Mancini Caterina, Menegazzi Giulio, Peppicelli Silvia, Versienti Giampaolo, Guasti Daniele, Pieraccini Giuseppe, Rovida Elisabetta, Lulli Matteo, Papucci Laura, Dello Sbarba Persio, Biagioni Alessio
Department of Experimental and Clinical Medicine, Università degli Studi di Firenze, Florence, Italy.
Department of Experimental and Clinical Biomedical Sciences "Mario Serio", Università degli Studi di Firenze, Florence, Italy.
Cancer Cell Int. 2025 May 14;25(1):176. doi: 10.1186/s12935-025-03805-y.
Chronic myeloid leukemia (CML) is influenced by microenvironmental nutrients, glucose (Glc), and glutamine (Gln) which regulate cell proliferation, viability, and the expression of the driver oncoprotein (BCR::ABL1).
Our study revealed that Glc, while partially supporting alone cell growth in normoxia, is essential in low oxygen conditions, whereas Gln is ineffective. Under low oxygen, Gln reduced oxidative respiratory activity while enhancing glycolysis. In these conditions, fatty acid (FA) metabolism becomes crucial, as evidenced by increased lipid droplets (LD) accumulation when Glc was absent. Gln, in particular, drives CD36-mediated FA uptake, suppressing the BCR::ABL1 oncoprotein and facilitating cell survival. By co-culturing leukemia cells with adipocytes, one of the main bone marrow (BM) cell components, we observed an enhanced FA release, suggesting a link between FA, microenvironmental BM cells, and the maintenance of leukemic stem cells (LSC).
K562 and KCL22 cell lines were subjected to Glc and/or Gln deprivation under hypoxic conditions (96 h at 0.1% O). Metabolic profiling was conducted through the Seahorse XFe96 analyzer, and the contribution of L-Glutamine-C to FA de novo synthesis was determined via GC/MS. Intracellular neutral LD were measured using BODIPY 493/503 in confocal microscopy and flow cytometry, with their presence and morphology further examined via transmission electron microscopy. BCR::ABL1 as well as several FA-related markers were evaluated via Western Blotting, whilst CD36 was determined through flow cytometry. LC2 assay was used for measuring leukemia stem cell potential by inhibiting FA uptake via the usage of the Sulfo-N-Succinimidyl Oleate, a CD36 inhibitor. qPCR was exploited to detect markers of FA secretion in CML-adipocytes co-culture together with Nile Red staining to assess free FA in the media.
These findings underscore the central role of FA in the regulation of the LSC compartment of CML, highlighting the importance of Gln in facilitating CML cell survival under restrictive metabolic conditions and preparing the cell population for expansion upon the release of these restrictions.
慢性粒细胞白血病（CML）受微环境营养物质、葡萄糖（Glc）和谷氨酰胺（Gln）的影响，这些营养物质调节细胞增殖、活力以及驱动癌蛋白（BCR::ABL1）的表达。
我们的研究表明，Glc虽然在常氧条件下仅能部分支持细胞生长，但在低氧条件下是必不可少的，而Gln则无效。在低氧条件下，Gln降低氧化呼吸活性，同时增强糖酵解。在这些条件下，脂肪酸（FA）代谢变得至关重要，当缺乏Glc时脂滴（LD）积累增加就证明了这一点。特别是，Gln驱动CD36介导的FA摄取，抑制BCR::ABL1癌蛋白并促进细胞存活。通过将白血病细胞与脂肪细胞（骨髓（BM）的主要细胞成分之一）共培养，我们观察到FA释放增加，这表明FA、微环境BM细胞与白血病干细胞（LSC）的维持之间存在联系。
将K562和KCL22细胞系在低氧条件下（0.1% O₂，96小时）进行Glc和/或Gln剥夺。通过Seahorse XFe96分析仪进行代谢分析，并通过气相色谱/质谱法测定L-谷氨酰胺-C对FA从头合成的贡献。在共聚焦显微镜和流式细胞术中使用BODIPY 493/503测量细胞内中性LD，通过透射电子显微镜进一步检查其存在和形态。通过蛋白质免疫印迹法评估BCR::ABL1以及几种FA相关标志物，同时通过流式细胞术测定CD36。使用LC2测定法通过使用CD36抑制剂磺基-N-琥珀酰亚胺油酸酯抑制FA摄取来测量白血病干细胞潜能。利用qPCR检测CML-脂肪细胞共培养中FA分泌的标志物，并使用尼罗红染色评估培养基中的游离FA。
这些发现强调了FA在CML的LSC区室调节中的核心作用，突出了Gln在限制代谢条件下促进CML细胞存活以及使细胞群体在这些限制解除后准备扩增方面的重要性。
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