title: Role of endometrial factors in regulating secretion of components of the immunoreactive human embryonic interleukin-1 system during embryonic development.
abstract: In the study reported here, we localized at the protein level the major components of the interleukin (IL)-1 system in the human embryo, and we investigated the endometrial factors influencing their secretion during embryonic development. To localize these components, we performed immunohistochemical experiments in 44 oocytes and 78 embryos. The following primary antibodies were used: monoclonal mouse anti-human IL-1 receptor type I (IL-1R tl), monoclonal mouse anti-human IL-1 beta, and polyclonal rabbit anti-human IL-1 receptor antagonist (IL-1ra). For embryo culture, human embryos at different developmental stages were cultured in 100-microliters drops of Ham's F-10 medium + 4 mg/ml BSA (n = 33), in 100-microliters drops of Menezo B2 culture medium (n = 18), or in wells with 1 ml of Menezo B2 culture medium (n = 8). For embryo coculture, endometrial stromal cells (ESC) and endometrial epithelial cells (EEC) were isolated from human secretory endometrium and cultured until confluence in 75% Dulbecco's Modified Eagle's Medium and 25% MCDB-105 containing antibiotics and supplemented with 10% charcoal-Dextran-treated fetal bovine serum. Individual human embryos were cocultured with experimental EEC and ESC (n = 23 and n = 4, respectively) for 5 days in 600-microliters drops of Menezo B2 medium, and conditioned medium was removed every 24 h. Human embryos were also cultured with EEC-conditioned medium (n = 9). IL-1 alpha, IL-1 beta, and IL-1ra levels were determined by ELISA in the 24-h culture- or coculture-conditioned media. Immunostaining confirmed the presence of IL-1 beta, IL-1ra, and IL-1R tl in oocytes and embryos in all stages analyzed, with no statistical differences. IL-1 alpha, IL-1 beta, and IL-1ra were absent in conditioned media of cultured embryos and embryos cocultured with ESC. However, when human embryos were cocultured with EEC or with EEC-conditioned medium alone, two different populations of embryos were observed: IL-1 producers (57% and 56%) and IL-1 nonproducers (43% and 44%, respectively). Finally, the IL-1 profile of a single human embryo cocultured with maternal EEC which successfully implanted and developed is presented, this pattern being similar to that described in the IL-1 producer population. These results demonstrate the presence of the IL-1 system in the human embryo. However, the selective release of IL-1 only when embryos were cocultured with EEC or EEC-conditioned medium indicates an obligatory role of the endometrium in the regulation of the embryonic IL-1 system. Furthermore, the differential embryonic production of IL-1 may be related to the implantation capability of the embryos.
authors.affiliation: Department of Pediatrics, Obstetrics and Gynecology, Valencia University School of Medicine, Spain.
authors.forename: M J
authors.initials: MJ
authors.lastname: De los Santos
authors.forename: A
authors.initials: A
authors.lastname: Mercader
authors.forename: A
authors.initials: A
authors.lastname: Francés
authors.forename: E
authors.initials: E
authors.lastname: Portolés
authors.forename: J
authors.initials: J
authors.lastname: Remohí
authors.forename: A
authors.initials: A
authors.lastname: Pellicer
authors.forename: C
authors.initials: C
authors.lastname: Simón
mesh_terms: D000328:Adult; D018920:Coculture Techniques; D017077:Culture Media, Conditioned; D046508:Culture Techniques; D010064:Embryo Implantation / Q000502:physiology; D004622:Embryo, Mammalian / Q000502:physiology; D005314:Embryonic and Fetal Development / Q000502:physiology*; D004717:Endometrium / Q000378:metabolism*; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D053590:Interleukin 1 Receptor Antagonist Protein; D007375:Interleukin-1 / Q000032:analysis / Q000378:metabolism*; D012795:Sialoglycoproteins / Q000032:analysis / Q000378:metabolism
chemical_list: D017077:Culture Media, Conditioned; C507763:IL1RN protein, human; C507764:Il1rn protein, mouse; D053590:Interleukin 1 Receptor Antagonist Protein; D007375:Interleukin-1; D012795:Sialoglycoproteins