In this study, we found that re-epithelialized skin after partial-thickness burns was thicker and had lower ultrasonographic echogenicity than normal skin and that an LEB was present, as in photoaging skin. We also found that the thickness of the LEB was closely correlated to healing time and was associated with the type of scar.
The water content of the dermis is principally regulated by the pressure of the interstitial fluid, which, in turn, is determined by the distensibility of the collagen network and the water-holding capacity of glycosaminoglycans [12, 14]. Comparative studies of ultrasonography and nuclear magnetic resonance spectroscopy showed that dermal water content can be directly correlated with dermal echogenicity [15]. Greater fluid content in tissue results in a decrease in echogenicity readily detectable by ultrasound [16]. In acute burn wounds, vasoactive mediators, such as prostaglandins, histamine, and bradykinin, can lead to oedema by altering endothelial cell and basement membrane function to increase permeability [17, 18]. In this study, we found that dermal echogenicity was decreased and that skin thickness was increased in re-epithelialized skin after partial-thickness burns compared to those in contralateral normal skin, which is thought to be the effect of oedema. This difference tended to be prominent when the healing time was long and decreased with wound maturation over time (data not shown).
In the wound-healing process, granulation tissue, comprising procollagen, elastin, proteoglycans, and hyaluronic acid, allows the ingrowth of new blood vessels [19, 20]. The extracellular matrix is immature, with fine bundles of collagen in a predominantly linear (horizontal) organization and an increased number of blood vessels [21]. Therefore, newly organized tissue during the healing process is highly hydrated and shows very low echogenicity on ultrasound images.
Hoffmann et al. [22] described the poor internal echoes in lesions after cryosurgery at the end of the healing process as associated with newly formed fine-fibrillary connective tissue (granulation tissue). Rippon et al. [23] analysed ultrasound scans of wounds and showed that early granulation tissue was echo-poor because of the low echogenicity of cellular infiltrates. The researchers also concluded that ultrasound can be used to visualize and quantify fibrous granulation tissue accumulation in wounds. In a human wound model, Dunkin et al. [11] demonstrated that the wound was represented as an area of echolucency in the dermis. In their study, the thickness of the low-echogenic area produced by wounds varied depending on the injury depth.
In the present study, we also found a low-echogenic area in the re-epithelialized skin after partial-thickness burns. Because most burn injuries cause surface damage, this low-echogenic area takes the form of a ‘band’ similar to the ‘subepidermal LEB’ observed in the ultrasound of photoaging skin [12,13,14].
This ultrasonographic LEB is found in all lesions, even in wounds with a healing time of fewer than 10 days. This band was thicker and more significant with longer healing time. Because each patient and each body site have variable skin thickness, we used the proportion of LEB thickness among the total skin thickness to assess the correlation with healing time, and a strong correlation was observed. is influenced not only by the depth of the wound but also by various factors; however, when the wound is deeper, the healing time becomes longer and the newly organized tissue becomes thicker, which can be observed in the form of an LEB through ultrasound. Based on these results, we suggest that ultrasound can be used to retrospectively estimate the depth wounds with uncertain clinical information.
Wound tissues change over time during the wound-healing process, so timing of the ultrasound evaluation may affect the result. Usually, hypertrophic changes begin 1 to 2 months after re-epithelialization. To assess the condition before hypertrophic change, we selected ROIs evaluated before 14 days after re-epithelialization. The time interval from the injury date to ultrasound was the sum of the healing time and the time interval from re-epithelialized date to ultrasound. Because a deviation in the time interval from the re-epithelialized date had relatively low deviation, the time interval from injury date was more affected by the healing time. Therefore, the proportion of LEB thickness was inevitably correlated to the time interval from the injury date as well as the healing time. In addition, the time interval from the injury date was statistically significant only in the univariate analysis, not in the multivariate analysis.
