In Vitro Evaluation of Novel Synthetic Benzimidazolium–Chalcone Hybrids: Antioxidant and Regenerative Effects in Diabetic Wound Healing

The prevalence of diabetes mellitus (DM) is increasing daily worldwide. DM patients suffer from numerous complications, including the development of chronic wounds that can lead to amputation. These complications necessitate innovative approaches. As part of these innovative approaches, this study synthesized four chalcone derivatives and evaluated their activities in an in vitro diabetic wound model. Several spectroscopic techniques, including 1 H and 13 C NMR and HR-MS, were used to confirm the structures of the newly synthesized compounds. After investigating the inhibition of α-glucosidase in HDF-1 cells treated with D-glucose (50 mM), MTT tests and scratch tests were performed. Tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β), total antioxidant status (TAS), and procollagen type-1 were analyzed using an ELISA kit. Additionally, proliferation (Ki67), inflammatory (Nuclear factor kappa B; NFκB), and growth factor (Platelet-derived growth factor subunit A; PDGFA) markers in fibroblasts were assessed by immunohistochemistry. All compounds exhibited strong α-glucosidase inhibitory activity, with IC₅₀ values ranging from 1.115 to 1.612 µg/mL. Cytotoxicity analysis demonstrated that all compounds were biocompatible, maintaining over 85% cell viability in HDF-1 cells. Under diabetic conditions, treatment significantly improved cell viability and promoted wound closure, particularly in C4. In addition, the compounds reduced pro-inflammatory cytokines TNF-α and IL-1β while increasing TAS and procollagen type I levels. Immunocytochemical findings revealed enhanced Ki67 and PDGFA expression and decreased NFκB activation in treated groups. Molecular docking analysis supported the experimental findings by demonstrating favorable binding interactions with α-glucosidase. In conclusion, chalcone derivatives-particularly compound C4-promote diabetic wound healing through multitarget mechanisms involving anti-inflammatory, antioxidant, and pro-regenerative effects, highlighting their strong therapeutic potential.