were cultured for 48 h in the presence of
increasing concentrations of IL-1, IL-1ß, and IL-1 + IL-1ß of
0, 1, 10, 100, and 1000 pg/mL, respectively. Then EEC monolayers were
trypsinized and analyzed for antihuman ß3 by FC as
described above. To assure that the results observed were not related
to endogenous production of the IL-1 system, IL-1, IL-1ß, and
IL-1ra were measured in the conditioned media of the dose-response
experiments.
Enzyme-linked immunosorbent assay
IL-1, IL-1ß, and IL-1ra concentrations were measured in the
conditioned media from dose-response experiments using a kit from R&D
Systems (Minneapolis, MN) according to the manufacturer’s
instructions. The sensitivity was 0.2 pg/mL, 0.3 pg/mL, and 22 pg/mL,
respectively. Intra- and interassay coefficients of variation were
3.2%, 4.3%, and 3.4% and 5.1%, 5.2%, and 5.3%, respectively.
Mouse embryonic adhesion assay
Blastocysts were flushed from the uteri on day 3.5 of pregnancy from pregnant mare serum gonadotropin/human CG-stimulated 8-week-old Swiss females (CFLP) that were mated and plugged with males of the same strain and age. Mouse blastocyst thereby retrieved were rinsed and placed in human EEC wells and were divided according to the experimental design in EEC previously cocultured with human embryos that achieved the blastocyst stage, EEC cocultured with arrested human embryos, and EEC without human embryos. Between two and five mouse blastocyst were cultured per EEC monolayer at 37 C in a 5% CO2/95% air-humidified incubator. The percentage of blastocysts attached to EEC monolayer was recorded after a 48-h incubation period. To identify embryo adhesion, a small amount of medium was gently flushed on each embryo by a glass pipette as described (22). Briefly, embryos that showed no movement while being observed under an Olympus inverted phase-contrast microscope were considered to be attached.
Statistical analysis
Percentage of cells stained were expressed as mean ±
SEM. For statistical comparison among groups, 
-square or
ANOVA followed by Fisher PLSD test were applied; a P
value 
0.05 was considered statistically significant.
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Results |
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TopAbstractIntroductionMaterial and MethodsResultsDiscussionReferences |
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4, and 1
To investigate the potential embryonic regulation of endometrial
integrins, we first localized the presence of immunoreactive
ß3, 4, and 1 in cultured
human EEC. Immunocytochemical experiments for ß3 were
performed in 12 EEC monolayers (3 EEC wells from embryos that reached
the blastocyst stage, 6 EEC wells from arrested embryos, and 3 EEC
wells without embryos). Also, 4 and 1
were each studied in 12 EEC monolayers (3 and 3 EEC wells,
respectively, from embryos that reached blastocyst stage; 2 and 2 EEC
wells, respectively, from arrested embryos; and 1 and 1 EEC wells,
respectively, without embryos).
Immunostaining for ß3 (Fig. 1
) was
positive in EEC cocultured with human embryos with increased intensity
in EEC wells from embryos that reached blastocyst stage (Fig. 1, F and
G) compared with those EEC wells from arrested embryos (Fig. 1, D and
E). ß3 subunit was morphologically localized to the
plasma membrane (Fig. 1G) with increased intensity in cell- to cell
contact sites (Fig. 1F). No labeling was observed in control
experiments (Fig. 1A). The expression of 4 and
1 subunits was undetectable above background staining in
EEC either with or without embryos.
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To quantify the embryonic regulation of endometrial ß3 integrin, FC analysis of EEC monolayers was performed. In total, 32 EEC monolayers were classified according to the embryonic stage reached by cocultured human embryos: 13 EEC monolayers from embryos that reached blastocyst stage, 12 EEC wells from arrested embryos, and 7 EEC wells without embryos.
In Fig. 2, we present a representative cytofluorometric
analysis of ß3 in EEC monolayers without an embryo (Fig. 2A), with an arrested embryo (Fig. 2B), or with a human blastocyst
(Fig. 2C). In the control EEC well, no appreciable staining of the
secondary antibody was detected under the same experimental conditions.
Data from four different experiments were combined and expressed as the
mean ± SEM of the percentage of
ß3-stained cells (Fig. 2D) in EEC monolayers cocultured
without embryos, with arrested embryos, or with human blastocysts.
