Interactions of silver nanoparticles with primary mouse fibroblasts and liver cells

Primary cells are ideal for in vitro toxicity studies since they closely resemble tissue environment. Here, we report a detailed study on the in vitro interactions of 7-20 nm spherical silver nanoparticles (SNP) with primary fibroblasts and primary liver cells isolated from Swiss albino mice. The intended use of silver nanoparticles is in the form of a topical antimicrobial gel formulation for the treatment of burns and wounds. Upon exposure to SNP for 24 h, morphology of primary fibroblasts and primary liver cells remained unaltered up to 25 mu g/mL and 100 mu g/mLSNP, respectively, although with minor decrease in confluence. IC50 values for primary fibroblasts and primary liver cells as revealed by XTT assay were 61 mu g/mL and 449 mu g/mL, respectively. Ultra-thin sections of primary cells exposed to 1/2 IC50 SNP for 24 h, visualized under Transmission electron microscope showed the presence of dark, electron dense. spherical aggregates inside the mitochondria, and cytoplasm, probably representing the intracellular SNP. When the cells were challenged with similar to 1/2 IC50 concentration of SNP (i.e. 30 mu g/mL and 225 mu g/mL for primary fibroblasts and primary liver cells, respectively), enhancement of GSH (similar to 1.2 fold) and depletion of lipid peroxidation (similar to 1.4 fold) were seen in primary fibroblasts which probably protect the cells from functional damage. In case of primary liver cells: increased levels of SOD (similar to 1.4 fold) and GSH (similar to 1.1 fold) as compared to unexposed cells were observed. Caspase-3 activity assay indicated that the SNP concentrations required for the onset of apoptosis were found to be much lower (3.12 mu g/mL in primary fibroblasts, 12.5 mu g/mL in primary liver cells) than the necrotic concentration (100 mu g/mL in primary fibroblasts, 500 mu g/mL in primary liver cells). These observations were confirmed by CLSM studies by exposure of cells to 1/2 IC50 SNP (resulting in apoptosis) and 2xIC(50)) cells (resulting in necrosis). These results clearly suggest that although silver nanoparticles seem to enter the eukaryotic cells, cellular antioxidant mechanisms protect the cells from possible oxidative damage. This property, in conjunction with the finding that primary cells possess much higher SNP tolerance than the concentration in the gel (similar to 20 mu g/g), indicates preliminary safety of the formulation and warrants further study for possible human application. (C) 2009 Elsevier Inc. All rights reserved.