Marković Ivana, Debeljak Željko, Dobrošević Blaženka, Lukić Maja, Mrđenović Stefan, Kotris Ana, Bošnjak Bojana, Dmitrović Branko
Clinical Institute of Laboratory Diagnostics, University Hospital Centre Osijek, J. Huttlera 4, 31 000 Osijek, Croatia; Faculty of Medicine Osijek, JJ Strossmayer University of Osijek, J. Huttlera 4, 31 000 Osijek, Croatia.
Clinical Institute of Laboratory Diagnostics, University Hospital Centre Osijek, J. Huttlera 4, 31 000 Osijek, Croatia; Faculty of Medicine Osijek, JJ Strossmayer University of Osijek, J. Huttlera 4, 31 000 Osijek, Croatia.
Clin Chim Acta. 2024 Jul 15;561:119758. doi: 10.1016/j.cca.2024.119758. Epub 2024 Jun 6.
Modern mass spectrometry imaging (MSI) enables single cells' metabolism exploration. Aims of this study were development of the single-cell MSI of human CD19+ lymphocytes and metabolic profiling of chronic lymphocytic leukemia (CLL).
Blood donor (BD) samples were used for the optimization of CD19+ lymphocyte isolation and single-cell matrix-assisted laser desorption/ionization time-of-flight (MALDI TOF) MSI. Independent set of 200 CD19+ lymphocytes coming from 5 CLL patients and 5 BD was used for the CD19+ lymphocytes classification assessment and the untargeted metabolic profiling. CLL vs BD lymphocyte classification was performed using partial least squares-discriminant analysis (PLS-DA) using normalized single-cell mass spectra recorded in 300-600 and 600-950 Da ranges was applied.
Accuracy assessed by 10-fold cross-validation of CD19+ lymphocyte PLS-DA classification reached >90.0 %. Volcano plots showed 106 significantly altered m/z signals in CLL of which 9 were tentatively annotated. Among tentatively annotated m/z signals formaldehyde and glutathione metabolites and tetrahydrofolate stand out.
A method for single-cell MALDI TOF MSI of CD19+ lymphocytes was successfully developed. The method confirmed the significance of oxidative stress and single-carbon metabolism, pyruvate and fatty acid metabolism and apoptosis in CLL and it provided metabolic candidates for diagnostic applications.
现代质谱成像（MSI）能够探索单细胞的代谢情况。本研究的目的是开发人CD19 +淋巴细胞的单细胞MSI，并对慢性淋巴细胞白血病（CLL）进行代谢谱分析。
使用献血者（BD）样本优化CD19 +淋巴细胞的分离及单细胞基质辅助激光解吸/电离飞行时间（MALDI TOF）MSI。来自5例CLL患者和5名BD的200个独立的CD19 +淋巴细胞用于CD19 +淋巴细胞分类评估和非靶向代谢谱分析。使用偏最小二乘判别分析（PLS - DA）对CLL与BD淋巴细胞进行分类，采用记录在300 - 600和600 - 950 Da范围内的归一化单细胞质谱。
通过对CD19 +淋巴细胞PLS - DA分类进行10倍交叉验证评估的准确率达到>90.0%。火山图显示CLL中有106个显著改变的m/z信号，其中9个进行了初步注释。在初步注释的m/z信号中，甲醛、谷胱甘肽代谢物和四氢叶酸较为突出。
成功开发了一种用于CD19 +淋巴细胞的单细胞MALDI TOF MSI方法。该方法证实了氧化应激、一碳代谢、丙酮酸和脂肪酸代谢以及凋亡在CLL中的重要性，并为诊断应用提供了代谢候选物。