These quantitative results demonstrate that individual human
blastocysts up-regulate endometrial epithelial expression of
ß3 compared with those EEC cocultured with arrested
embryos.
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To explore whether the IL-1 system and LIF were involved in the
blastocyst up-regulation of endometrial ß3 integrin,
neutralization experiments were performed as described in
Materials and Methods. Immunostaining for ß3
was analyzed by FC and expressed as percentage of ß3
stained EEC (Fig. 3). Interestingly, the percentage of
stained cells for ß3 was significantly decreased when
IL-1 and IL-1ß from the EEC/blastocyst-conditioned media was
blocked as compared with EEC/blastocysts-conditioned media alone
(2.3 ± 0.7 vs. 14.3 ± 0.7 (P <
0.05). Furthermore, no effect was observed when the antagonist, IL-1ra,
was neutralized (13.9 ± 4) or with the addition of recombinant
hIL-1ra (14.2 ± 3). Finally, LIF neutralization showed a trend
towards an increase in EEC ß3 regulation (23.9 ±
1.9), although it was not statistically significant.
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Figure 4 shows the morphological differences by
optical microscopy among EEC cultured with conditioned media from
cocultured blastocysts alone (Fig. 4A), in the presence of
anti-hIL-1ß + anti-hIL-1 (Fig. 4B) or with anti-IL-1ra (Fig. 4C).
Notice that when the IL-1 system was blocked (Fig. 4B), EEC
became rounded and started to detach from the plates.
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) were healthy and their
shape was more rounded compared with the control EEC cultured without
embryos (Fig. 5A). Also, retraction fibers (a sign of cell migration)
and specifications of the plasma membrane such as short-stubby and
long-hairy microvilli (Fig. 5E) were much more abundant compared with
the control (Fig. 5D). Strikingly, EEC in contact with conditioned
media from cocultured blastocysts developed bulging of the membranes
resembling to pinopods (Fig. 5H), and this feature was not found in the
control EEC (Fig. 5G). When the IL-1 system was blocked in those EEC
cultured with conditioned media from cocultured blastocysts, SEM
demonstrated that antibodies clearly interrupted cell to plastic
adhesion (Fig. 5C) but not cell-to-cell adhesion (Fig. 5, F and I). EEC
appeared generally similar to those cultured without antibodies with
the exception of the retraction fibers (Fig. 5, F and I).
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The percentage of endometrial epithelial
ß3-expressing cells after IL-1, IL-1ß, and IL-1 +
IL-1ß stimulation, as well as the endogenous production of IL-1,
IL-1ß, and IL-1ra by experimental EEC, are shown in Fig. 6. Increasing doses of IL-1 (Fig. 6A) and IL-1ß
(Fig. 6B) did not augment the percentage of ß3-stained
cells. However, when IL-1 + IL-1ß were added, a peak of
ß3-positive stained cells was observed at 10 ng/mL
(37.9 ± 1.2 vs. 17.8 ± 1.4) (Fig. 6C), although
it was not statistically significant. Interestingly, endogenous
production of the IL-1 system by EEC was not regulated by exogenous
IL-1 except in one case. Increasing doses of exogenous IL-1ß
up-regulated endogenous IL-1ra production by EEC (Fig. 6A).
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The effect of previous EEC-blastocysts coculture on the increase of EEC adhesiveness was investigated using an assay for the attachment of mouse blastocysts on cultured EEC. Adhesion of mouse blastocysts was significantly increased in EEC previously cocultured with human blastocysts (12 of 17; 72.7%), as compared with EEC cultured without embryos (3 of 9; 33%), or with EEC previously cocultured with arrested embryos (7 of 15; 40%).
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Discussion |
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TopAbstractIntroductionMaterial and MethodsResultsDiscussionReferences |
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+ IL-1ß.
If we accept the relevance of ß3 as a marker of uterine
receptivity (7, 8, 9), these observations may imply an active role for the
blastocyst in preparing the endometrium and regulating its own ability
to implant. Immunocytochemical experiments have shown weak staining for ß3 in EEC monolayers cultured without embryos. Because this integrin is progesterone-driven (10, 11), and because the endometrial cultures were performed in the absence of steroid hormones, these results were expected. However, we observed increased staining intensity in those EEC wells cocultured with embryos that achieved the blastocyst stage compared with EEC cocultured with arrested embryos in the absence of hormones.
