Lepur Adriana, Carlsson Michael C, Novak Ruđer, Dumić Jerka, Nilsson Ulf J, Leffler Hakon
Section MIG (Microbiology, Immunology, Glycobiology), Department of Laboratory Medicine, Lund University, 223 62 Lund, Sweden.
Biochim Biophys Acta. 2012 Jul;1820(7):804-18. doi: 10.1016/j.bbagen.2012.02.018. Epub 2012 Mar 17.
Galectin-3 (the Mac-2 antigen) is abundantly expressed in both macrophage like cells and certain non-macrophage cells. We have studied endocytosis of galectin-3 as one important step relevant for its function, and compared it between variants of a macrophage like cell line, and non-macrophage cells.
Endocytosis of galectin-3 was observed by fluorescence microscopy and measured by flow cytometry. The endocytosis mechanism was analysed using galectin-3 mutants, galectin-3 inhibitors and endocytic pathways inhibitors in the human leukaemia THP-1 cell line differentiated into naïve (M0), classical (M1) or alternatively activated (M2) macrophage like cells, and the non-macrophage cell lines HFL-1 fibroblasts and SKBR3 breast carcinoma.
Galectin-3 endocytosis in non-macrophage cells and M2 cells was blocked by lactose and a potent galectin-3 inhibitor TD139, and also by the R186S mutation in the galectin-3 carbohydrate recognition domain (CRD). In M1 cells galectin-3 endocytosis could be inhibited only by chlorpromazine and by interference with the non-CRD N-terminal part of galectin-3. In all the cell types galectin-3 entered early endosomes within 5-10 min, to be subsequently targeted mainly to non-degradative vesicles, where it remained even after 24 h.
Galectin-3 endocytosis in M1 cells is receptor mediated and carbohydrate independent, while in M2 cells it is CRD mediated, although the non-CRD galectin-3 domain is also involved. General significance The demonstration that galectin-3 endocytosis in M1 macrophages is carbohydrate independent and different from M2 macrophages and non-macrophage cells, suggests novel, immunologically significant interactions between phagocytic cells, galectin-3 and its ligands.
半乳糖凝集素-3（Mac-2抗原）在巨噬细胞样细胞和某些非巨噬细胞中均大量表达。我们研究了半乳糖凝集素-3的内吞作用，将其作为与其功能相关的重要步骤之一，并在巨噬细胞样细胞系的变体与非巨噬细胞之间进行了比较。
通过荧光显微镜观察半乳糖凝集素-3的内吞作用，并通过流式细胞术进行测量。使用半乳糖凝集素-3突变体、半乳糖凝集素-3抑制剂和内吞途径抑制剂，在分化为幼稚型（M0）、经典型（M1）或替代性活化型（M2）巨噬细胞样细胞的人白血病THP-1细胞系，以及非巨噬细胞系HFL-1成纤维细胞和SKBR3乳腺癌细胞中分析内吞作用机制。
乳糖和一种有效的半乳糖凝集素-3抑制剂TD139，以及半乳糖凝集素-3碳水化合物识别结构域（CRD）中的R186S突变，均可阻断非巨噬细胞和M2细胞中的半乳糖凝集素-3内吞作用。在M1细胞中，仅氯丙嗪和对半乳糖凝集素-3非CRD N端部分的干扰可抑制半乳糖凝集素-3内吞作用。在所有细胞类型中，半乳糖凝集素-3在5-10分钟内进入早期内体，随后主要靶向非降解性囊泡，即使在24小时后仍保留在其中。
M1细胞中的半乳糖凝集素-3内吞作用是受体介导的且不依赖碳水化合物，而在M2细胞中是CRD介导的，尽管半乳糖凝集素-3的非CRD结构域也参与其中。一般意义 M1巨噬细胞中的半乳糖凝集素-3内吞作用不依赖碳水化合物且不同于M2巨噬细胞和非巨噬细胞，这一发现提示了吞噬细胞、半乳糖凝集素-3及其配体之间新的、具有免疫学意义的相互作用。