In human burns, delayed wound healing is a significant risk factor for hypertrophic scarring [1], and healing time is an important factor in the prediction of scarring and the selection of scar prevention methods [2, 3]. Because of the strong relationship between the proportion of LEB thickness and healing time, we hypothesize that the proportion of LEB thickness is also a useful prognostic factor in the prediction of scarring. In the univariate logistic regression analysis, both the healing time and the proportion of LEB thickness were statistically significant factors affecting scar status. The proportion of LEB thickness was the only statistically significant factor in the multivariate model (Table 2). These results indicate that the proportion of LEB thickness of post-burn skin may have a prognostic value for scarring similar to that of healing time.
In this study, the LEB thickness was closely related to the healing time associated with wound depth. The LEB thickness is expected to be related to the wound depth. Thin LEB lesions without hypertrophy may be limited to papillary dermal injury, and thick LEB lesions with hypertrophy may include reticular dermal injury. However, 22 MHz ultrasound cannot distinguish between the papillary dermis and the reticular dermis. Ultrasonography with a higher MHz is required for more detailed inspection.
The ultrasound examination, which is reproducible and valid, results in no damage to the wound and allows serial scans to be obtained, allowing temporal changes within the wound to be monitored [8]. In objective scar assessment, the ultrasound technique is an effective modality for measuring scar thickness [5]. Ultrasonography provides objective and quantitative information with which to identify the current state of the scar and to evaluate treatment results [9]. Ultrasound examination of early re-epithelialized skin after partial-thickness burns is valuable not only for the estimation of scarring but also for the serial objective scar assessment to observe the progress of the scar.
Clinical practice to prevent hypertrophic scars requires great effort and cost. For effective clinical practice, it is necessary to select the wound to which such efforts and costs apply. Early ultrasound evaluation of re-epithelialized skin after partial-thickness burns can inform the potential to form a hypertrophic scar and help to determine the management strategy. Re-epithelialized skin after partial-thickness burns with thick LEB in early ultrasound evaluation may be associated with a longer healing time and higher scar potential and need a strict scar prevention strategy. In a human model study, Dunkin et al. [11] reported that the mean threshold depth to leave a scar on the lateral aspect of the hip was 33.1% of total skin thickness. In our data, the mean value of the proportion of LEB thickness in the hypertrophic scar group was 29.97% (SD=4.30). In our centre, a strict scar management protocol is recommended for re-epithelialized skin after partial-thickness burns with a proportion of LEB thickness above 25%.
The proposed ultrasound device operates at a frequency of 22 MHz. Agabalyan et al. [24] showed that a 20 MHz probe could not provide sufficient penetration and information to evaluate scar thickness. These authors recommended using a transducer set between 5 and 10 MHz for measuring deeper dermal thickness. In our experience, the penetration thickness of the 22 MHz probe that we used was limited to 3 mm although the manufacturer described between 6 and 8 mm. In this study, however, our ROI was the recently re-epithelialized skin; therefore, it was sufficient to use the 22 MHz probe rather than the hypertrophic scar.
The ultrasound examination was simple to set up and use and required little training to operate. The time taken to record a scan was very short. We also performed the ultrasound examinations on an outpatient basis. A typical scan took only approximately 1 min and caused no discomfort to the patient. These scans allow us to observe the status of re-epithelialized skin after a burn and determine the most cost-effective management plan for scar treatment. In addition, the scanning results provide a computerized two dimensional (2D) image that is intuitive and understandable to the patient in real time, which can be an effective way to explain the patient’s current condition and treatment plan.
This study is a retrospective analysis. Because the data on healing time were obtained through medical records and the ROI was set manually, it is possible that information bias exists. Another possible limitation is that the scar status was evaluated by dichotomy, and other factors, such as erythema and pliability, were not considered. There was no control for other factors (e.g., wound location, sex, or age) that could affect skin thickness and echogenicity or for other factors that could affect scar formation. Further studies are needed to evaluate the prognostic value of skin ultrasonography after re-epithelialization of burn injuries.