Next-Generation ‘Smart’ Diamond-Silk Dressings for Early Monitoring of Infection and Healing Progression in Burn Wounds

<b>Motivation:</b> According to the World Health Organization, more than 300,000 deaths occur each year as a consequence of burn injuries¹. Each year in Australia, over 8000 patients are hospitalised for burn injuries, with acute treatment costing over $106M¹. Assessing wound repair requires removal of the dressings, which is a painful, time-consuming process based on subjective visual observation of the wound, and is disruptive to healing. Hence there is a critical need to improve this process as wound monitoring has a key role in identifying progress or deterioration for optimal patient management.<br/><b>Biomarkers of interest:</b> Temperature and pH of wound surface are two vital indicators of wound healing and progression^2,3. A clinically diagnosed wound infection elevates the temperature by +3° C to +5° C compared to the healthy skin². The pH of wound fluids is found to rise³ prior to the onset of more vital signs of local infection. Strongly alkaline pH at the wound surface is known to enable the invasion of microorganisms into the tissue layers below the dermis. In contrast an acidic wound environment aids healing through protease activity, increased oxygen release and reduction in microbial growth.<br/><b>Aim</b>: Our work aims to develop a smart and biocompatible silk dressing, integrated with temperature and pH sensors, capable of monitoring early signs of infections via non-invasive light-based measurements.<br/><b>Materials and methods </b><br/><u>Diamond particles for temperature sensing -</u> The nitrogen vaccancy centre (NV) in diamond is highly sensitive to temperature due to changes in its electron spin energy levels. Diamond micro and nanoparticles with NV centres can be used as photostable, biocompatible and non-invasive sensor of temperature changes. The temperature changes (down to millikelvin resolution) can be detected via fluorescence using the optically detected magnetic resonance technique⁴.<br/><u>Organic fluorophore 5,6-carboxynapthofluorescein</u> (CNF) can measure pH within a wound-relevant pH window of 6.5-8.0, with a high sensitivity of 0.2⁵. CNF is a ratiometric pH sensor and its emission spectrum changes with respect to pH, providing a measurement technique that is robust even when the intensity of the fluorescence varies significantly. The ratio of the emission peaks around 550nm (green) and 680 nm (red) is a function of pH.<br/><u>Optically transparent silk</u> fibroin performs the crucial role of encapsulating the two sensors whilst preserving their sensing capabilities and preventing their release (uptake) into the healthy cells and tissues. Transparency of silk dressings will also enable non-invasive optical coherence tomography imaging to assess wound repair, scar formation and healing progression.<br/><b>Results:</b> We have obtained preliminary results indicating that silk membranes embedded with nanodiamond sensors are able to monitor temperature changes in the wound relevant window of 32-40 °C⁴. The <i>in vitro</i> tests revealed that the surface of the hybrid diamond-silk membranes enables eukaryotic cells attachment and promotes their growth. When applied to an <i>in vivo</i> wound model, the hybrid membranes enabled healing and the presence of nanodiamonds did not cause adverse effects on wound healing and closure. The hybrid membranes were found to be highly biocidal towards major skin wound infecting bacteria, P. aeruginosa and E. coli. We have also established that the pH-sensitive fluorophore CNF can be encapsulated within a silk matrix to provide <i>in vivo </i>measurements of acidity or basicity in vivo⁵. Hence hybrid sensor embedded silk dressings can be used as a multifunctional platform for biosensing as well as provide a network of fibres that supports a healthy wound healing process.<br/>[1] Australian Institute of Health and Welfare 2019.<br/>[2] A Chanmugam, et al, Advances in Skin & Wound Care, 30 (9), 2017.<br/>[3] S Ono, et al, Burns,<i> 41 </i>(4), 2015.<br/>[4] A Khalid, et al, ACS Applied Materials & Interfaces, 12(43), 2020.<br/>[5] A Khalid, et al, Sensor and actuators B. Chemicals, 311, 2020.