Although immunocytochemistry is a potent tool to visualize the morphological localization, it is at best a semiquantitative technique. For this reason, we used FC analysis as an additional approach to quantify the embryonic regulation of ß3 in EEC. Using this approach, quantitative results confirmed what immunocytochemical experiments suggested, that human blastocysts up-regulate EEC ß3 expression compared with arrested embryos, indicating an embryonic regulation of endometrial ß3 integrin in addition to the already described hormonal regulation (8, 9, 10). Furthermore, EEC ß3 expression decreased when cells were in contact with arrested embryos, possibly suggesting that an antiimplantation factor might be released by dead embryos, or that continued secretion of a proimplantation factor is necessary for optimal ß3 expression.
Having demonstrated that blastocysts up-regulate EEC ß3
expression, we tested the hypothesis that the embryonic IL-1 system and
LIF were involved. This hypothesis was based on two different lines of
evidence. First, the IL-1 system (16) and LIF (13) are produced by the
human embryo and are implicated in the adhesion phase of implantation
in mice (12, 13). Second, IL-1ß is a potent inducer of adhesion
molecules in several systems (23, 24, 25, 26, 27, 28, 29), including decidualized human
stromal cells (30). In fact, the inhibition of human endothelial cell
adhesiveness for human neutrophils and eosinophils is an established
bioassay for recombinant human IL-1ra activity (31). Of particular
interest is the inhibition of the blastocyst-mediated up-regulation of
endometrial epithelial ß3 when both agonists, IL-1 +
IL-1ß, were neutralized and the opposite effect when the antagonist,
IL-1ra, was blocked. No significant effect was detected when LIF was
blocked. These neutralization experiments suggest that the
embryo-endometrial communication in the adhesion phase of human
embryonic implantation is mediated, at least in part, by the embryonic
IL-1 system. Morphological results using light microscopy and SEM
reinforce this biochemical finding showing that anti-IL-1 and
IL-1ß antibodies have clearly interrupted cell-to-plastic and not
cell-to-cell adhesion. Further, dose-response experiments suggest that
the optimal dose of IL-1 + IL-1ß to up-regulate EEC
ß3 is 10 pg/mL, which corresponds to the level of these
cytokines secreted by single human embryos cocultured with EEC (16).
Lastly, we assessed the functional relevance of the EEC
ß3 up-regulation induced by a human blastocyst. For this
purpose, we performed a mouse blastocyst adhesion assay on these EEC
monolayers. Although this assay may be biased by species differences
between human EEC and mouse blastocyst, we learned that, under these
conditions, attachment of mouse blastocysts increases when EEC were
previously cocultured with human blastocysts as compared with EEC
previously cultured with arrested embryos or without embryos.
In vitro studies are limited by their own conditions. First, purity and functionality were of great concern. In our cultures, purity has been assessed by immunohistochemical markers such as cytokeratin, vimentin, and CD68. The functionality of EEC monolayers has been analyzed by PGE2 secretion in response to IL-1ß, and by the morphological changes at SEM induced in those EEC by the addition of blastocyst-conditioned media. Second, the origin of cultured EEC is from both lumenal and glandular EEC and the contribution of each cell populations to our cultures cannot be determined.
The purpose of this study was to gain knowledge about embryonic ability
to modulate human endometrial receptivity. This crucial concept has
already been demonstrated in experimental animals such as mice (32),
rabbits (33), and sheep (34, 35); however the role of the human embryo
in regulating human endometrial receptivity has never been
investigated. Based on this and previous studies (12, 14, 15, 16, 17), our
hypothesis (Fig. 7) is that the human embryo secretes
the complete IL-1 system (IL-1 and IL-1ß/IL-1ra) in response to an
unknown endometrial factor, as demonstrated by the selective release in
the presence of EEC (16). The human blastocyst up-regulates endometrial
epithelial ß3 subunit (as demonstrated by FC), and this
activation is triggered by the binding and activation of embryonic
IL-1 + IL-1ß to the endometrial epithelial IL-1R type I (as
demonstrated by neutralizing experiments and SEM). Finally, this
endometrial ß3 up-regulation is functionally relevant
because it increases the ability of the blastocyst to adhere to the EEC
monolayer (as shown in the blastocyst attachment assay). Because
ß3 integrin is consistently expressed by the human embryo
throughout the preimplantation period (36), it would be interesting to
know whether a simultaneous ß3 up-regulation in both
embryo and endometrium occurs. We hypothesize that the normal
hormonally regulated human endometrium is the trigger of molecular
events preparing the blastocyst to efficiently communicate and regulate
the endometrium to implant.
