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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
In the present study, we examined the embryonic regulation of
ß3 integrin in human endometrial epithelial cells (EEC)
at the protein level and analyzed putative embryonic factors
responsible for this regulation. The model employed is based on a
clinical in vitro fertilization program in which single
human embryos were cocultured with EEC until blastocyst stage and then
transferred back to the uterus. After embryo transfer, EEC wells were
divided according to the embryonic status reached: EEC with embryos
that achieved the blastocyst stage, EEC with arrested embryos, and EEC
without embryos. Immunostaining for ß3 was positive in
plasma membrane of EEC. Flow cytometry showed a mean percentage of
ß3-stained cells of 24.1 ± 5.7 in EEC cocultured
with embryos that achieved the blastocyst stage (n = 13)
vs. 9.5 ± 1.6 (P < 0.05) in
those EEC cultured with arrested embryos (n = 12). Immunostaining
for 
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.
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