基于Keap1/Nrf2/HO-1信号通路探讨毛兰素对前列腺癌细胞铁死亡的影响

赵海英; 刘建荣; 向阳

基于Keap1/Nrf2/HO-1信号通路探讨毛兰素对前列腺癌细胞铁死亡的影响

1.
中国人民解放军联勤保障部队第 961医院泌尿外科，黑龙江齐齐哈尔　161000
2.
齐齐哈尔市第一医院泌尿外科，黑龙江齐齐哈尔　161000

基金项目: 黑龙江省卫生健康委员会科研项目（H186150198）

详细信息

作者简介:
赵海英（1978～），男，黑龙江齐齐哈尔人，医学学士，主治医师，主要从事前列腺癌治疗工作

通讯作者:
赵海英，E-mail：dl08888483@163.com

中图分类号: R737.25
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- 被引次数: 0
出版历程
- 收稿日期: 2025-07-02
- 网络出版日期: 2026-03-29

Effects of Erianin on Ferroptosis in Prostate Cancer Cells via the Keap1/Nrf2/HO-1 Signaling Pathway

1.
Department of Urology，The 961st Hospital of the Joint Service Support Force of the People’s Liberation Army of China，Qiqihar Heilongjiang 161000
2.
Department of Urology，Qiqihar First Hospital，Qiqihar Heilongjiang 161000，China

摘要

摘要: 目的探讨毛兰素是否通过调控Keap1/Nrf2/HO-1信号通路，进而影响前列腺癌细胞的铁死亡进程，以阐明其潜在的分子机制。方法培养DU145和PC-3前列腺癌细胞系。CCK8、集落形成测定、细胞侵袭实验和流式细胞术检测细胞活力、迁移、侵袭和凋亡。使用ROS特异性探针DCFH-DA分析细胞内ROS含量，检测丙二醛（malondialdehyde，MDA）和谷胱甘肽（glutathione，GSH）含量，蛋白质印迹法测量细胞中转移性表型标志物的表达（vimentin、N-cadherin、slug、snail和MMP-9）表达、铁死亡阴性调控蛋白（GPX4、CHAC2、SLC40A1、SLC7A11和glutaminase）的表达和Keap1/Nrf2/HO-1信号通路表达。结果与对照组比较，毛兰素显著降低前列腺癌细胞（DU145、PC-3）活力，抑制其增殖、迁移及侵袭能力（P < 0.05）；毛兰素可下调间充质标志物vimentin、N-cadherin、slug、snail和MMP-9并上调上皮标志物E-cadherin来抑制前列腺癌细胞的迁移，ROS积累、谷胱甘肽消耗和脂质过氧化显著增加，毛兰素诱导的前列腺癌细胞死亡可以通过与ROS抑制剂n-乙酰-l-半胱氨酸（NAC）和GSH共处理来挽救。毛兰素处理后，Phen Green SK（检测细胞内游离 Fe²⁺）阳性细胞比例显著下降（P < 0.05）。透射电子显微镜（transmission electron microscope，TEM）结果发现，经毛兰素处理的前列腺癌细胞出现线粒体基质凝结和嵴增大。毛兰素处理后，铁死亡阴性调控蛋白GPX4、CHAC2、SLC40A1、SLC7A11和glutaminase的表达明显降低，与铁死亡抑制剂利蒲他汀-1 （Lip-1）或铁他汀-1 （Fer-1）联合治疗，可以阻断毛兰素诱导的前列腺癌细胞死亡，毛兰素提高了Keap1蛋白表达，而细胞中的Nrf2，HO-1和NQO1蛋白表达降低。CPUY192018可提高前列腺癌细胞铁死亡，表明毛兰素靶向Keap1/Nrf2/HO-1信号通路诱导细胞铁死亡。结论毛兰素靶向Keap1/Nrf2/HO-1信号通路诱导细胞死亡和抑制前列腺癌细胞转移。
- 毛兰素 /
- Keap1/Nrf2/HO-1 /
- 前列腺癌 /
- 铁死亡
Abstract: Purpose To investigate whether erianin induces ferroptosis in prostate cancer cells by regulating the Keap1/Nrf2/HO-1 signaling pathway, and to elucidate its potential molecular mechanisms. Methods Human prostate cancer cell lines DU145 and PC-3 were cultured. Cell viability was detected by CCK-8 assay, cell proliferation by colony formation assay, cell invasion by Transwell assay, and cell apoptosis by flow cytometry. Intracellular reactive oxygen species （ROS） content was analyzed using the ROS-specific probe DCFH-DA, and the levels of malondialdehyde （MDA） and glutathione （GSH） were determined. Western blot was used to detect the expression of metastatic phenotypic markers （vimentin, N-cadherin, slug, snail, and MMP-9）, ferroptosis-negative regulatory proteins （GPX4, CHAC2, SLC40A1, SLC7A11, and glutaminase）, and proteins related to the Keap1/Nrf2/HO-1 signaling pathway. Results Compared with the control group, erianin significantly reduced the viability of prostate cancer cells （DU145, PC-3） and inhibited their proliferation, migration, and invasion abilities （P < 0.05）. Erianin inhibited the migration of prostate cancer cells by downregulating the mesenchymal markers （vimentin, N-cadherin, slug, snail, and MMP-9） and upregulating the epithelial marker E-cadherin. ROS accumulation, GSH depletion, and lipid peroxidation were significantly increased. Erianin-induced prostate cancer cell death could be rescued by co-treatment with the ROS inhibitor N-acetyl-L-cysteine （NAC） and glutathione （GSH）. After erianin treatment, the proportion of Phen Green SK-positive cells （detecting intracellular free Fe²⁺） was significantly decreased （P < 0.05）. Transmission electron microscopy （TEM） revealed mitochondrial matrix condensation and cristae enlargement in erianin-treated prostate cancer cells. Additionally, erianin treatment significantly downregulated the expression of ferroptosis-negative regulatory proteins （GPX4, CHAC2, SLC40A1, SLC7A11, and glutaminase）. Co-treatment with ferroptosis inhibitors （Liproxstatin-1, Lip-1; or Ferrostatin-1, Fer-1） blocked erianin-induced prostate cancer cell death. Erianin increased Keap1 protein expression while decreasing the expression of Nrf2, HO-1, and NQO1 proteins in cells. CPUY192018 enhanced ferroptosis in prostate cancer cells, indicating that erianin induces ferroptosis through targeting the Keap1/Nrf2/HO-1 signaling pathway. Conclusion: Erianin induces cell death and suppresses prostate cancer cell migration by targeting the Keap1/Nrf2/HO-1 signaling pathway.
- Erianin /
- Keap1/Nrf2/HO-1 Signaling pathway /
- Prostate cancer /
- Ferroptosis

