Pérez-Montero Andrea, Zaragoza Oscar, Luque Alfonso, Hortelano Sonsoles, Acebo Paloma
Unidad de Terapias Farmacológicas, Instituto de Investigación de Enfermedades Raras (IIER), Instituto de Salud Carlos III, Madrid, Spain.
Programa de Doctorado UNED-ISCIII Ciencias Biomédicas y Salud Pública, Universidad Nacional de Educación a Distancia (UNED), Madrid, Spain.
Bio Protoc. 2023 Mar 5;13(5):e4629. doi: 10.21769/BioProtoc.4629.
Lipid droplets (LD), triglycerides and sterol esters among them, are well known for their capacity as lipid storage organelles. Recently, they have emerged as critical cytoplasmic structures involved in numerous biological functions. LD storage is generated de novo by the cell and provides an energy reserve, lipid precursors, and cell protection. Moreover, LD accumulation can be observed in some pathologies as obesity, atherosclerosis, or lung diseases. Fluorescence imaging techniques are the most widely used techniques to visualize cellular compartments in live cells, including LD. Nevertheless, presence of fluorophores can damage subcellular components and induce cytotoxicity, or even alter the dynamics of the organelles. As an alternative to fluorescence microscopy, label-free techniques such as stimulated Raman scattering and coherent anti-stokes Raman scattering microscopy offer a solution to avoid the undesirable effects caused by dyes and fluorescent proteins, but are expensive and complex. Here, we describe a label-free method using live-cell imaging by 3D holotomographic microscopy (Nanolive) to visualize LD accumulation in the MH-S alveolar macrophage cell line after treatment with oleic acid, a monounsaturated fatty acid that promotes lipid accumulation.
脂滴（LD），其中包括甘油三酯和固醇酯，作为脂质储存细胞器的能力广为人知。最近，它们已成为参与众多生物学功能的关键细胞质结构。脂滴储存由细胞重新产生，并提供能量储备、脂质前体和细胞保护。此外，在一些疾病如肥胖、动脉粥样硬化或肺部疾病中可以观察到脂滴积累。荧光成像技术是在活细胞中可视化细胞区室（包括脂滴）最广泛使用的技术。然而，荧光团的存在会损害亚细胞成分并诱导细胞毒性，甚至改变细胞器的动态。作为荧光显微镜的替代方法，诸如受激拉曼散射和相干反斯托克斯拉曼散射显微镜等无标记技术提供了一种解决方案，以避免由染料和荧光蛋白引起的不良影响，但它们昂贵且复杂。在这里，我们描述了一种无标记方法，使用三维全息断层显微镜（Nanolive）进行活细胞成像，以可视化油酸处理后MH-S肺泡巨噬细胞系中的脂滴积累，油酸是一种促进脂质积累的单不饱和脂肪酸。
