Dental implants are commonly used to replace missing teeth. The success of such an implant relies on solid anchorage into the alveolar bone that contains the tooth sockets. But many patients do not have sufficient bone volume to secure the implant and require bone reconstruction before tooth implantation.

The reconstruction procedure involves infilling a bone substitute into the alveolar socket to initiate bone formation. Infection, however, remains a major concern in dental surgery. A barrier membrane can help prevent infection, while also blocking soft-tissue ingrowth as the new bone forms. Now, a research team headed up at the University of Western Australia has fabricated a silver nanoparticle (AgNP)-coated collagen membrane, exploiting the natural anti-bacterial properties of silver to prevent infection (Biomed. Mater. 10.1088/1748-605X/aae15b).

Silver coating

The researchers used CelGro collagen membrane, which is approved for dental guided bone regeneration, and coated it with AgNPs using two low-temperature fabrication methods: sonication and sputtering. Images of the coated membranes showed that the AgNPs were evenly coated on both sides using sonication, but on only one side using sputtering.

Scanning electron microscopy revealed that sonication could accurately deposit AgNPs on the membrane, with higher AgNP concentrations depositing more nanoparticles on collagen fibres. Sputtering, however, was difficult to control and led to large uneven deposition of AgNPs.

To test the membrane’s anti-bacterial properties, the researchers prepared AgNP-coated collagen membranes with different nanoparticle concentrations and placed them on bacterial inoculation plates. After four days, samples fabricated via either sonication or sputtering exhibited excellent anti-bacterial effect against two common strains of bacteria, with maximum effect achieved at a concentration of 1.0 mg/ml.

Next, the team seeded mesenchymal stem cells (which can differentiate into a variety of cell types, including bone cells) on AgNP-coated collagen membranes. After 24 hr in culture, they saw a AgNP-dose dependent decline in cell numbers on sonication-coated samples; however, proliferation rates after day 1 were similar. They note that sputter-coated collagen severely inhibited cell growth — suggesting that this technique is not suitable for coating collagen membranes for cell proliferation.

The researchers also assessed the cell membrane integrity using an LDH leakage assay. After 24 hr, they saw an increase in the amount of leaked LDH, correlating to the concentration of AgNP on the membrane. They noted a significant increase between the 1.0 and 1.2 mg/ml sonication groups, indicating that AgNPs can damage the cell membrane.

To maximize antibacterial effectiveness while minimizing cytotoxicity, the team chose 1.0 mg/ml AgNP as the optimal coating concentration. At this dose, confocal imaging showed that cells seeded on AgNP-coated collagen membrane were morphologically comparable to cells on uncoated membranes.

Anti-inflammatory capacity

Inflammation during bone reconstruction can result in a less reliable preparation for the tooth implant. To assess the anti-inflammatory effects of AgNP coating, the researchers examined the expression of two inflammatory cytokines, IL-6 and TNF-a, in macrophages seeded on collagen membranes.

When the cells were stimulated to initiate inflammation, IL-6 expression was lower on AgNP-coated than on uncoated membranes, 1 and 2 hr after stimulation; TNF-alpha expression was only suppressed 1 hr after. Released IL-6 and TNF-alpha were further suppressed 2, 4 and 8 hr after stimulation. These findings demonstrate the anti-inflammatory properties of the coated membranes.

Finally, the researchers examined the osteogenic differentiation of mesenchymal stem cells seeded on AgNP-coated collagen membranes. Expression of early osteogenic markers was far higher in cells cultured on AgNP-coated membranes than on uncoated membranes at days 3 and 6. However, there was no significant difference when cells continued to be cultured to day 9.

The authors conclude that the optimized AgNP-coated collagen membrane showed the ability to guide bone regeneration, as well as exhibiting anti-bacterial and anti-inflammatory capacity, with limited cellular toxicity. They emphasize the potential application of such membranes in dental surgery, particularly for alveolar bone augmentation and bone graft integration.