J. Xu1,2, C. Zgheib1,2, J. Hu1,2, K. W. Liechty1,2 1University Of Colorado Denver,Surgery,Aurora, CO, USA 2Children’s Hospital Colorado,Pediatric Surgery,Aurora, CO, USA
Introduction: Impaired wound healing represents a significant complication of diabetes. The etiology of this wound healing impairment is multifactorial and includes an increased and chronic inflammatory response. We have previously shown that murine diabetic wounds are deficient in SDF-1α and that treatment of diabetic wounds with SDF-1α can improve healing, however, the mechanisms of this correction are not well characterized. MicroRNAs (miRNAs) are novel RNA molecules that regulate the translation of mRNAs at the post-transcriptional level. In particular, microRNA-146a (miR-146a) has been shown to be a key repressor of the inflammatory response by targeting interleukin-1 receptor associated kinease (IRAK1) and tumor necrosis factor receptor associated factor 6 (TRAF6) and repressing NFκB. We have recently shown that miR-146a expression is decreased in murine diabetic wounds. We hypothesized that the improved diabetic wound healing following treatment with SDF-1α is due, in part, to correction of the dysregulated miR-146a gene expression resulting in decreased inflammatory response.
Methods: To test this hypothesis, 8mm full-thickness wounds were created on the flank of diabetic (Db/Db) and non-diabetic (Db/+) mice with a dermal punch instrument. At the time of wounding, the wounds were treated with an intradermal injection with either 108 plaque-forming units (PFU) of a lentivirus containing the SDF-1α or GFP transgene. The wounds were harvested 7days after injury and processed for histology and isolation of total cellular RNA. In order to examine the role of the fibroblast in the dysregulation of the inflammatory response, we isolated diabetic and non-diabetic dermal fibroblasts and treated them with either 106 PFU of a lentivirus containing the SDF-1α or GFP transgene. Gene expression was analyzed using Real-time PCR analysis.
Results: At 7 days, diabetic wounds treated with lenti-SDF-1α exhibited a significant decrease in wound surface area compared to lenti-GFP treated wounds. Diabetic wounds treated with SDF-1α also demonstrated a significant increase in miR-146a expression, and a significant decrease in the miR-146a targets IRAK-1 and TRAF-6, compared to GFP treated diabetic wounds. At the cellular level, MiR-146a was significantly down-regulated in diabetic fibroblasts at baseline, compared to non-diabetic fibroblasts. In addition, SDF-1α treatment corrected the miR-146a gene expression to levels similar to the non-diabetic fibroblast.
Conclusion: SDF-1α mediated correction of the diabetic wound healing impairment is due, in part, to correction of dysregulated microRNA-146a expression. Increased expression of miR-146a may result in decreased inflammatory response and less oxidative stress resulting in improved healing. Further studies are needed to define the role of abnormal miRNA regulation in the pathogenesis of the diabetic wound healing impairment.