PO.MCB11.01 · 分子与细胞生物学
DDX3调控线粒体功能以抑制HCC进展
DDX3 modulates mitochondrial function to inhibit HCC progression
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
背景：线粒体是负责产生ATP的重要细胞器，在多种细胞过程中发挥关键作用。线粒体功能障碍与包括癌症发生在内的多种病理状况有关。作为营养代谢的中枢器官，肝脏含有特别高密度的线粒体。线粒体功能受损导致活性氧（ROS）过度产生和氧化应激，造成与肝细胞癌（HCC）密切相关的细胞损伤。癌细胞常发生代谢重编程以维持其快速增殖期间的高能量需求。鉴于我们既往发现DDX3是HCC中的一个肿瘤抑制因子，本研究旨在阐明DDX3对线粒体功能和氧化还原调控的影响，以及其对HCC肿瘤发生的意义。
方法：使用超分辨共聚焦显微镜分析DDX3失调HCC细胞中的线粒体形态。通过免疫印迹和定量实时PCR评估线粒体动力学相关蛋白的表达。通过MitoTracker Red/Green染色和Seahorse胞外通量分析检测代谢活性。通过MitoSOX和DCFH-DA染色，以及GSH/GSSG比值和NADPH/NADP⁺水平的测定评估氧化还原稳态。通过增殖、集落形成和transwell迁移/侵袭实验（在有或无代谢抑制剂的情况下）研究DDX3调控的细胞功能。使用成球、化疗耐药和基于流式细胞术的实验评估癌症干性。
结果：HCC细胞中DDX3表达降低导致线粒体形态显著改变和线粒体氧化磷酸化（OXPHOS）增加，从而导致线粒体超氧化物积累升高。出乎意料的是，DDX3敲低细胞中的细胞总ROS水平降低。进一步分析显示，低DDX3表达增强了细胞维持氧化还原平衡的能力，赋予更强大的抗氧化防御。此外，DDX3表达降低的HCC细胞在不同营养条件下表现出更强的增殖和自我更新潜能，而这些效应在代谢抑制剂处理后减弱。
结论：我们的结果提示DDX3可能通过抑制线粒体活性来抑制HCC进展，从而强化其作为HCC肿瘤抑制因子的作用。
查看英文原文 English abstract
Background: Mitochondria are essential organelles responsible for ATP generation and play pivotal roles in diverse cellular processes. Mitochondrial dysfunction contributes to various pathological conditions, including cancer development. As the central organ for nutrient metabolism, the liver contains a particularly high density of mitochondria. Impaired mitochondrial function leads to excessive generation of reactive oxygen species (ROS) and oxidative stress, resulting in cellular damage closely associated with hepatocellular carcinoma (HCC). Cancer cells frequently undergo metabolic reprogramming to sustain their high energy demands during rapid proliferation. Given our previous findings identifying DDX3 as a tumor suppressor in HCC, this study aims to elucidate the effects of DDX3 on mitochondrial function and redox regulation, as well as its implications for HCC tumorigenesis.
Methods: Mitochondrial morphology in DDX3-deregulated HCC cells was analyzed using super-resolution confocal microscopy. The expression of mitochondrial dynamics-related proteins was assessed by immunoblotting and quantitative real-time PCR. Metabolic activity was examined by MitoTracker Red/Green staining and Seahorse extracellular flux analysis. Redox homeostasis was evaluated by MitoSOX and DCFH-DA staining, along with measurements of the GSH/GSSG ratio and NADPH/NADP⁺ levels. DDX3-regulated cellular functions were investigated through proliferation, colony formation, and transwell migration/invasion assays, with or without metabolic inhibitors. Cancer stemness was assessed using sphere-formation, chemoresistance, and flow cytometry-based assays.
Results: Reduced DDX3 expression in HCC cells led to marked alterations in mitochondrial morphology and increased mitochondrial oxidative phosphorylation (OXPHOS), resulting in elevated mitochondrial superoxide accumulation. Unexpectedly, the total cellular ROS levels were decreased in DDX3-knockdown cells. Further analyses revealed that low DDX3 expression enhances the cellular capacity to maintain redox balance, conferring a more robust antioxidant defense. Additionally, HCC cells with reduced DDX3 expression exhibited greater proliferation and self-renewal potential under varying nutrient conditions, effects that were attenuated upon treatment with metabolic inhibitors.
Conclusion: Our results suggested that DDX3 may suppress HCC progression by inhibiting mitochondrial activity, thereby reinforcing its role as a tumor suppressor in HCC.
利益披露 Disclosure
J. Huang, None..
H. Lin, None..
Y. Su, None..
R. Mai, None.