| 引用本文: |
毛昀, 丁致薰, 付玲, 胡贝尔, 朱世杰, 曹文.不同补肾类中药方含药血清抑制乳腺癌细胞MDA-MB-231间质-上皮转化的机制研究比较[J].湖南中医药大学学报,2025,45(8):1434-1442[点击复制] |
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| 不同补肾类中药方含药血清抑制乳腺癌细胞MDA-MB-231间质-上皮转化的机制研究比较 |
| 毛昀,丁致薰,付玲,胡贝尔,朱世杰,曹文 |
| (湖南中医药大学第二附属医院肿瘤血液科, 湖南 长沙 410005;中国中医科学院望京医院肿瘤科, 北京 100102) |
| 摘要: |
| 目的 探讨补肾阳方、补肾阴方、阴阳双补方含药血清干预乳腺癌细胞间质-上皮转化的疗效差异与分子机制。方法 建立乳腺癌细胞MDA-MB-231和成骨前体细胞MC3T3-E1共培养模型，分组为对照组、模型组、补肾阳方组、补肾阴方组、阴阳双补方组，其中对照组为乳腺癌细胞单培养，其他组为乳腺癌细胞与成骨前体细胞共培养模型。对照组和模型组培养液中加入10%普通血清，补肾阳方组、补肾阴方组、阴阳双补方组培养液中加入10%相应含药血清。给药干预24、48、72 h后，采用CCK-8法检测乳腺癌细胞活力；结晶紫染色观察细胞形态。共培养48 h后，细胞划痕实验检测乳腺癌细胞迁移能力，流式细胞术检测乳腺癌细胞周期和凋亡；培养48 h后qRT-PCR和Western blot检测E-钙黏蛋白（E-cad）、N-钙黏蛋白（N-cad）、缝隙连接蛋白43（Cx43）和dickkopf相关蛋白1（Dkk-1） mRNA相对表达水平和蛋白表达量。结果 与对照组比较，模型组乳腺癌细胞活力增加（P<0.05）、细胞总凋亡率下降（P<0.05），细胞周期检测示G2/M期细胞增多（P<0.05）,G0/G1期、S期细胞减少（P<0.05）;E-cad、N-cad、Cx43和Dkk-1 mRNA相对表达水平和蛋白表达量均上升（P<0.05）。与模型组比较，补肾阳方组、补肾阴方组、阴阳双补方组细胞活力、迁移愈合率降低（P<0.05），细胞总凋亡率增加（P<0.05），细胞周期检测示G0/G1期细胞增多（P<0.05）,G2/M期细胞减少（P<0.05）;E-cad、N-cad、Cx43和Dkk-1 mRNA相对表达水平和蛋白表达量下降（P<0.05）。结论 成骨前体细胞MC3T3-E1通过间质-上皮转化促进乳腺癌细胞MDA-MB-231存活；采用补肾类中药干预共培养模型后，可抑制乳腺癌细胞活力、促进凋亡，机制与抑制间质-上皮转化进程，下调关键蛋白mRNA（E-cad、N-cad、Cx43和Dkk-1）的表达相关。 |
| 关键词: 乳腺癌 骨转移 MDA-MB-231细胞 MC3T3-E1细胞 间质-上皮转化 补肾类中药 |
| DOI：10.3969/j.issn.1674-070X.2025.08.006 |
| 投稿时间：2025-06-03 |
| 基金项目:国家自然科学基金面上项目(81973640);国家自然科学基金青年项目(82405165);湖南省自然科学基金青年项目(2023JJ40497);湖南省教育厅优秀青年项目(22B0356);长沙市自然科学基金项目(kq2403109);湖南中医药大学校级重点课题(2022XYLH020) |
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| Comparative study on the mechanisms of inhibiting mesenchymal-epithelial transition in breast cancer MDA-MB-231 cells by drug-containing sera from different kidney-tonifying Chinese medicine formulas |
| MAO Yun, DING Zhixun, FU Ling, HU Bei'er, ZHU Shijie, CAO Wen |
| (Department of Oncology and Hematology, The Second Hospital of Hunan University of Chinese Medicine, Changsha, Hunan 410005, China;Oncology Department, Wang Jing Hospital of China Academy of Chinese Medical Sciences, Beijing 100029, China) |
| Abstract: |
| Objective To explore the efficacy differences and molecular mechanisms of drug-containing sera from Bushenyang Formula（BSYAF）, Bushenyin Formula（BSYIF）, and Yin-yang Shuangbu Formula（YYSBF） in intervening mesenchymalepithelial transition（MET） in breast cancer cells. Methods A co-culture model of breast cancer cells MDA-MB-231 and osteoblast precursor cells MC3T3-E1 was established. Groups included the control group, model group, BSYAF group, BSYIF group, and YYSBF group. In the control group, breast cancer cells were cultured alone, while the other groups adopted the co-culture model.The culture medium of the control group and the model group was supplemented with 10% normal serum, while that of the BSYAF,BSYIF, and YYSBF groups was supplemented with 10% corresponding drug-containing sera. After 24, 48, and 72 hours of drug intervention, the viability of breast cancer cells was determined using the CCK-8 assay, and cell morphology was observed through crystal violet staining. After 48 hours of co-culture, the migration ability of breast cancer cells was assessed using a wound healing assay, and the cell cycle and apoptosis of breast cancer cells were tested by flow cytometry. After 48 hours of culture, the relative m RNA expression levels and protein expression amounts of E-cadherin（E-cad）, N-cadherin（N-cad）, connexin 43（Cx43）, and dickkopfrelated protein 1（Dkk-1） were measured by qRT-PCR and Western blot respectively. Results Compared with the control group, the breast cancer cell viability in the model group increased（P＜0.05）, the total cell apoptosis rate decreased（P＜0.05）. Cell cycle analysis showed an increase in the number of cells in the G2/M phase（P＜0.05） and a decrease in the number of cells in the G0/G1and S phases（P＜0.05）. The relative mRNA expression levels and protein expression amounts of E-cad, N-cad, Cx43, and Dkk-1all increased（P＜0.05）. Compared with the model group, the cell viability and migration healing rate in the BSYAF, BSYIF, and YYSBF groups decreased（P＜0.05）, while the total cell apoptosis rate increased（P＜0.05）. Cell cycle analysis revealed an increase in the number of cells in the G0/G1phase（P＜0.05） and a decrease in the G2/M phase（P＜0.05）. The relative mRNA expression levels and protein expression amounts of E-cad, N-cad, Cx43, and Dkk-1 decreased（P＜0.05）. Conclusion Osteoblast precursor cells MC3T3-E1promote the survival of breast cancer cells MDA-MB-231 through mesenchymal-epithelial transition. After intervention with kidneytonifying Chinese medicines in the co-culture model, the viability of breast cancer cells can be effectively inhibited and apoptosis can be promoted. The mechanism is related to the downregulation of the expression of key proteins and mRNAs（E-cad, Cx43, and Dkk-1） involved in mesenchymal-epithelial transition. |
| Key words: breast cancer bone metastasis MDA-MB-231 cells MC3T3-E1 cells mesenchymal-epithelial transition kidney-tonifying Chinese medicines |
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