| Objective①To investigate the effects of nordihydroguaiaretic acid (NDGA) on the mice embryos growth;②To observe the change of light microscope structure and ultramicrostructure in pre-implantation mice endometrium after treating with NDGA;③To investigate the change of the expression of matrix metalloproteinase-9 (MMP-9) and tissue inhibitor of metalloproteinase-1 (TIMP-1) in pre-implantation mice endometrium after treating with NDGA. Methods We took mature and nulliparous Kunming female mice as subject, three Kunming adult female mice were caged with a male overnight. The following morning, females were checked for the presence of a vaginal sperm plug. The day of vaginal plug was designated as day 1 of pregnancy. 120 Kunming pregnant mice were randomly divided into four groups (low-dose group, middle-dose group, high-dose group and control); low- dose group, middle-dose group, high-dose group were given NDGA (10, 30, 90 mg/kg) by s.c. injection on the day 2 of pregnancy and the control group were given the equal volume of PBS. Half of mice were put to death through cervical vertebrae luxation on the day 4 of pregnancy, then the uteri were removed, scratched one cubic centimeters of endometrium and fixed with glutaraldehyde to observe the change of ultramicrostructure in it. The remaining mice uteri fixed with 4% paraformaldehyde; HE staining was used to observe the chang of structure in mice endometrium on the day 4 of pregnancy and immunohistochemical SP staining was used to measure the expression of MMP-9 and TIMP-1 in mice endometrium on day 4 of pregnancy. Microscopic imaging system took the pictures and then analysed by computer. On the day 8 of pregnancy, the other half was killed to calculate the uterine organ coefficient and the number of the embryos. Results①In the control group, the uterine organ coefficient was (2.387±0.376)% and the number of the embryos were 15.87±1.92; In the low-dose group, the uterine organ coefficient was (2.341±0.106)% and the number of the embryos were 14.93±1.79; There were no significant differences in the uterine organ coefficient and the number of the embryos between the low-dose group and the control; In the middle-dose group, the uterine organ coefficient was (1.350±0.010)% and the number of the embryos were 8.27±0.96; Compared with the control, the uterine organ coefficient and the number of the embryo were decreased in the middle- dose group; In high-dose group, the uterine organ coefficient was(0.850±0.287)%;Compared with the control, the uterine organ coefficient were distinctly decreased in high-dose group; No embryo was observed in high-dose group.②Effect of NDGA on morphological change of mice endometrium: HE staining showed distinctly endometrial development with loosened stroma, big stromal cell, high columnar glandular epithelial cell and luminal epithelial cell in the control and low-dose groups mice endometrium; Compared with the control, the mice endometrium showed clear charges with compactible stroma, small stromal cell, low glandular epithelial cell and luminal epithelial cell in the middle-dose and high-dose groups; Under TEM, In the control and low-dose groups, the membrane of mice endometrial stromal cell was normal; Rough endoplasmic reticula(RER) and mitochondria enriched in mice endometrial stromal cell; Mitochondria enriched in glandular epithelial cell and luminal epithelial cell with short microvilli. Compared with the control, the membrane of mice endometrial stromal cell was broken and mitochondria in most glandular epithelial cell and luminal epithelial cell exhibited vacuole degeneration in the middle-dose and high-dose groups.③Effect of NDGA on the expression of MMP-9 and TIMP-1 in mice endometrium: The result of immunohistochemisty showed MMP-9 and TIMP-1 expressed in the control group mice endometrial stromal cell, glandular epithelial cell and luminal epithelial cell. Compared with the control, the mean optical density (MOD) of MMP-9 and TIMP-1 of the low-dose groups mice endometrial stromal cell, glandular epithelial cell and luminal epithelial cell had no significant difference. Compared with the control, the mean optical density (MOD) of MMP-9 and TIMP-1 of the middle-dose and high-dose groups mice endometrial stromal cell, glandular epithelial cell and luminal epithelial cell had significant difference. Conclusion①NDGA may inhibite mice blastocysts implantation with a dose-dependent manner.②After treating with NDGA, endometrial stromal cell, glandular epithelial cell and luminal epithelial cell in mice endometrium were degeneration in varying degrees.③NDGA may inhibit the expression of MMP-9 and TIMP-1 in the endometrial stromal cell, glandular epithelial cell and luminal epithelial cell.④The expression of MMP-9 and TIMP-1 were imbalance in mice endometrium after treating with NDGA and it may be one of the mechanism induced the failure of implantation.
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