Background: Wound age determination is a critical
aspect of forensic investigations. The primary
objective aimed to determine the appropriateness of
vascular endothelial growth factor (VEGF), alphasmooth muscle actin (α-SMA), T tumour necrosis factor-alpha (TNF-α) and
transforming growth factor beta (TGF-β1) biomarkers for wound age determination in
rats. We conducted histopathological, immunohistochemical, and molecular
assessments to investigate changes in these markers over different post-wounding time
intervals. Methods: Forty-eight healthy adult male albino rats were separated into 8
equal groups based on the post-wounding time interval. Histopathological examinations,
immunohistochemical staining, and gene expression analyses were performed on skin
samples to assess the statement of TGF-β1, TNF-α, α-SMA, and VEGF. Data were
analysed statistically to determine the significance of changes over time. Results: The
histopathological analysis revealed distinct changes in skin tissue over time, ranging
from ulceration to reepithelialization and tissue regeneration. Immunohistochemical
examination demonstrated varying levels of VEGF and α-SMA expression, with
marked increases in later post-wounding periods. Molecular analysis indicated a
significant upregulation of TNF-α and TGF-β1 expression, reaching peak levels around
seven days post-wounding. Conclusions: The use of VEGF, α-SMA, TNF-α, and TGFβ1 as potential biomarkers for wound age determination. The expression patterns of
these markers in the rat skin suggest their suitability for forensic applications.