Emaus Katlynn J, Dunbar Carmen A, Caruso Joseph, Ruotolo Brandon T, Wider Joseph M, Sanderson Thomas H
Neuroscience Graduate Program, University of Michigan, Ann Arbor, MI, United States.
Department of Emergency Medicine, University of Michigan, Ann Arbor, MI, United States.
Front Physiol. 2025 May 29;16:1592008. doi: 10.3389/fphys.2025.1592008. eCollection 2025.
The mitochondrial phospholipid cardiolipin (CL) is essential for proper mitochondrial function and energy production. Cardiolipin has four distinct fatty acid tails with varying expression compositions, resulting in a highly variable tissue-specific distribution of isomer expression. Neuronal cardiolipin has a remarkable variety of subspecies and has recently been used as a biomarker to predict brain injury severity following cardiac arrest and traumatic brain injury. Multiple conditions have been associated with disordered cardiolipin remodeling, including Alzheimer's disease, Parkinson's disease, Barth syndrome, and astrocytoma. The clinical relevance of cardiolipin as a biomarker and the importance of the mechanistic role of cardiolipin remodeling in disease emphasize the demand for a reliable and accurate means of the identification and quantification of cardiolipin. In this study, we outline the use of a novel method of cardiolipin analysis using cyclic ion mobility mass spectrometry (cIMS-MS) to isolate and identify cardiolipin subspecies in several biological samples. Furthermore, cIMS-MS established the composition of the cardiolipin profile by individual subspecies across biological samples under basal conditions. Monolysocardiolipin (MLCL), the precursor of mature cardiolipin and a primary diagnostic biomarker of Barth syndrome, was isolated from cardiolipin and identified. The monolysocardiolipin:cardiolipin ratio was quantified in brain samples from tafazzin-knockout (KO) mice, demonstrating accumulation of MLCL and providing direct evidence for the validity of this cIMS-MS methodology through genetic loss-of-function. The novel, multiple-pass feature of cIMS-MS enabled the isolation and amplification of less abundant cardiolipin subspecies in both standards and biological samples. This protocol enables rapid analysis of biological samples, allowing researchers to further dissect the mechanistic role of cardiolipin in injury pathology, with simplified sample preparation and reduced potential for artifact introduction.
线粒体磷脂心磷脂（CL）对于线粒体的正常功能和能量产生至关重要。心磷脂有四条不同的脂肪酸尾巴，其表达组成各异，导致异构体表达在组织特异性分布上高度可变。神经元心磷脂有多种亚型，最近已被用作预测心脏骤停和创伤性脑损伤后脑损伤严重程度的生物标志物。多种疾病与心磷脂重塑紊乱有关，包括阿尔茨海默病、帕金森病、巴氏综合征和星形细胞瘤。心磷脂作为生物标志物的临床相关性以及心磷脂重塑在疾病中的机制作用的重要性，凸显了对可靠、准确的心磷脂鉴定和定量方法的需求。在本研究中，我们概述了一种使用循环离子淌度质谱（cIMS-MS）分析心磷脂的新方法，用于分离和鉴定几种生物样品中的心磷脂亚型。此外，cIMS-MS在基础条件下确定了跨生物样品的各个心磷脂亚型的心磷脂谱组成。从心磷脂中分离并鉴定了单溶血心磷脂（MLCL），它是成熟心磷脂的前体，也是巴氏综合征的主要诊断生物标志物。在tafazzin基因敲除（KO）小鼠的脑样本中对单溶血心磷脂与心磷脂的比例进行了定量，证明了MLCL的积累，并通过基因功能丧失为这种cIMS-MS方法的有效性提供了直接证据。cIMS-MS的新型多通道特性能够在标准品和生物样品中分离和扩增丰度较低的心磷脂亚型。该方案能够快速分析生物样品，简化了样品制备过程，减少了引入假象的可能性，使研究人员能够进一步剖析心磷脂在损伤病理中的机制作用。
