Increased uptake of silica nanoparticles in inflamed macrophages but not upon co-exposure to micron-sized particles

Silica nanoparticles (NPs) are widely used in various industrial and biomedical applications. Little is known about the cellular uptake of co-exposed silica particles, as can be expected in our daily life. In addition, an inflamed microenvironment might affect a NP’s uptake and a cell’s physiological response. Herein, prestimulated mouse J774A.1 macrophages with bacterial lipopolysaccharide were post-exposed to micron- and nanosized silica particles, either alone or together, i.e., simultaneously or sequentially, for different time points. The results indicated a morphological change and increased expression of tumor necrosis factor alpha in lipopolysaccharide prestimulated cells, suggesting a M1-polarization phenotype. Confocal laser scanning microscopy revealed the intracellular accumulation and uptake of both particle types for all exposure conditions. A flow cytometry analysis showed an increased particle uptake in lipopolysaccharide prestimulated macrophages. However, no differences were observed in particle uptakes between single- and co-exposure conditions. We did not observe any colocalization between the two silica (SiO₂) particles. However, there was a positive colocalization between lysosomes and nanosized silica but only a few colocalized events with micro-sized silica particles. This suggests differential intracellular localizations of silica particles in macrophages and a possible activation of distinct endocytic pathways. The results demonstrate that the cellular uptake of NPs is modulated in inflamed macrophages but not in the presence of micron-sized particles.二氧化硅纳米颗粒（silica nanoparticles，NPs）广泛应用于各类工业与生物医学领域。目前，人们对日常生活中可接触到的联合暴露二氧化硅颗粒的细胞摄取情况仍知之甚少。此外，炎症微环境可能会影响纳米颗粒的细胞摄取过程以及细胞的生理应答反应。本研究中，我们使用细菌脂多糖预先刺激小鼠J774A.1巨噬细胞，随后将其分别以单一暴露或联合暴露的方式接触微米级与纳米级二氧化硅颗粒，联合暴露又分为同时暴露与序贯暴露两种模式，并设置了不同的暴露时长。实验结果显示，经脂多糖预先刺激的巨噬细胞出现了形态学改变，且肿瘤坏死因子α（tumor necrosis factor alpha）的表达水平显著上调，表明细胞呈现M1极化表型。共聚焦激光扫描显微镜观测结果表明，所有暴露条件下，两类颗粒均能在细胞内发生蓄积并被细胞摄取。流式细胞术分析结果显示，经脂多糖预先刺激的巨噬细胞对颗粒的摄取量有所升高。但在单一暴露与联合暴露组之间，颗粒摄取量并未观察到显著差异。我们未观测到两类二氧化硅（SiO₂）颗粒之间存在任何共定位现象。不过，溶酶体与纳米级二氧化硅颗粒之间存在显著共定位，而与微米级二氧化硅颗粒的共定位事件则极少。这表明巨噬细胞内不同尺寸的二氧化硅颗粒的细胞内定位存在差异，且可能通过不同的内吞途径被摄取。本研究结果证实，炎症状态下的巨噬细胞对纳米颗粒的摄取过程会受到调控，但微米级二氧化硅颗粒的摄取则不受该因素影响。