| 摘要: |
| ［摘要］ 目的 探讨抗肿瘤天然产物Garcinone E对卵巢癌细胞自噬功能的影响及作用机制。方法 选择卵巢癌细胞HeyA8和SKOV3进行实验。以不同浓度Garcinone E对卵巢癌细胞进行干预，采用CCK-8实验检测细胞活力，通过单丹磺酰尸胺（MDC）染色实验检测细胞自噬小体形成情况。构建稳定表达stubRFP-sensGFP-LC3慢病毒的HeyA8细胞和SKOV3细胞，检测细胞自噬流变化。通过溶酶体染色实验检测卵巢癌细胞溶酶体与sensGFP-LC3共定位情况。应用Western blot实验检测卵巢癌细胞细胞周期相关蛋白p21、周期蛋白依赖性激酶4（CDK4）、周期蛋白依赖性激酶7（CDK7）、蛋白激酶B（Akt）、微管相关蛋白1-轻链3B（LC3B）和自噬相关蛋白p62的表达水平。结果 经Garcinone E干预后，卵巢癌细胞活力显著下降（P<0.05），其对HeyA8细胞、SKOV3细胞的半数抑制量(IC50)分别为5.93 μmol/L、8.14 μmol/L。MDC染色实验发现，经Garcinone E干预后，卵巢癌细胞内自噬小体数量增加。荧光显微镜下发现Garcinone E阻断稳定转染stubRFP-sensGFP-LC3慢病毒的HeyA8细胞和SKOV3细胞的自噬流。溶酶体染色实验结果显示Garcinone E阻断了自噬小体与溶酶体的结合。Western blot实验结果显示，Garcinone E干预可上调卵巢癌细胞LC3B、p62和p21的蛋白表达水平（P<0.05），下调Akt、CDK4和CDK7的蛋白表达水平（P<0.05）。结论 Garcinone E能够抑制卵巢癌细胞的自噬流，阻滞细胞周期，具有抗卵巢癌的作用，有作为卵巢癌化疗药物研发先导药物的潜力。 |
| 关键词: Garcinone E 卵巢癌 自噬流 细胞周期 |
| DOI：10.3969/j.issn.1674-3806.2025.05.09 |
| 分类号: |
| 基金项目:国家自然科学基金项目（编号：82260721，81903644）；广西自然科学基金重点项目（编号：2024GXNSFDA010045） |
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| A study on mechanism of anti-tumor natural product Garcinone E inhibiting autophagic flux in ovarian cancer cells |
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SUN Xuemei1, LEI Shunmei1, WANG Chunlai1, DAI Yiping1, WEI Dan1, YIN Fuqiang2, LIU Xia1
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1.School of Basic Medicine, Key Laboratory of Longevity and Aging-Related Diseases of Chinese Ministry of Education, Guangxi Medical University, Nanning 530021, China; 2.Life Sciences Institute, Guangxi Medical University, Nanning 530021, China
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| Abstract: |
| ［Abstract］ Objective To explore the effect of anti-tumor natural product Garcinone E on autophagy function of ovarian cancer cells and its mechanism. Methods Ovarian cancer cells HeyA8 and SKOV3 were selected for experiments. Ovarian cancer cells were intervened with different concentrations of Garcinone E. Cell viability was detected by CCK-8 assay, and autophagic vesicle formation was detected by monodansylcadaverine(MDC) staining assay. The HeyA8 cell line and SKOV3 cell line stably expressing stubRFP-sensGFP-LC3 lentivirus were constructed to detect the changes in cellular autophagic flux. Co-localization of lysosomes with sensGFP-LC3 in ovarian cancer cells was detected by using lysosomal staining assay. Western blot assay was applied to detect the expression levels of cell cycle-related proteins p21, cyclin-dependent kinase 4(CDK4), cyclin-dependent kinase 7(CDK7), protein kinase B(Akt), microtubule-associated protein 1-light chain 3B(LC3B) and autophagy-related protein p62 in ovarian cancer cells. Results The viability of ovarian cancer cells was significantly decreased(P<0. 05) after intervention with Garcinone E. The half maximal inhibitory concentration(IC50) of Garcinone E on HeyA8 cells and SKOV3 cells were 5.93 μmol/L and 8.14 μmol/L, respectively. The MDC staining assay revealed an increase in the number of autophagosomes in the ovarian cancer cells after intervention with Garcinone E. Fluorescence microscopy revealed that Garcinone E blocked the autophagic flux in HeyA8 cells and SKOV3 cells stably transfected with stubRFP-sensGFP-LC3 lentivirus. The results of lysosomal staining assay showed that Garcinone E blocked the binding of autophagosomes to lysosomes. The results of Western blot assay showed that the intervention with Garcinone E up-regulated the protein expression levels of LC3B, p62 and p21 in ovarian cancer cells(P<0. 05) and down-regulated the protein expression levels of Akt, CDK4 and CDK7(P<0. 05). Conclusion Garcinone E can inhibit the autophagic flux of ovarian cancer cells and block the cell cycle of ovarian cancer cells, which has an anti-ovarian cancer effect and the potential to serve as a lead compound in the development of chemotherapeutic drugs for ovarian cancer. |
| Key words: Garcinone E Ovarian cancer Autophagic flux Cell cycle |