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Acknowledgments |
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1, B-5410
mouse antihuman 4, and AP3 mouse antihuman
ß3 antibodies. We are also indebted to Dr. George Nikas
(Hammersmith Hospital, London) for the SEM studies. |
Footnotes |
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Received February 12, 1997.
Revised April 9, 1997.
Accepted April 18, 1997.
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References |
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TopAbstractIntroductionMaterial and MethodsResultsDiscussionReferences |
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, TNF-, IL-1ß, IL-4) on porcine endothelial
cells: induction of MHC and adhesion molecules and functional
significance of these changes. Immunology. 187:127–133.
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E. Dimitriadis, C.A. White, R.L. Jones, and L.A. Salamonsen Cytokines, chemokines and growth factors in endometrium related to implantation Hum. Reprod. Update, November 1, 2005; 11(6): 613 - 630. [Abstract] [Full Text] [PDF]
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M.R. Kim, D.W. Park, J.H. Lee, D.S. Choi, K.J. Hwang, H.S. Ryu, and C.K. Min Progesterone-dependent release of transforming growth factor-beta1 from epithelial cells enhances the endometrial decidualization by turning on the Smad signalling in stromal cells Mol. Hum. Reprod., November 1, 2005; 11(11): 801 - 808. [Abstract] [Full Text] [PDF]
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R. Matorras, F. Matorras, R. Mendoza, M. Rodriguez, J. Remohi, F. J. Rodriguez-Escudero, and C. Simon The implantation of every embryo facilitates the chances of the remaining embryos to implant in an IVF programme: a mathematical model to predict pregnancy and multiple pregnancy rates Hum. Reprod., October 1, 2005; 20(10): 2923 - 2931. [Abstract] [Full Text] [PDF]
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Z. Strakova, P. Mavrogianis, X. Meng, J. M. Hastings, K. S. Jackson, P. Cameo, A. Brudney, O. Knight, and A. T. Fazleabas In Vivo Infusion of Interleukin-1{beta} and Chorionic Gonadotropin Induces Endometrial Changes that Mimic Early Pregnancy Events in the Baboon Endocrinology, September 1, 2005; 146(9): 4097 - 4104. [Abstract] [Full Text] [PDF]
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 F. Dominguez, M. Yanez-Mo, F. Sanchez-Madrid, and C. Simon Embryonic implantation and leukocyte transendothelial migration: different processes with similar players? FASEB J, July 1, 2005; 19(9): 1056 - 1060. [Abstract] [Full Text] [PDF]
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R. R. Gonzalez, B. R. Rueda, M. P. Ramos, R. D. Littell, S. Glasser, and P. C. Leavis Leptin-Induced Increase in Leukemia Inhibitory Factor and Its Receptor by Human Endometrium Is Partially Mediated by Interleukin 1 Receptor Signaling Endocrinology, August 1, 2004; 145(8): 3850 - 3857. [Abstract] [Full Text] [PDF]
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K. Deb, M. M Chaturvedi, and Y. K Jaiswal A 'minimum dose' of lipopolysaccharide required for implantation failure: assessment of its effect on the maternal reproductive organs and interleukin-1{alpha} expression in the mouse Reproduction, July 1, 2004; 128(1): 87 - 97. [Abstract] [Full Text] [PDF]
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 A. Cervero, J. A. Horcajadas, J. MartIn, A. Pellicer, and C. Simon The Leptin System during Human Endometrial Receptivity and Preimplantation Development J. Clin. Endocrinol. Metab., May 1, 2004; 89(5): 2442 - 2451. [Abstract] |