HTML全文

图 1 毛兰素对前列腺癌细胞生长的影响

A～B：CCK8检测细胞活力；C：有代表性的细胞形态变化；D～E：流式细胞术检测；与对照组比较，^*P < 0.05，^**P < 0.01。

Figure 1. Efects of erianin on prostate cancer cell growth

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图 2 毛兰素对前列腺癌细胞迁移的影响

A：伤口愈合测定；B～C：Transwell测定细胞迁移；D：蛋白印迹检测转移性表型标志物的表达；与对照组比较，^*P < 0.05；^**P < 0.01。

Figure 2. Effect of erianin on prostate cancer cell migration

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3 毛兰素对前列腺癌细胞铁死亡的影响

A～B：流式细胞仪检测细胞ROS水平；C：细胞内GSH水平；D：细胞内MDA水平；E～F：ROS抑制剂NAC和GSH处理后细胞活力；G：用荧光指示剂Phen Green SK（绿色）测定细胞内的螯合铁；H：透射电镜观察细胞的铁死亡；I：蛋白印迹检测铁死亡标志物的表达；J：铁死亡抑制剂Lip-1或Fer-1的细胞抑制率；与对照组比较，^*P < 0.05；^**P < 0.01。

3. Effect of erianin on ferroptosis in prostate cancer cells

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3 毛兰素对前列腺癌细胞铁死亡的影响

A～B：流式细胞仪检测细胞ROS水平；C：细胞内GSH水平；D：细胞内MDA水平；E～F：ROS抑制剂NAC和GSH处理后细胞活力；G：用荧光指示剂Phen Green SK（绿色）测定细胞内的螯合铁；H：透射电镜观察细胞的铁死亡；I：蛋白印迹检测铁死亡标志物的表达；J：铁死亡抑制剂Lip-1或Fer-1的细胞抑制率；与对照组比较，^*P < 0.05；^**P < 0.01。

3. Effect of erianin on ferroptosis in prostate cancer cells

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图 4 毛兰素对前列腺癌细胞Keap1/Nrf2/HO-1信号通路的影响

A：蛋白印迹检测PC3细胞中Keap1/Nrf2/HO-1信号通路；B：蛋白印迹定量分析PC3细胞信号通路表达；C：蛋白印迹检测DU145细胞中Keap1/Nrf2/HO-1信号通路；D：蛋白印迹定量分析DU145 细胞信号通路表达；与对照组比较，^*P < 0.05；^**P < 0.01。

Figure 4. Effect of erianin on the Keap1/Nrf2/HO-1 signaling pathway in prostate cancer cells

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图 5 抑制Keap1/Nrf2/HO-1信号通路对前列腺癌细胞铁死亡的影响

A～B：检测细胞活力；C：用荧光指示剂Phen Green SK （绿色）测定细胞内的螯合铁；D：蛋白印迹检测铁死亡标志物的表达；与对照组比较，^*P < 0.05；^**P < 0.01。

Figure 5. Effects of inhibiting the Keap1/Nrf2/HO-1 signaling pathway on ferroptosis in prostate cancer cells

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