M. M. Hodges1, C. Zgheib1, J. Hu1, J. Xu1, K. W. Liechty1 1University Of Colorado Denver,Laboratory For Fetal And Regenerative Biology, Department Of Surgery,Aurora, CO, USA
Introduction: In 2014, the United States Center for Disease Control and Prevention (CDC) estimated that 29.1 million Americans (9.3% of Americans) were living with diabetes, with the economic burden of diabetes related wound care estimated as $38.6 billion in 2007. With the global prevalence of diabetes expected to double between 2000 and 2030, there will be an increasing need for innovative technologies to address the commensurate rise in chronic wounds. We have previously demonstrated that diabetic wounds exhibit markedly decreased levels of the anti-inflammatory microRNA miR-146a. Furthermore, we have shown improved healing of diabetic wounds after treatment with a lentiviral construct expressing stromal derived growth factor (SDF-1α). We hypothesize that the improved healing observed in diabetic wounds after SDF-1α treatment is partly due to increased miR-146a expression and the subsequent decrease in inflammation.
Methods: To test our hypothesis we isolated dermal fibroblasts from the skin of 10 week old diabetic (Db/Db) and non-diabetic (Db/+) mice. These fibroblasts were then cultured and transfected with a lentivirus expressing either SDF-1α or green fluorescent protein (GFP) as a control. RNA was extracted from these fibroblasts, and real time PCR analysis was utilized to quantify the gene expression of NFkB, TRAF6, IRAK1, IL-6, MIP2 (IL-8), and miR-146a.
Results: When compared to fibroblasts from the skin of non-diabetic mice, fibroblasts isolated from the skin of diabetic mice demonstrated significantly decreased levels of miR-146a and increased levels of NFkB, TRAF6, IRAK1, IL-6, and MIP2 (IL-8). When transfected with a lentivirus expressing SDF-1α, fibroblasts from the skin of diabetic mice demonstrated significantly increased expression of miR-146a, and significantly decreased expression of NFkB, TRAF6, IRAK1, IL-6, and MIP2 (IL-8).
Conclusions: The significantly increased expression of inflammatory mediators NFkB, TRAF6, IRAK1, IL-6, and MIP2 (IL-8) observed in diabetic fibroblasts is nearly reversed with lenti-SDF-1α transfection. Similarly, the decreased level of miR-146a observed in diabetic fibroblasts is completely reversed when diabetic fibroblasts are transfected with lenti-SDF-1α. The accelerated wound healing previously observed in diabetic skin after lenti-SDF-1α treatment may be due in part to the increased expression of miR-146a and decreased expression of inflammatory mediators NFkB, TRAF6, IRAK1, IL-6, and MIP2 (IL-8). These results support further investigation of the role of SDF-1α in the pathogenesis of diabetic wound healing impairment and as a possible therapeutic target in the treatment of diabetic wounds.