黄酮通过激活 AMPK/PGC-1α 通路抑制喉癌细胞线粒体代谢重塑及增殖的机制研究

Flavonoids Inhibiting Mitochondrial Metabolic Remodeling and Proliferation of Laryngeal Cancer Cells by Activating the AMPK/PGC-1α Pathway

摘要:

目的 探讨黄酮类化合物对喉癌细胞增殖的影响,并深入研究其是否通过调控 AMPK/PGC-1α 信号通路干预线粒体代谢重塑,进而发挥抗肿瘤作用。方法 以人喉癌 Hep-2 细胞系为研究对象并采用木犀草素为黄酮代表性物质进行梯度浓度处理。CCK-8和克隆形成实验检测细胞增殖能力；流式细胞术测定线粒体聚集体的变化；Seahorse 能量代谢分析仪检测细胞耗氧率(OCR)与细胞外酸化率(ECAR)；Western blot 技术检测 AMPK 磷酸化(p-AMPK)、PGC-1α、线粒体生物合成相关蛋白(TFAM、NRF1)及代谢关键蛋白(SDHA)的表达水平。结果 黄酮可浓度依赖性地抑制 Hep-2 细胞增殖与克隆形成能力,并抑制细胞线粒体膜电位的发生。此外,Hep-2细胞的基础呼吸和基础糖酵解能力同样受到抑制。机制层面,黄酮能显著促进 AMPK 磷酸化,上调下游转录共激活因子 PGC-1α 及其靶基因 TFAM、NRF1 的表达。回复实验显示,Compound C 抑制 AMPK 活性或 siRNA 敲低 PGC-1α 表达后,黄酮对线粒体功能的抑制作用及抗增殖效应均被显著削弱。结论 黄酮可有效抑制喉癌细胞增殖,其作用机制与激活 AMPK/PGC-1α 信号通路相关。该通路的活化虽能上调线粒体生物合成相关蛋白表达,但最终会引发线粒体功能紊乱,抑制细胞代谢重塑,本研究为黄酮类化合物作为喉癌潜在辅助治疗剂提供了新的理论依据。

Abstract:

Objective: To investigate the effect of flavonoids on the proliferation of laryngeal cancer cells and to explore whether they exert anti-tumor effects by regulating the AMPK/PGC-1α signaling pathway and interfering with mitochondrial metabolic remodeling. Methods: The human laryngeal cancer cell line Hep-2 was used as the research object, and luteolin was selected as the representative flavonoid for gradient concentration treatment. CCK-8 and colony formation assays were used to detect cell proliferation ability; flow cytometry was used to measure the changes in mitochondrial aggregates; Seahorse energy metabolism analyzer was used to detect the oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) of cells; Western blot was used to detect the expression levels of AMPK phosphorylation (p-AMPK), PGC-1α, mitochondrial biogenesis-related proteins (TFAM, NRF1), and metabolic key proteins (SDHA). Results: Flavonoids could inhibit the proliferation and colony formation ability of Hep-2 cells in a concentration-dependent manner, and also inhibit the occurrence of mitochondrial membrane potential. In addition, the basal respiration and basal glycolysis ability of Hep-2 cells were also inhibited. At the mechanism level, flavonoids could significantly promote AMPK phosphorylation and up-regulate the expression of downstream transcriptional co-activator PGC-1α and its target genes TFAM and NRF1. Rescue experiments showed that the inhibitory effect of flavonoids on mitochondrial function and anti-proliferative effect were significantly weakened after Compound C inhibited AMPK activity or siRNA knocked down PGC-1α expression. Conclusion: Flavonoids can effectively inhibit the proliferation of laryngeal cancer cells, and their mechanism of action is related to the activation of the AMPK/PGC-1α signaling pathway. Although the activation of this pathway can up-regulate the expression of mitochondrial biogenesis-related proteins, it ultimately leads to mitochondrial dysfunction and inhibits cell metabolic remodeling. This study provides a new theoretical basis for flavonoids as potential adjuvant therapeutic agents for laryngeal cancer.