Estimation of vitality of skin injuries by immunohistochemistry using TNF-α

Introduction. Estimation of wound vitality is one of the most challenging and urgent tasks in forensic medicine, having undeniable significance. Conventional histological methods, based on the assessment of inflammatory response elements, are often insufficiently sensitive for diagnosing injuries inflicted shortly before death. The development and implementation of highly sensitive molecular morphological methods is an important direction for advancing expert practice. Aim. To evaluate the diagnostic value of such immunohistochemical marker as tumor necrosis factor-alpha (TNF-α) and the method of quantitative digital analysis for the objective determination of the vitality of skin blunt force injuries. Materials and methods. Skin samples from 14 cadavers with documented antemortem injuries (28 sections, 140 fields of view) were analyzed. Samples were taken from the area of external injury, and a control tissue sample was also taken from an undamaged area. Paraffin-embedded sections were stained with monoclonal antibodies to TNF-α. Digital images were analyzed using Fiji software using color deconvolution to isolate DAB staining and subsequently determine the area of marker expression in the epidermis and dermis separately. Results. A statistically significant increase in the area of TNF-α expression was found in the damaged area compared to intact skin in control samples. In the epidermis, the difference in medians was –16,66% (p = 0,0002), and in the dermis –7,93% (p = 0,0072). Conclusion. Immunohistochemical detection of TNF-α in combination with quantitative analysis of digital image is highly informative and objective method for ascertaining the skin wound vitality. This approach allows for the identification of early molecular signs of alteration preceding morphological changes and minimizes the subjective factor during the interpretation of results, which increases the evidentiary value of the forensic medical report.