Abstract
Due to the vigorous development of nanotechnology, nanomaterials are used in a variety of fields including commercial products and biomedical applications. Silver nanoparticles (AgNPs) are applied to daily utilities due to antimicrobial activity, which increase the chance of AgNPs exposure in human. AgNPs can get into human body via dermal contact, ingestion and inhalation, and distribute to various organs via blood circulation. AgNPs can cross through blood brain barrier (BBB) to potentially induce neurotoxicity for neurodegenerative diseases. However, there is not clearly how AgNPs disrupt BBB. Thus, this study investigated the potential effects of 3-5 nm AgNPs on inflammation and tight junctions individually in mouse brain endothelial bEnd3 cells and in vitro model of mouse BBB. In vitro model of mouse BBB was constrcted by bEnd3 cells, mouse brain ALT astrocytes and mouse neuroblastoma N2a cells to investigate the potential effects of 3-5 nm AgNPs on gene expression of inflammation and tight junctions. The results showed AgNPs exposure increased the gene expression relevant to inflammation (CXC motif chemokine 13 (CXCL13), macrophage receptor with collagenous structure (MARCO), glutathione synthetase (GSS), tumor necrosis factor α (TNFα), interlukin-6 (IL-6) and interlukin-1β (IL-1β)) and decreased the gene expression of claudin-5 (Cldn-5) and Lrp5 in bEnd.3 cells. The zona occludens-1 (ZO-1) and occludin (Ocln) gene expression had no significant effect in bEnd.3 cells after AgNPs exposure. In bEnd.3 cells and ALT cells co-culture system, as AgNPs exposure concentration increased, the lower the TEER value in bEnd.3 cells which lead to AgNPs cross through BBB. Additionally, immunofluorescence images showed reduction of claudin-5 and ZO-1 protein for BBB disruption in bEnd.3 cells of co-culture system. After the triple cell co-culture system exposed to AgNPs (2, 3 and 4 g/ml) for 24 hour, the gene expression relevant to inflammation (interleukin-2 (IL-2)、interleukin-17A (IL-17A)、interferon gamma (IFN-γ)、IL-6、IL-1β and monocyte chemoattractant protein-1 (MCP-1) decreased in bEnd.3 cells；The gene expression of IL-2, IL-17A, IFN-γ and MCP-1 increased in ALT cells；Only the MCP-1 gene expression increased in N2a cells. Besides, 4 g/mL AgNPs exposure increased MCP-1 and IL-6 cytokine secreation and decreased IL-1β cytokine secreation in the upper medium of in triple cell co-culture system. This study found that AgNPs exposure increased the gene expression relevant to inflammation and decreased the gene expression of Cldn-5 in bEnd.3 cells. In triple cell co-culture system, AgNPs exposure reduced claudin-5 and ZO-1 protein expression leading to permeability of bEnd.3 cells rise. AgNPs can cross through BBB leading to inflammation in ALT cells. Besides, AgNPs exposure obviously altered the gene expression of tight junctions and inflammation in vitro model of mouse BBB. Therefore the effect of AgNPs to human body should be concern.