(E-197) Spectroscopic assessment of burn depth in a live pig model of second and third-degree burns

Introduction:: Burn-related injuries led to roughly 371,000 emergency department visits in the US in 2020 [1]. Healing can often vary from several days to years depending on the depth and the extent of the injury [2], and deciding the best course of treatment as early as possible is critical. Sub-optimal assessment can lead to challenges in choosing appropriate treatment and worse outcomes, including slow wound healing, infection, pain, and hypertrophic scarring [3]. Clinical assessment of burns has a low success rate (64-76%) even for experienced surgeons [4] or is dependent on highly invasive and painful biopsies [5]. Dynamic changes in the acute healing phase may further complicate assessment to guide a potential surgical approach [6]. Thus, there is an urgent need to develop a rapid, non-invasive, and objective metric of assessment that is easy to implement in the clinic. An ideal method does not destroy the sample, such as in the case of biopsies, thus allowing frequent wound sampling in the acute phase. Early and accurate assessment may help treatment decisions and reduce the duration of hospitalization and costs in addition to improved outcomes.

We propose a fluorescence spectroscopy-based method for the assessment of burn wounds. A non-contact, non-destructive, benchtop device is developed that utilizes a 441 nm laser for excitation and uses a spectrometer to measure the fluorescence emission of the wound in < 3 minutes. In the current study, we test this device in a novel model of burns to evaluate efficacy in diagnosing second vs. third-degree burns.

Materials and Methods:: We used Yucatan pigs to create a model of burn injuries due to similarities with human skin [7], [8]. Brass blocks of 4cm radius were used to create contact burns by preheating to three different temperatures - 21°C for sham burns, 63°C for 2nd-degree burns (partial thickness), and 95°C for 3^rd-degree burns (full thickness) as described before for the miniature swine model of burns [9]. Burns were created in a pattern of four burns per row in two rows for eight burns (3% Body Surface Area or BSA) evenly distributed between both sides of the pigs in the dorsal paravertebral region, as shown in Figure 1A. The eight locations were randomly assigned to either sham, second-degree, or third-degree categories to avoid any potential differences in healing due to the location of wounds. In full-thickness burns, debridement was performed 4 days after the burn injury. The animal was followed for 8 weeks which is considered sufficient for the long-term assessment. Each wound was assessed using a bench-top device immediately (< 2 hours) after and 4 days after the initial injury. This device uses a 441 nm laser pump connected with an optical fiber to a probe used for excitation. This same probe also carries the emission spectrum for analysis and filters the incident wavelength. The resulting spectrum is analyzed for the total fluorescence intensity for correlation with burn depth.

Results, Conclusions, and Discussions:: We were successfully able to distinguish between sham, second, and third-degree burns based on the fluorescence intensity of burns immediately after initial injuries, as seen in Figure 1B. The fluorescence intensity followed the same trend on the postoperative day (POD) 4, as shown in Figure 1C. The fluorescence intensity from third-degree burns was significantly higher than the second-degree and sham burns. In contrast, fluorescence from second-degree burns was lower than sham burns. A proposed reason for this trend is the combined effect of the contributions of denatured collagen and hemoglobin to the fluorescence intensity spectrum. Denatured collagen in burns is known to have higher autofluorescence compared to healthy collagen, as previously seen in mice models [10]. This is in contrast to hemoglobin which acts as a strong absorber of the 441 nm wavelength and thus lowers the fluorescence intensity. Denatured collagen is the predominant contributor to the overall fluorescence spectrum of third-degree burns as the vasculature is severely disrupted, greatly diminishing hemoglobin content in the wound. Both denatured collagen and hemoglobin, however, contribute to the fluorescence intensity of second-degree burns. Net absorption of the incident photons by higher hemoglobin, when compared to sham burns, was observed because of localized inflammation. This increased inflammation is also confirmed clinically by the erythema/hyperemia, as seen in Figure 1A. Thus, fluorescence intensity can accurately predict the depth of the burn with high confidence early in the wound healing process. A facile quantification of burn depth using our benchtop device has the potential to become a convenient and portable assessment in the clinic in addition to visual observation.


Acknowledgements (Optional): : The authors would like to sincerely thank the Knight Surgery Research Laboratory (KSRL) and the Center for Comparative Medicine (CCM) at Massachusetts General Hospital (MGH) for their support in pig surgeries and housing. The authors would also like to thank the generous funding provided by the Shriners Children's Hospital to support this work.

References (Optional): : 1. National Hospital Ambulatory Medical Care Survey: 2020 Emergency Department Summary Tables (accessed on April 30, 2020, 2023) National Hospital Ambulatory Medical Care Survey: 2020 Emergency Department Summary Tables (cdc.gov)
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