Introduction
The antimicrobial properties of silver are well known. Silver nanoparticles (AgNPs) are increasingly being investigated as a non-antibiotic alternative for bone grafting. This research investigated the potential cytotoxicity and intra-cellular uptake and gene expression of bone related cells exposed to AgNPs.
Methods
Lipoic-capped-AgNPs were synthesised and assessed via inductively coupled plasma-mass spectrometry and TEM. Osteoblast (Saos-2) and osteoclast (RAW 264.7) cells were tested with AgNPs from 0.0225-50 μg/ml with controls of carrier only, chlorhexidine digluconate and silver nitrate for cell viability and IC50’s calculated. TEM was conducted without heavy metal staining to preserve the AgNPs. Gene expression assays with qRT2-PCR were undertaken on osteoblasts for stress-related genes and a STRING analysis presented.
Results
AgNPs were generated with a mean hydrodynamic size of 7.5 nm. Cell viability assays showed osteoclasts were more susceptible to AgNPs than osteoblasts. TEM found AgNPs both as nano-particles and nano-chain assemblies within the cytosol of cells. AgNPs at 10 μg/ml for 48 h on osteoblasts activated a process of autophagy into vacuoles and autolysosomes. There were 28 significantly regulated genes with HMOX1 (86-fold upregulation) as a hub gene, and the only gene regulated at 4 h with 1 μg/ml of AgNPs.
Conclusion
AgNPs on osteoblasts and osteoclasts causes cytotoxic effects with particles sequestered within cells via autophagy. The gene expression prolife suggests the osteoblasts work to suppress reactive oxygen species levels as a survival mechanism and that autophagic cell death is active as a response to environmental stress. HMOX1 acts as a protective anti-apoptotic protein and its early regulation indicates cells rapidly activate protective pathways. Osteoclasts were particularly suspectable to AgNPs, which they internalise as nano-chains or aggregates. AgNPs differential effect on bone cells may provide an antimicrobial environment with preference to osteoblasts over osteoclasts.