Scaffold-mediated SIRT1 transgene expression enhances wound healing in diabetes

Introduction: Impaired wound healing due to diabetes leads to severe clinical complications such as infection and limb amputation. The objective of this study was to assess whether localized transgene expression of SIRT1 via a citrate-based hydrogel would modulate cell inflammatory and senescence states leading to accelerated dermal wound healing in diabetes.

Materials and

Methods: Poly(polyethyleneglycol citrate co N-isopropylacrylamide) (PPCN) was synthesized via sequential polycondensation and free radical polymerization reactions as previously described. Dermal tissues from diabetic and control mice were homogenized and assayed for protein using BCA. Immunoblotting employed anti-Sirt1 and anti-β actin antibodies. Deacetylase activity was quantified via SAMDI mass spectrometry. Lentiviruses encoding Sirt1, pWPI (blank vector) and Sirt1 mutant (Sirt1 inhibitor) were entrapped in PPCN and their release profile was analyzed using qPCR. Human healthy and diabetic dermal fibroblasts (hHDFs/dHDFs) were transduced with lentiviruses encoding Sirt1, pWPI and Sirt1 mutant. Cell migration was assessed by scratch assay; cell senescence was evaluated using a β-galactosidase assay post hydrogen peroxide treatment. In vivo, full-thickness dermal wounds were created on diabetic mice and treated with PPCN, Sirt1, pWPI, PPCN-Sirt1, PPCN-pWPI and saline, and monitored for closure via digital imaging. Digital images of the wound were taken every 3 days until closure. Histology and immunohistochemistry analyses were conducted on skin samples collected at 3 and 27 days post-wounding.

Results, Conclusions, and Discussions: Sirt1 overexpression significantly enhanced the 8-hour wound closure rates. In hHDFs, rates increased from approximately 20.6% with pWPI to 39.7% with Sirt1 (p < 0.001). In dHDFs, the improvement was even more pronounced, with rates rising from 6.9% in pWPI to 39.1% with Sirt1 (p < 0.0001). Histological examination revealed lower fibrosis and inflammation in PPCN-Sirt1 wounds (2-fold lower than wounds treated with PPCN, p< 0.05), alongside enhanced regeneration of skin structures. In vivo, wound closure rates improved significantly when treated with PPCN-Sirt1 compared to saline, increasing from 27% to 47%, 36% to 76%, and 59% to 95% on days 12, 15, and 18, respectively (p < 0.01), demonstrating accelerated healing. f These findings suggest that localized Sirt1 overexpression could be a promising approach for treating chronic wounds in diabetic patients. Future work will elucidate Sirt1’s role and mechanisms in dermal tissue regeneration.
The findings suggest that localized SIRT1 transgene expression using PPCN hydrogels may be a viable strategy to enhance wound healing in diabetic conditions. By effectively delivering therapeutic genes directly to the wound site, this approach not only accelerates wound closure but also improves the quality of healed tissue, potentially offering a significant advancement in the treatment of diabetic ulcers.

Acknowledgements (Optional):