This study was undertaken to discover any influences of hyperestrogenism on implantation of mouse embryos in vivo and in vitro.
I. In. Vivo Implantation Experiment: Mouse oocytes were collected from superovulated Crj-CD1 (ICR) females. In vitro fertilization was performed according to the method of Toyoda et.al. Pronuclear stage eggs were transferred to m-WM medium added 100 μM EDTA six hours after insemination. Blastocysts developed after ninety-six hours of culture, were transferred into uterine horn of recipients on Day4 of pseudopregnancy which mated with BDF1 sterilized male mice by a modified McLaren's technique. Each amounts (0, 0.1, 1, 10, 100μg/mouse) of estradiol-17β were injected subcutaneously to recipient mice during Day1 to Day5 (Group I) or on Day4 and 5 (Group II) of pseudopregnancy. On the other hand, 1 or 5mg of progesterone was injected simultaneously with
E2 injection to determine the effect of luteal support on implantation. The recipients were killed on Day 9 of pseudopregnancy by cervical dislocation. The proportion of implantation sites and live embryos were examined in bilateral uterine horn.
II An Vitro Implantation Experiment: Uteri were removed from mice at D
4 of pseudopregnancy. The monolayer cells were made by the digestion of uterine epithelium with a trypsin-EDTA-DNase solution according to the technique of Salomon et. al. The uterine cells were seeded with a concentration of 6×10
6 cells/dish in a 35mm culture dish (Falcon 3001) containing BME+AA medium (Spindle, A. I., et. al.), and were allowed to settle and attach to the dish for 24 hours. Five blastocysts fertilized in vitro were transferred directly onto the monolayer cells. Hormonal treatment was performed from the day 1 of co-culture with addition of E
2 or E
2 & P in the medium. Attachment, TbOG and two layered ICM were scored by counting under a dissecting microscope.
Results; In Group I, 1μg or more of E
2 treatment had of on implantation in vivo. In Group II, 10μg or more of E
2 had of on implantation. However, the injection of 1 or 5 mg of progesterone with E
2 overcame the influences. Addition of E
2 or E
2 and P had no affect on attachment, TbOG and two-layered ICM up to a concentration of 10ng/ml E
2. 10
2 ng/ml or more of E
2 had influence on the number of two-layered ICM, and 10
3ng/ml or more of E
2 had affect on TbOG. However, progesterone had no influence on these phenomena.
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