Department of Chemical and Biomedical Engineering, FAMU-FSU College of Engineering, 2525 Pottsdamer Street, Tallahassee, FL 32310-2870, USA.
College of Medicine, Florida State University, Tallahassee, FL 32306-4370, USA.
Acta Biomater. 2022 Mar 15;141:344-353. doi: 10.1016/j.actbio.2022.01.028. Epub 2022 Jan 19.
Phagocytosis performed by a macrophage involves complex membrane trafficking and reorganization among various membranous cellular structures including phagosomes and vesicles derived from the phagosomes known as phagosome-derived vesicles. The present work reports on development of a technique that allows to specifically label the phagosome-derived vesicles in macrophages with a membrane dye. The technique is based on the use of microfabricated microparticles that are made of a thermosensitive nonbiodegradable polymer poly(N-isopropylacrylamide) (PNIPAM) or its derivative and contain a membrane dye 1,1'-dialkyl-3,3,3',3'-tetramethylindodicarbocyanine (DiI). The microparticles can be phagocytosed by RAW264.7 macrophages into their phagosomes, resulting in formation of intracellular DiI-positive vesicles derived from the phagosomes. The DiI-positive vesicles are motile and acidic; can be stained by fluorescently labelled dextran added in the culture medium; and can accumulate around new phagosomes, indicating that they possess properties of lysosomes. This technique is also applicable to another membrane dye 3,3'-dioctadecyloxacarbocyanine (DiO) and holds great potential to be useful for advancing our understanding of phagocytosis. STATEMENT OF SIGNIFICANCE: Phagocytosis performed by macrophages is a cellular process of great importance to various applications of biomaterials such as drug delivery and medical implantation. This work reports on a technique for characterizing phagocytosis based on the use of poly(N-isopropylacrylamide), which is a major biomaterial with numerous applications. This technique is the first of its kind and has generated an original finding about phagocytosis. In addition to drug delivery and medical implantation, phagocytosis plays critical roles in diseases, injuries and vaccination. This work could thus attract immediate and widespread interests in the field of biomaterials science and engineering.
巨噬细胞的吞噬作用涉及到各种膜结构之间复杂的膜运输和重排，包括吞噬体和来自吞噬体的囊泡，这些囊泡称为吞噬体衍生的囊泡。本工作报道了一种技术的发展，该技术允许用膜染料特异性标记巨噬细胞中的吞噬体衍生的囊泡。该技术基于使用由热响应性不可生物降解的聚合物聚（N-异丙基丙烯酰胺）（PNIPAM）或其衍生物制成的微制造微颗粒，该微颗粒包含膜染料 1,1'-二烷基-3,3,3',3'-四甲基吲哚羰花青（DiI）。微颗粒可以被 RAW264.7 巨噬细胞吞噬到它们的吞噬体中，导致来自吞噬体的细胞内 DiI 阳性囊泡的形成。DiI 阳性囊泡是运动的和酸性的；可以用添加在培养基中的荧光标记葡聚糖染色；并且可以围绕新的吞噬体积累，表明它们具有溶酶体的特性。该技术也适用于另一种膜染料 3,3'-二辛基氧杂羰花青（DiO），并且对于深入了解吞噬作用具有很大的潜力。意义声明：巨噬细胞的吞噬作用是一种对生物材料的各种应用（如药物输送和医学植入）非常重要的细胞过程。这项工作报道了一种基于使用聚（N-异丙基丙烯酰胺）的吞噬作用的特征化技术，聚（N-异丙基丙烯酰胺）是一种具有众多应用的主要生物材料。该技术是首创的，并且产生了关于吞噬作用的原始发现。除了药物输送和医学植入之外，吞噬作用在疾病、损伤和疫苗接种中也起着至关重要的作用。因此，这项工作可能会立即引起生物材料科学和工程领域的广泛关注。
