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Objective <p>This study aimed to examine the preventive impact of Liuwei Dihuang pill (LWDH) on ovarian injury and to elucidate its mechanism through in vivo and in vitro tests.</p> Methods <p>Cyclophosphamide (CTX) was injected intraperitoneally and LWDH suspension was administered intragastrically to create a mouse model of primary ovarian insufficiency. Serum concentrations of hormones including AMH, E2 and FSH, alongside oxidative stress-related biomarkers such as SOD, GSH-Px and MDA, were measured using ELISA. By counting the autophagosomes in granulosa cells using electron microscopy and LC3 immunofluorescence labeling, the degree of autophagy was assessed. Finally, detect the expression levels of genes and proteins related to autophagy.</p> Results <p>LWDH elevated the ovarian weight and improved the ovarian index in mice with ovarian injury. The LWDH-treated group exhibited considerably lower MDA levels and significantly higher SOD and GSH-Px levels than the CTX group. Meanwhile, the group treated with LWDH reduced the granulosa cells' excessive apoptosis. In addition, the LWDH-treated group exhibited reduced LC3 fluorescence intensity and fewer autophagosome counts in comparison to the CTX group. Similarly, the expression of LC3, an autophagy-related marker, was decreased and p62 was markedly elevated in comparison to the CTX group. These findings suggest that LWDH has a positive protective effect on ovarian injury mice by increasing antioxidant enzyme activity, improving ovarian tissue pathological damage, inhibiting granulosa cell apoptosis, and balancing granulosa cell autophagy. In addition, the expression levels of SIRT1 and PI3K/AKT signaling pathway related proteins in the LWDH treatment group were significantly higher than those in the CTX group, suggesting that LWDH may exert its protective effect by activating the SIRT1/PI3K/AKT signaling pathway.</p> Conclusion <p>LWDH may suppress granulosa cell autophagy through the activation of the SIRT1/PI3K/AKT signaling pathway, hence enhancing ovarian function.</p> Graphical Abstract <p></p>

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Mechanism study on the regulation of autophagy and oxidative stress through SIRT1/PI3K/AKT pathway by Liuwei Dihuang pill to improve ovarian function

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年6月2日

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  • Jiawen Zhong,
  • Guoxia Lan,
  • Xiaoqing Ma,
  • Wenyan Zhang,
  • Bo Jiang,
  • Ling Qin,
  • Xiaorong Li,
  • Yan Nian

摘要

Objective

This study aimed to examine the preventive impact of Liuwei Dihuang pill (LWDH) on ovarian injury and to elucidate its mechanism through in vivo and in vitro tests.

Methods

Cyclophosphamide (CTX) was injected intraperitoneally and LWDH suspension was administered intragastrically to create a mouse model of primary ovarian insufficiency. Serum concentrations of hormones including AMH, E2 and FSH, alongside oxidative stress-related biomarkers such as SOD, GSH-Px and MDA, were measured using ELISA. By counting the autophagosomes in granulosa cells using electron microscopy and LC3 immunofluorescence labeling, the degree of autophagy was assessed. Finally, detect the expression levels of genes and proteins related to autophagy.

Results

LWDH elevated the ovarian weight and improved the ovarian index in mice with ovarian injury. The LWDH-treated group exhibited considerably lower MDA levels and significantly higher SOD and GSH-Px levels than the CTX group. Meanwhile, the group treated with LWDH reduced the granulosa cells' excessive apoptosis. In addition, the LWDH-treated group exhibited reduced LC3 fluorescence intensity and fewer autophagosome counts in comparison to the CTX group. Similarly, the expression of LC3, an autophagy-related marker, was decreased and p62 was markedly elevated in comparison to the CTX group. These findings suggest that LWDH has a positive protective effect on ovarian injury mice by increasing antioxidant enzyme activity, improving ovarian tissue pathological damage, inhibiting granulosa cell apoptosis, and balancing granulosa cell autophagy. In addition, the expression levels of SIRT1 and PI3K/AKT signaling pathway related proteins in the LWDH treatment group were significantly higher than those in the CTX group, suggesting that LWDH may exert its protective effect by activating the SIRT1/PI3K/AKT signaling pathway.

Conclusion

LWDH may suppress granulosa cell autophagy through the activation of the SIRT1/PI3K/AKT signaling pathway, hence enhancing ovarian function.

Graphical Abstract