Aim: To study the effect of prenatal exposure to diethylstilbestrol (DES) and the role of actin and proliferating cell nuclear antigen (PCNA) on testicular gubernaculum development in fetal male Kunming mice. Methods:...Aim: To study the effect of prenatal exposure to diethylstilbestrol (DES) and the role of actin and proliferating cell nuclear antigen (PCNA) on testicular gubernaculum development in fetal male Kunming mice. Methods: Pregnant mice were randomly assigned to 6 groups and injected with DES subcutaneously from gestational day 9 (E9) to day 17 (E17) at doses of 0, 25, 50, 100, 200 μg·kg-1·d-1 in 0.2 mL dimethyl sulfoxide (DMSO). On E17 they were sacrificed and fetuses quickly removed for fixation. Male fetuses were sliced on serial coronal plane. Histologi-cal changes were observed under the light microscope (LM) and ultrastructural changes with the scanning and transmission electron microscopes (SEM & TEM). The expression intensity of actin and PCNA in the gubernacula was quantitated by immunohistochemistry. Results: The mortality of the fetuses was higher in the DES-treated groups than that in the DMSO and saline controls (P<0.05). Under LM the gubernacula were seen to be poorly developed with smaller bulbs. On SEM the bulbs lose the clear demarcation between the mesenchymal inner core and the muscular outer layer and looked like a small cone instead of the normal cylindrical appearance. On TEM there were some smaller disordered myofibrils and sparse cytoplasmic organelles in the gubernacular muscular cells of the treated groups. The expression intensity of actin and PCNA in the gubernacula was significantly weaker in the treated groups than that in the DMSO and saline controls (P<0.05). Conclusion: DES induces underdevelopment of the gubernacula in a dose-dependent manner in fetal male mice and down regulates the actin and PCNA expression.展开更多
Objective To evaluate the effect of dynein inhibitor on mouse oocyte in vitro maturation and its cyclin B1 transcription level. Methods Immature mouse oocytes were cultured in vitro with a known dynein ATPase activ...Objective To evaluate the effect of dynein inhibitor on mouse oocyte in vitro maturation and its cyclin B1 transcription level. Methods Immature mouse oocytes were cultured in vitro with a known dynein ATPase activity inhibitor-sodium orthovanadate (SOV) to detect the changes of maturation rate, and semi-quantitative RT-PCR and single cell RT-PCR were performed to detect the changes of cyclin B1 mRNA level. Results In dose-dependent experiments, the maturation rates of oocytes were significantly different between 5 mmol/L SOV and control groups (P<0.05), and decreased with SOV increasing doses. However, the elevation of cyclin B1 mRNA level of immatured oocytes cultured for 12 h depended on SOV concentrations ranging from 50 to 500 mmol/L. In incon- tinuity exposed SOV experiments, the maturation rates of oocytes markedly reduced after the first incubation with 400 mmol/L SOV at least for 1 h and were first cultured in SOV-free medium for 4 h or 8 h before exposure to SOV (P<0.05). In time-course experiment, the opposite changes of cyclin B1 mRNA level in oocytes between SOV and control groups were observed. Conclusion Dynein inhibitor might delay oocytes meiosis process, and cause ectopic expression of cyclin B1 in oocytes. Most Oocytes incubated with SOV blocked at germinal vesicles (GV) stage or MⅠto anaphase transition due to dynein dysfunction and ectopic transcription level of cyclin B1.展开更多
文摘Aim: To study the effect of prenatal exposure to diethylstilbestrol (DES) and the role of actin and proliferating cell nuclear antigen (PCNA) on testicular gubernaculum development in fetal male Kunming mice. Methods: Pregnant mice were randomly assigned to 6 groups and injected with DES subcutaneously from gestational day 9 (E9) to day 17 (E17) at doses of 0, 25, 50, 100, 200 μg·kg-1·d-1 in 0.2 mL dimethyl sulfoxide (DMSO). On E17 they were sacrificed and fetuses quickly removed for fixation. Male fetuses were sliced on serial coronal plane. Histologi-cal changes were observed under the light microscope (LM) and ultrastructural changes with the scanning and transmission electron microscopes (SEM & TEM). The expression intensity of actin and PCNA in the gubernacula was quantitated by immunohistochemistry. Results: The mortality of the fetuses was higher in the DES-treated groups than that in the DMSO and saline controls (P<0.05). Under LM the gubernacula were seen to be poorly developed with smaller bulbs. On SEM the bulbs lose the clear demarcation between the mesenchymal inner core and the muscular outer layer and looked like a small cone instead of the normal cylindrical appearance. On TEM there were some smaller disordered myofibrils and sparse cytoplasmic organelles in the gubernacular muscular cells of the treated groups. The expression intensity of actin and PCNA in the gubernacula was significantly weaker in the treated groups than that in the DMSO and saline controls (P<0.05). Conclusion: DES induces underdevelopment of the gubernacula in a dose-dependent manner in fetal male mice and down regulates the actin and PCNA expression.
基金This work was supported by a grant from the National Natural Science Foundation of China (No. 30170481).
文摘Objective To evaluate the effect of dynein inhibitor on mouse oocyte in vitro maturation and its cyclin B1 transcription level. Methods Immature mouse oocytes were cultured in vitro with a known dynein ATPase activity inhibitor-sodium orthovanadate (SOV) to detect the changes of maturation rate, and semi-quantitative RT-PCR and single cell RT-PCR were performed to detect the changes of cyclin B1 mRNA level. Results In dose-dependent experiments, the maturation rates of oocytes were significantly different between 5 mmol/L SOV and control groups (P<0.05), and decreased with SOV increasing doses. However, the elevation of cyclin B1 mRNA level of immatured oocytes cultured for 12 h depended on SOV concentrations ranging from 50 to 500 mmol/L. In incon- tinuity exposed SOV experiments, the maturation rates of oocytes markedly reduced after the first incubation with 400 mmol/L SOV at least for 1 h and were first cultured in SOV-free medium for 4 h or 8 h before exposure to SOV (P<0.05). In time-course experiment, the opposite changes of cyclin B1 mRNA level in oocytes between SOV and control groups were observed. Conclusion Dynein inhibitor might delay oocytes meiosis process, and cause ectopic expression of cyclin B1 in oocytes. Most Oocytes incubated with SOV blocked at germinal vesicles (GV) stage or MⅠto anaphase transition due to dynein dysfunction and ectopic transcription level of cyclin B1.
