Controlled Release of Thymosin Beta 4 Using a Collagen–Chitosan Sponge Scaffold Augments Cutaneous Wound Healing and Increases Angiogenesis in Diabetic Rats with Hindlimb Ischemia

It is important to establish an efficient vascularization for the long-term acceptance of bioengineered skin equivalents treating the cutaneous wounds of diabetic rats with hindlimb ischemia. This study investigates the possible use of a collagen-chitosan sponge scaffold encapsulated with thymosin beta 4 (CCSS-eTβ4), an angiogenic factor, to accelerate cutaneous wound healing in streptozotocin (STZ)-induced diabetic rats with hindlimb ischemia. CCSSs-eTβ4 was fabricated using a freeze-drying method. The scaffolds were analyzed by scanning electron microscopy, swelling and degradation assays, mechanical properties, and scaffolds of 50:50 collagen-chitosan were selected and applied. The controlled release of Tβ4 from the scaffolds elicited localized and prolonged effects over 12 days, as shown by an enzyme-linked immunosorbent assay (ELISA). In vivo, CCSSs-eTβ4 improved diabetic cutaneous wound healing, with faster wound reepithelialization, better dermal reorganization, and higher wound vascularization. Furthermore, CCSSs-eTβ4 downregulated inflammatory genes and upregulated angiogenic genes in the wound tissue. Significant increases in CD31-positive endothelial cells and new vessel density were also observed. In vitro, Tβ4 increased the migratory and proliferative activity of high glucose (HG)-treated human umbilical vein endothelial cells (HUVECs). Meanwhile, we found that Tβ4 could promote HG-treated HUVECs migration and improve angiogenesis by activation of the VEGF/AKT pathway. Overall, these findings demonstrated the promising potential of CCSSs-eTβ4 to promote more effective wound healing and suggest its possible application for diabetic cutaneous wound treatment.