The regulatory definition(s) of nanomaterials (NMs) frequently uses the term ‘agglomerates and aggregates’ (AA) despite the paucity of evidence that AA are significantly relevant from a nanotoxicological perspective. This knowledge gap greatly affects the safety assessment and regulation of NMs, such as synthetic amorphous silica (SAS). SAS is used in a large panel of industrial applications. They are primarily produced as nano-sized particles (1–100 nm in diameter) and considered safe as they form large aggregates (> 100 nm) during the production process. So far, it is indeed believed that large aggregates represent a weaker hazard compared to their nano counterpart. Thus, we assessed the impact of SAS aggregation on in vitro cytotoxicity/biological activity to address the toxicological relevance of aggregates of different sizes. We used a precipitated SAS dispersed by different methods, generating 4 ad-hoc suspensions with different aggregate size distributions. Their effect on cell metabolic activity, cell viability, epithelial barrier integrity, total glutathione content and, IL-8 and IL-6 secretion were investigated after 24 h exposure in human bronchial epithelial (HBE), colon epithelial (Caco2) and monocytic cells (THP-1). We observed that the de-aggregated suspension (DE-AGGR), predominantly composed of nano-sized aggregates, induced stronger effects in all the cell lines than the aggregated suspension (AGGR). We then compared DE-AGGR with 2 suspensions fractionated from AGGR: the precipitated fraction (PREC) and the supernatant fraction (SuperN). Very large aggregates in PREC were found to be the least cytotoxic/biologically active compared to other suspensions. SuperN, which contains aggregates larger in size (> 100 nm) than in DE-AGGR but smaller than PREC, exhibited similar activity as DE-AGGR. Overall, aggregation resulted in reduced toxicological activity of SAS. However, when comparing aggregates of different sizes, it appeared that aggregates > 100 nm were not necessarily less cytotoxic than their nano-sized counterparts. This study suggests that aggregates of SAS are toxicologically relevant for the definition of NMs.

中文翻译：

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聚集的合成无定形二氧化硅是否具有毒理学意义？

尽管从纳米毒理学的角度来看，AA的意义不大，但纳米材料（NMs）的监管定义经常使用术语“附聚物和聚集体”（AA）。这种知识差距极大地影响了诸如合成无定形二氧化硅（SAS）之类的NM的安全评估和监管。SAS用于大量工业应用。它们主要以纳米级颗粒（直径为1–100 nm）生产，由于在生产过程中会形成大的聚集体（> 100 nm），因此被认为是安全的。到目前为止，确实可以相信，与纳米纳米颗粒相比，大的聚集体危害较小。因此，我们评估了SAS聚集体对体外细胞毒性/生物学活性的影响，以解决不同大小聚集体的毒理学相关性。我们使用了通过不同方法分散的沉淀SAS，生成了4种具有不同聚集体大小分布的临时悬浮液。在人支气管上皮细胞（HBE），结肠上皮细胞（Caco2）和单核细胞中暴露24小时后，研究了它们对细胞代谢活性，细胞活力，上皮屏障完整性，总谷胱甘肽含量以及IL-8和IL-6分泌的影响（ THP-1）。我们观察到，主要由纳米尺寸的聚集体组成的解聚集悬浮液（DE-AGGR）在所有细胞系中均比聚集悬浮液（AGGR）产生更强的作用。然后，我们将DE-AGGR与从AGGR分离的2种悬浮液进行了比较：沉淀级分（PREC）和上清液级分（SuperN）。与其他悬浮液相比，发现PREC中非常大的聚集体具有最低的细胞毒性/生物学活性。SuperN包含比DE-AGGR大（> 100 nm）但小于PREC的聚集体，其活性与DE-AGGR类似。总体而言，聚集导致SAS的毒理活性降低。但是，当比较不同大小的聚集体时，似乎> 100 nm的聚集体不一定比其纳米大小的聚集体具有更低的细胞毒性。这项研究表明，SAS的聚集体与NMs的毒理学相关。