J. Xu1, J. Hu1, C. Zgheib1, M. M. Hodges1, K. W. Liechty1 1University Of Colorado Anschtuz Medical Campus,Pediatric Surgery,Aurora, CO, USA
Introduction: Impaired diabetic wound healing is associated with abnormal long non-coding RNA GAS5 (Growth Arrest-Specific 5) and chronic inflammation. We have previously shown that lncRNA GAS5 was up-regulated in diabetic wounds, and the persistence of the proinflammatory macrophage (M1) phenotype was mediated partly by GAS5/STAT1 pathway, indicating a potential role for GAS5 in the pathogenesis of diabetic wounds. We hypothesized that inhibition of GAS5 would promote the transition of macrophages from M1 to M2 phenotype and enhance the wound-healing process.
Methods: Db/Db diabetic mice and Db/+ non-diabetic mice were wounded with an 8-mm punch biopsy and the wounds treated with a lentiviral vector containing either the green fluorescent protein (GFP) or sh-GAS5 transgene, an shRNA technology to knock down GAS5. Computerized planimetry was used to measure wound size daily. Loss function of GAS5 in RAW macrophage was achieved by transfection. Real-time PCR was applied to measure genes expression.
Results: Inhibition of GAS5 resulted in a significant increase in the rate of diabetic wound healing, (59% vs 99% in GFP-treated wounds at day 8 after injury; P < 0.002), and also improved the early phase of non-diabetic wound healing. GAS5 inhibition resulted in significantly decreased production of pro-inflammatory cytokines (M1 macrophage marker genes) including iNOS, IL1-beta, TNF-α and IL6 in RAW macrophage.
Conclusion: The relative level of lncRNA GAS5 in the wound plays a key role in the wound-healing response. Alterations in the wound level of GAS5 by inhibition, enhance healing by potentially transition of prolonged M1 macrophage to M2 macrophage in diabetic wounds, leading to decreased production of inflammatory cytokines and inflammation.