Estimation of Tissue Homogenate Cytokines and MicroRNAs might Help to Determine Wound Vitality and Dating

Abstract

Aim of the work:To assess the applicability of estimated levels of tumor necrosis factor-α (TNF-α) and interleukin (IL)-6, and gene expression levels of microRNAs (MiR)92a and MiR-214a in tissue homogenate (TH) of skin biopsies harvested from wound for discrimination between antemortem and post-mortem wounds and to suggest the post-injury interval (PII). Material and methods:A 2-cm skin incision was made under anesthesia and full thickness punches were obtained from wound edge immediately (C-group) and at 30min, 2-h, 6-h and 24-h after wounding in living animals (L-group) or animals were decapitated immediately after wounding and biopsies were obtained at the same periods after decapitation (D-group). Tissues were homogenized to be used for ELISA estimation of TNF-α and IL-6 levels and qRT-PCR expression levels of MiR-92a and MiR-214a.Results: TNF-α, IL-6 and MiR-92a levels were significantly higher in L-group than other groups. Estimated TNF-α and IL-6 levels showed biphasic increases at 30-min and 2h, respectively and at 24h for both, while the peak levels of MiR -214a and MiR -92a were at 2h and 6h, respectively. MicroRNAs levels showed non-significant differences between all D-group specimens. Regression analysis defined high IL-6 levels as the significant variate to identify PII as either 2h or 24h and high levels of MiR-214a could suggest PII of 2h, while high levels of MiR-92a and TNF-α as the significant variate to suggest PII of 30-min and 6h, respectively. Multivariate analysis defined high IL-6 as the persistently significant predictor for victim's vitality at wounding, while ROC curve analysis defined high MiR-214a levels as the sensitive identifier for victim's viability during wounding. Conclusion:Estimation of expression levels of MiR-92a and MiR-214a in TH might define the probable PII and differentiate antemortem from postmortem wounds, respectively. However, estimated TH levels of TNF-α and IL-6 alone are undependable for provision of knowledge about vitality and timing of wound, so combined markers might increase the accuracy of wound-dating.