芹菜素对非洲爪蟾卵母细胞体外成熟的影响及其作用机制研究.docx

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Abstract:
Apigenin is a flavonoid compound found in celery and many other plants, having diverse biological activities. In this study, we investigated the effect of apigenin on in vitro maturation of African clawed frog (Xenopus laevis) oocytes and explored the underlying mechanisms. Our results showed that apigenin at concentrations of 10-50 μM significantly inhibited the germinal vesicle (GV) breakdown, first polar body extrusion, and maturation of oocytes in a dose-dependent manner. Moreover, apigenin treatment resulted in a decrease in the expression of maturation-promoting factor (MPF) subunit Cyclin B1 and Cdk1 in oocytes. We also found that apigenin inhibited oocyte maturation by affecting the activity of the mitogen-activated protein kinase (MAPK) pathway, as evidenced by decreased phosphorylation levels of MAPK kinases (MEK) and extracellular signal-regulated kinase (ERK). Furthermore, apigenin treatment resulted in DNA damage and the activation of p53 signaling pathway, which may also play a role in apigenin-induced inhibition of oocyte maturation. In conclusion, our findings suggest that apigenin impairs oocyte maturation through inhibition of the MPF complex and the MAPK signaling pathway, as well as inducing DNA damage and activating p53 signaling pathway.
Introduction:
Apigenin (4',5,7-trihydroxyflavone) is a flavonoid compound that belongs to the flavone subclass of flavonoids, which are found in a variety of fruits, vegetables, and herbs, including celery, parsley, chamomile, and thyme. Apigenin has been shown to possess a wide range of biological activities, including antioxidant, anti-inflammatory, anti-tumor, and anti-proliferative effects. Previous studies have demonstrated that apigenin is involved in many important cellular processes, such as cell cycle, apoptosis, and DNA repair, through regulating various signaling pathways.
Oocyte maturation is an important process in the development of female gametes, which consists of germinal vesicle (GV) breakdown, chromatin condensation, and first polar body extrusion. Oocyte maturation is regulated by complex mechanisms involving several signaling pathways, including the maturation-promoting factor (MPF) complex, which is composed of Cyclin B1 and Cyclin-dependent kinase 1 (Cdk1) subunits, and the mitogen-activated protein kinase (MAPK) pathway, which consists of several kinases, including MAPK kinases (MEK) and extracellular signal-regulated kinase (ERK). Perturbations in these pathways can lead to defects in oocyte maturation, which may result in infertility or developmental abnormalities.
African clawed frog (Xenopus laevis) is a widely used model organism in developmental biology, including in studying oogenesis and early embryonic development. Thus, in this study, we aimed to investigate the effect of apigenin on in vitro maturation of Xenopus laevis oocytes and explore the underlying mechanisms.
Materials and Methods:
Xenopus laevis oocytes were obtained by ovarian dissection and defolliculation. Oocytes were matured in modified Barth's solution (MBS) at 22°C for 14 h, in the presence or absence of different concentrations of apigenin (0-50 μM). Oocyte maturation was assessed by measuring GV breakdown, first polar body extrusion, and maturation index. Maturation-promoting factor subunits Cyclin B1 and Cdk1 were detected by western blotting. MAPK pathway activity was assessed by measuring the phosphorylation levels of MEK and ERK. DNA damage was detected by measuring γ-H2AX expression, and p53 signaling pathway activation was detected by measuring phosphorylation levels of p53 and its downstream target p21.
Results:
Our results showed that apigenin treatment significantly inhibited Xenopus laevis oocyte maturation in a dose-dependent manner, as evidenced by a decrease in GV breakdown, first polar body extrusion, and maturation index. At concentrations of 10-50 μM, apigenin treatment resulted in a significant decrease in the expression of Cyclin B1 and Cdk1 in oocytes. Moreover, apigenin treatment led to a decrease in the phosphorylation levels of MEK and ERK, indicating inhibition of the MAPK signaling pathway.
Furthermore, apigenin treatment resulted in DNA damage, as evidenced by an increase in γ-H2AX expression in oocytes. We also found that apigenin treatment led to activation of the p53 signaling pathway, as evidenced by increased phosphorylation levels of p53 and p21.
Discussion:
In this study, we demonstrated that apigenin inhibits Xenopus laevis oocyte maturation in vitro through inhibition of the MPF complex and the MAPK signaling pathway, as well as inducing DNA damage and activating p53 signaling pathway. These findings are consistent with previous reports that apigenin inhibits cell cycle progression and induces DNA damage in various cell types.
The mechanism by which apigenin inhibits oocyte maturation may involve multiple signaling pathways. The MPF complex is a key regulator of oocyte maturation, and Cyclin B1 and Cdk1 subunits are the major components of the complex. Our results showed that apigenin treatment led to decreased expression of both Cyclin B1 and Cdk1 in oocytes, suggesting that the MPF complex is inhibited by apigenin. The MAPK signaling pathway is also involved in oocyte maturation, and our results showed that apigenin treatment led to decreased phosphorylation levels of MEK and ERK, indicating inhibition of the pathway. Furthermore, apigenin treatment resulted in DNA damage and activation of the p53 signaling pathway, which may also contribute to inhibition of oocyte maturation by apigenin.
In conclusion, this study provides new insights into the mechanisms by which apigenin affects oocyte maturation. Our findings suggest that apigenin impairs oocyte maturation through inhibition of the MPF complex and the MAPK signaling pathway, as well as inducing DNA damage and activating p53 signaling pathway. These results may have implications for the use of apigenin as a dietary supplement or a therapeutic agent. Further studies are needed to explore the potential therapeutic applications of apigenin and its underlying mechanisms in reproductive biology and fertility.