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The Journal of Clinical Endocrinology & Metabolism Vol. 82, No. 8 2607-2616
Copyright © 1997 by The Endocrine Society

Experimental Studies

Embryonic Regulation of Integrins ß3, {alpha}4, and {alpha}1 in Human Endometrial Epithelial Cells in Vitro1

Carlos Simón, MarÍa José Gimeno, Amparo Mercader, José Enrique O’Connor, José RemohÍ, Mary Lake Polan and Antonio Pellicer

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 {alpha}1 and {alpha}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{alpha} + 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{alpha} + 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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