Bacteriostatic SiONx Coating Regulates Osteogenic Capacity via NRF2 Antioxidant Activity

Objectives: Compromised and complicated bone defects often have prolonged and delayed healing due to lack of bioactivity of the fixative device, secondary infections, and associated oxidative stress. Here, we propose amorphous silicon oxynitride (SiONx) as coatings for the fixative devices to improve bioactivity, bacteriostatic activity, and reduce oxidative stress. We aim to study the effect of increasing N/O atom ratio in the SiONx to fine tune the cellular activity and the antioxidant effect via NRF2 pathway under oxidative stress conditions.
Methods: 5 compositions of SiONx were fabricated as thin-film coatings on implant surfaces by changing N/O ratio. In vitro studies involved human mesenchymal stem cells (MSC) to study the effect of SiONx coatings on osteogenesis with and without toxic oxidative stress condition. Additionally, bacterial growth was studied on SiONx surfaces using Methicillin-Resistant Staphylococcus Aureus (MRSA) colonies. We evaluated the samples for cell proliferation, osteogenic differentiation, and antioxidant activity. Studies were conducted for 1-7 days, and all studies were repeated to yield n = 9 /group. NRF2 siRNA transfection was performed for the hMSCs (NRF2-KD) to study the antioxidant response to silicon ions.
Results: SiONx implant surfaces showed >4-fold decrease in bacterial growth vs control. Increasing N/O ratio in SiONx implants increased alkaline phosphatase activity >1.5 times and > 2-fold for other osteogenic markers (Osteocalcin, RUNX2, and Osterix) under normal conditions. Increasing N/O ratio in SiONx increased protective effects and improved cell viability against toxic oxidative stress conditions. There was a significant increase in osteocalcin activity compared to the uncoated group along with increased antioxidant activity, under oxidative stress conditions. In NRF2-KD cells, there was a stunted effect on the upregulation of antioxidant markers with silicon ions, indicating the role of NRF2.
Conclusions: SiONx coatings displayed bacteriostatic properties and promoted osteogenic markers under oxidative stress by enhancing antioxidant NRF2 activity that induced ROS reduction and robust osteogenic marker expression leading to rapid bone regeneration. These results indicate the potential for inducing in vivo bone regeneration in a challenging oxidative stress environment.