Jia Hailan, Wang Chi, Fu Yan, Wang Yalin, Zhang Xiaoyu, Tang Yuezhou, Ding Jiahao, He Kun, Wang Jing, Shen Yanting
School of Pharmacy, Key Laboratory of Innovative Drug Development and Evaluation, Hebei Medical University, Shijiazhuang, People's Republic of China.
Core Facilities and Centers, Hebei Medical University, Shijiazhuang, People's Republic of China.
Anal Chim Acta. 2025 Apr 1;1345:343748. doi: 10.1016/j.aca.2025.343748. Epub 2025 Jan 30.
Mitophagy is a selective way to eliminate dysfunctional mitochondria and recycle their constituents, which plays an important role in regulating and maintaining intracellular homeostasis. Real-time monitoring mitophagy process is of great importance for cellular physiological and pathological processes related to mitochondria. Howbeit, most of the current methods only focus on single-parameter detection of mitochondrial microenvironmental changes such as pH, viscosity and polarity. The mitochondrial molecular responses under mitophagy are not clear. Therefore, developing a new and simple method for molecular profiling is of great importance for accurately and comprehensively visualizing mitophagy.
In this work, Au NPs-based mitochondria-targeting nanoprobe was developed and the nanoprobe-based label-free surface enhanced Raman spectroscopy (SERS) method was proposed to track starvation induced mitophagy process at molecular level. The nanoprobe displayed good SERS performance and low cytotoxicity. Based on the developed strategy, the molecular response within mitochondria under mitophagy was validated. Meanwhile, the protein denaturation, conformational change, lipid degradation and DNA fragmentation within mitochondria under mitophagy were revealed for the first time, which provides molecular evidence for mitophagy. The changes in reactive oxygen species level and mitochondrial membrane potential further confirmed the damage of mitochondria. Moreover, the developed label-free SERS strategy was used to detect mitophagy in drug (cisplatin)-induced liver injury (DILI) cell model, and obvious mitophagy in DILI cells was observed.
The molecular biochemical signature dynamic changes within mitochondria during mitophagy process were revealed by SERS for the first time. Moreover, compared with the current research, our study can provide new insights into mitophagy and mitophagy-involved diseases at molecular level. This study will provide new insights into the molecular mechanism of mitophagy and offer a simple and effective method for mitochondrial molecular event monitoring in mitophagy-involved cellular processes.
线粒体自噬是一种消除功能失调的线粒体并回收其成分的选择性方式，在调节和维持细胞内稳态中发挥重要作用。实时监测线粒体自噬过程对于与线粒体相关的细胞生理和病理过程至关重要。然而，目前大多数方法仅关注线粒体微环境变化的单参数检测，如pH、粘度和极性。线粒体自噬下的分子反应尚不清楚。因此，开发一种新的简单分子谱分析方法对于准确、全面地可视化线粒体自噬具有重要意义。
在本研究中，开发了基于金纳米粒子的线粒体靶向纳米探针，并提出了基于纳米探针的无标记表面增强拉曼光谱（SERS）方法，以在分子水平上追踪饥饿诱导的线粒体自噬过程。该纳米探针表现出良好的SERS性能和低细胞毒性。基于所开发的策略，验证了线粒体自噬下线粒体内的分子反应。同时，首次揭示了线粒体自噬下线粒体内的蛋白质变性、构象变化、脂质降解和DNA片段化，为线粒体自噬提供了分子证据。活性氧水平和线粒体膜电位的变化进一步证实了线粒体的损伤。此外，所开发的无标记SERS策略用于检测药物（顺铂）诱导的肝损伤（DILI）细胞模型中的线粒体自噬，并观察到DILI细胞中明显的线粒体自噬。
SERS首次揭示了线粒体自噬过程中线粒体内分子生化特征的动态变化。此外，与当前研究相比，我们的研究可以在分子水平上为线粒体自噬和涉及线粒体自噬的疾病提供新的见解。本研究将为线粒体自噬的分子机制提供新的见解，并为涉及线粒体自噬的细胞过程中线粒体分子事件的监测提供一种简单有效的方法。
