Chang Eun Sol, Song Kyoung, Song Ji-Young, Sung Minjung, Lee Mi-Sook, Oh Jung Han, Kim Ji-Yeon, Park Yeon Hee, Jung Kyungsoo, Choi Yoon-La
Department of Health Sciences and Technology, SAIHST, Sungkyunkwan University, Seoul, South Korea.
Laboratory of Molecular Pathology and Theranostics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Irwon-Ro 81, Gangnam-Go, Seoul, 06351, South Korea.
Cancer Metab. 2024 Aug 29;12(1):25. doi: 10.1186/s40170-024-00353-3.
Mitochondria are known to synthesize adenosine triphosphate (ATP) through oxidative phosphorylation. Understanding and accurately measuring mitochondrial ATP synthesis rate can provide insights into the functional status of mitochondria and how it contributes to overall cellular energy homeostasis. Traditional methods only estimate mitochondrial function by measuring ATP levels at a single point in time or through oxygen consumption rates. This study introduced the relative mitochondrial ATP synthesis response against inhibiting and stimulating substrates (MitoRAISE), designed to detect real-time changes in ATP levels as the cells respond to substrates.
The sensitivity and specificity of the MitoRAISE assay were verified under various conditions, including the isolation of mitochondria, variations in cell numbers, cells exhibiting mitochondrial damage, and heterogeneous mixtures. Using peripheral blood mononuclear cells (PBMCs), we analyzed MitoRAISE data from 19 patients with breast cancer and 23 healthy women.
The parameters observed in the MitoRAISE data increased depending on the quantity of isolated mitochondria and cell count, whereas it remained unmeasured in mitochondrial-damaged cell lines. Basal ATP, rotenone response, malonate response, and mitochondrial DNA copy numbers were lower in PBMCs from patients with breast cancer than in those from healthy women.
The MitoRAISE assay has demonstrated its sensitivity and specificity by measuring relative ATP synthesis rates under various conditions. We propose MitoRAISE assay as a potential tool for monitoring changes in the mitochondrial metabolic status associated with various diseases.
已知线粒体通过氧化磷酸化合成三磷酸腺苷（ATP）。了解并准确测量线粒体ATP合成速率有助于深入了解线粒体的功能状态及其对细胞整体能量稳态的作用。传统方法仅通过在单个时间点测量ATP水平或通过氧消耗率来估计线粒体功能。本研究引入了针对抑制性和刺激性底物的相对线粒体ATP合成反应（MitoRAISE），旨在检测细胞对底物反应时ATP水平的实时变化。
在各种条件下验证了MitoRAISE检测的敏感性和特异性，包括线粒体的分离、细胞数量的变化、表现出线粒体损伤的细胞以及异质混合物。我们使用外周血单核细胞（PBMC）分析了19例乳腺癌患者和23名健康女性的MitoRAISE数据。
MitoRAISE数据中观察到的参数随分离的线粒体数量和细胞计数的增加而增加，而在受损线粒体的细胞系中未检测到该参数。乳腺癌患者PBMC中的基础ATP、鱼藤酮反应、丙二酸反应和线粒体DNA拷贝数均低于健康女性。
MitoRAISE检测通过在各种条件下测量相对ATP合成速率，证明了其敏感性和特异性。我们建议将MitoRAISE检测作为监测与各种疾病相关的线粒体代谢状态变化的潜在工具。
