A three-dimensional human skin model to evaluate the inhibition ofStaphylococcus aureusby antimicrobial peptide-functionalized silver carbon nanotubes

Abstract

Objective To investigate the toxicity and antibacterial application of antimicrobial peptide-functionalized silver-coated carbon nanotubes against Staphylococcus infection using a full thickness human three-dimensional skin model. Materials and methods The three-dimensional skin formation on the scaffolds was characterized by electron microscopy and investigation of several skin cell markers by real time–reverse transcriptase polymerase chain reaction. Functionalized silver-coated carbon nanotubes were prepared using carboxylated silver-coated carbon nanotubes with antimicrobial peptides such as TP359, TP226 and TP557. Following the characterization and toxicity evaluation, the antibacterial activity of functionalized silver-coated carbon nanotubes against Staphylococcus aureus was investigated using a bacterial enumeration assay and scanning electron microscopy. For this purpose, a scar on the human three-dimensional skin grown on Alvetex scaffold using keratinocytes and fibroblasts cells was created by taking precaution not to break the scaffold beneath, followed by incubation with 5 µg/mL of functionalized silver-coated carbon nanotubes re-suspended in minimum essential medium for 2 h. Post 2-h incubation, 200 µL of minimum essential medium containing 1 × 104 colony forming units of Staphylococcus aureus were incubated for 2 h. After incubation with bacteria, the colony forming unit/gram (cfu/g) of skin tissue were counted using the plate count assay and the samples were processed for scanning electron microscopy analysis. Results MTT assay revealed no toxicity of functionalized silver-coated carbon nanotubes to the skin cells such as keratinocytes and fibroblasts at 5 µg/mL with 98% cell viability. The bacterial count increased from 104 to 108 cfu/g in the non-treated skin model, whereas skin treated with functionalized silver-coated carbon nanotubes showed only a small increase from 104 to 105 cfu/g (1000-fold viable cfu difference). Scanning electron microscopy analysis showed the presence of Staphylococcus aureus on the non-treated skin as opposed to the treated skin. Conclusion Thus, our results showed that functionalized silver-coated carbon nanotubes are not only non-toxic, but also help reduce the infection due to their antibacterial activity. These findings will aid in the development of novel antibacterial skin substitutes.

The Advances in Biomedical Applications of Carbon Nanotubes

“…As an alternative to a classic approach based on the use of antibiotics, some researchers proposed to utilize noble metals such as silver and gold in the combination with CNTs [120]. The noble metals, particularly silver, possess the antibacterial and antifungal activity towards various pathogen microorganisms, including antibiotic-resistant strains [121,122].…”

Nanosystems as Vehicles for the Delivery of Antimicrobial Peptides (AMPs)

“…Furthermore, the gene expression profile of indolicidin was different depending on the carrier, the use of CNT presented or activated more pro-inflammatory genes, while the AuNP conjugate activated Il-10, a gene with anti-inflammatory function. Most recently, Chaudhari et al (2019) have analyzed the toxicity and antimicrobial activity of different AMPs (TP359, TP226, and TP557) supported in silver-coated CNTs against Staphylococcus aureus infection using a full thickness human 3D skin model [43].…”

Biofabrication using maize protein: 3D printing using zein formulations

“…We performed assays to evaluate the antimicrobial activity of NA released from 3D-printed zein tablets on Staphylococcus aureus strains. The bacteria were cultured in tryptic soy broth overnight at 37 °C under continuous shaking until the optical density (OD) reached 0.6-0.8 (at 600 nm) [29,30]. Tablet dissolution methods were used to determine the antimicrobial effect of 3D-printed discs according to antimicrobial testing guidelines [31].…”