Heilala Maria, Turpin Rita, Pahimanolis Nikolaos, Ikkala Olli, Klefström Juha, Munne Pauliina M
Department of Applied Physics, Aalto University. P.O. Box 15100, FI-00076 Aalto, Espoo Finland.
Cancer Cell Circuitry Laboratory, Translational Cancer Medicine, Medical Faculty, University of Helsinki. P.O. Box 63, Haartmaninkatu 8, FI-00014 Helsinki, Finland.
ACS Appl Mater Interfaces. 2025 Jul 16;17(28):40082-40100. doi: 10.1021/acsami.5c06549. Epub 2025 Jul 3.
Anti-inflammatory M2 macrophages are highly relevant in various physiological processes ranging from tissue regeneration to cancer progression. However, conventional two-dimensional (2D) cell cultures limit our understanding of macrophage phenotypes and how they can be modulated for immunotherapeutic approaches. Moreover, there is a growing demand for scalable, animal-free hydrogels to replace animal-derived materials in three-dimensional (3D) models. In this study, we explore hydrogels based on plant-derived nanofibrillar cellulose (NFC), also known as cellulose nanofibrils (CNFs) or microfibrillated cellulose (MFC), for generating 3D models of M2-like macrophages from human blood monocytes. Notably, flow cytometry analysis shows that cells cultured in 3D phosphorylated NFC hydrogels show enhanced expression of the M2 macrophage marker CD206 compared to cells cultured in other negatively charged hydrogels prepared from native NFC or NFCs with carboxylate or sulfate modifications. Furthermore, the upregulation of CD206 expression in 3D phosphorylated NFC is comparable to the induction of CD206 in interleukin 4 (IL-4)-differentiated M2a macrophages. In addition, the cells in the phosphorylated NFC hydrogel show a differential cytokine profile compared to 2D cultured cells, secreting similar levels of tumor necrosis factor α (TNF-α), but 2.6-fold higher amounts of IL-1β and 1.2-fold higher amounts of IL-10. The results suggest that the conversion of monocytes to M2-like macrophages can be controlled by the phosphorylation of NFC, a strategy which does not require the addition of polarization factors like growth factors and cytokines conventionally used to generate macrophages . The findings highlight the importance of surface chemistry in matrix-guided macrophage polarization, paving the way for xeno-free yet bioactive 3D macrophage culture scaffolds for immunological research.
抗炎性M2巨噬细胞在从组织再生到癌症进展的各种生理过程中高度相关。然而，传统的二维（2D）细胞培养限制了我们对巨噬细胞表型以及如何针对免疫治疗方法对其进行调节的理解。此外，对可扩展的、无动物的水凝胶的需求日益增长，以在三维（3D）模型中替代动物源材料。在本研究中，我们探索了基于植物源纳米原纤化纤维素（NFC）的水凝胶，也称为纤维素纳米原纤（CNF）或微纤化纤维素（MFC），用于从人血单核细胞生成M2样巨噬细胞的3D模型。值得注意的是，流式细胞术分析表明，与在由天然NFC或具有羧酸盐或硫酸盐修饰的NFC制备的其他带负电荷的水凝胶中培养的细胞相比，在3D磷酸化NFC水凝胶中培养的细胞显示出M2巨噬细胞标志物CD206的表达增强。此外，3D磷酸化NFC中CD206表达的上调与白细胞介素4（IL-4）分化的M2a巨噬细胞中CD206的诱导相当。此外，与2D培养的细胞相比，磷酸化NFC水凝胶中的细胞显示出不同的细胞因子谱，分泌相似水平的肿瘤坏死因子α（TNF-α），但IL-1β的量高2.6倍，IL-10的量高1.2倍。结果表明，单核细胞向M2样巨噬细胞的转化可通过NFC的磷酸化来控制，该策略不需要添加通常用于生成巨噬细胞的极化因子如生长因子和细胞因子。这些发现突出了表面化学在基质引导的巨噬细胞极化中的重要性，为用于免疫学研究的无动物但具有生物活性的3D巨噬细胞培养支架铺平了道路。
