Targeted Detection of Phospholipid Aldehydes in Living Cells by Oxime Ligation.

IF 16.9
Jacob A Vance, Wei-An Chen, Palina Nepachalovich, Caroline Knittel, Maria Fedorova, Neal K Devaraj
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Abstract

Oxidatively damaged lipids play critical roles in numerous human pathologies, yet methods to directly identify these species within living cells remain limited due to their transient and low-abundance nature. Among lipid oxidation products, aldehyde-containing lipids are significant due to their heightened reactivity and strong links to disease pathology. Here, we introduce OxiLox (oxime ligation to oxidized lipids), a method that leverages fluorescent hydroxylamine-based probes to enable direct, chemoselective tagging of aldehyde-containing lipids within living cells via rapid oxime ligation. Using confocal microscopy, we observe the subcellular localization of these oxidized lipids, highlighting prominent accumulation within perinuclear membranes, particularly under oxidative stress conditions induced by the ferroptosis activator RSL3. Coupled with high-resolution mass spectrometry, our method identifies aldehyde lipid species, notably revealing the abundance of oxidized plasmenyl phosphatidylethanolamines in ferroptotic cells. We demonstrate that cellular labeling by hydroxylamine probes can be modulated by altering the expression of antioxidant enzymes such as aldose reductase (AKR1B1), underscoring the relationship between enzymatic antioxidant defenses and lipid aldehyde metabolism. Our work establishes OxiLox as a robust approach for direct, sensitive, and chemically precise detection of lipid aldehydes in live cells, offering new insights into the role of lipid oxidation in health and disease.

肟结扎法靶向检测活细胞中磷脂醛。
氧化损伤的脂类在许多人类疾病中起着关键作用,但由于其短暂和低丰度的性质,直接在活细胞内识别这些物种的方法仍然有限。在脂质氧化产物中,含醛脂质因其高反应性和与疾病病理的密切联系而具有重要意义。在这里,我们介绍OxiLox(肟连接氧化脂质),这是一种利用荧光羟胺探针通过快速肟连接在活细胞内实现含醛脂质的直接化学选择性标记的方法。使用共聚焦显微镜,我们观察到这些氧化脂质的亚细胞定位,突出了核周膜内的显著积累,特别是在由铁下垂激活剂RSL3诱导的氧化应激条件下。结合高分辨率质谱分析,我们的方法鉴定了醛类脂质,特别是揭示了铁致细胞中氧化plasmenyl磷脂酰乙醇胺的丰度。我们证明了羟胺探针的细胞标记可以通过改变醛糖还原酶(AKR1B1)等抗氧化酶的表达来调节,强调了酶抗氧化防御与脂质醛代谢之间的关系。我们的工作建立了OxiLox作为一种直接、敏感和化学精确检测活细胞中脂质醛的强大方法,为脂质氧化在健康和疾病中的作用提供了新的见解。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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