## 4 Discussion  
Wound healing is a complex process involving four overlapping phases: hemostasis, inflammation, proliferation, and remodeling [3]. The dysregulation of these stages leads to delayed wound healing. DEX is one of the most common corticosteroids used to treat inflammatory diseases but has adverse effects on wound healing by delaying the inflammation phase and inhibiting the proliferation phase of this process [5]. In this study, DEX impaired the wound healing process. A treatment with puerarin recovered the impaired wound healing process in both the in vivo and in vitro models. Puerarin is a major isoflavonoid from the root of _Pueraria montana_ var. _lobata_ (Wild.) [PERSON] & [PERSON] [6]. Several studies suggest that isoflavonoids have beneficial effects on the wound healing process [25; 26]. An extract from the flower of _Pueraria thunbergiana_ Benth (synonymous with _Pueraria montana_ var. _lobata_ (Wild.) [PERSON] & [PERSON]), which contains isoflavonoids and essential oil components, has the potential for skin regeneration by stimulating the migration, proliferation, and collagen synthesis of epidermal keratinocytes [10]. A previous study reported that the oral administration of puerarin pro  
Figure 4: Effects of puerarin on the ERK and Akt signaling pathways in HaCaT cells stimulated with DEX. (**A**) HaCaT cells were incubated with DEX (100 nM) and puerarin (1, 10 nM) or _Centella siaatica_ extract (100 mg/mL) for 30 min. Protein expression of p-ERK, p-Akt, ERK, Akt was determined by Western blotting. (**B**) HaCaT cells were pretreated with U0126 (10 nM) or wortmannin (1 nM) for 1 h and then treated with DEX (100 nM) and puerarin (1 nM) for 48 h. The proliferation rate was assessed using BrdU assays. (**C**) HaCaT cells were pretreated with U0126 (10 nM) or wortmannin (1 nM) for 1 h and scratched and treated with dexamethasone (DEX, 100 nM) and puerarin (1 nM). Images of the scratch gap width were captured at 0 and 24 h post-scratching. The wound closure rates were defined as the difference between the wound width at 0 and 24 h. The results are presented as the means \(\pm\) SDs (\(n\) = 3 per experiment). Bars labeled by the same letter on the graph are not significantly different from each other (\(p\) < 0.05) by one-way ANOVA followed by post hoc Tukey’s test. moted wound healing in a diabetic rat model by enhancing the connective tissue and new vessel formation [16]. In the current study, the topical treatment of Puerrin for 14 days also promoted the wound closure process in DEX-treated mice. _Centella asiatica_ is a well-known wound healing agent in both clinical and experimental studies [18; 27]. _Centella asiatica_ used as the positive control drug is known to improve DEX-impaired wound healing ability [20]. In our experiments, the topical treatment of _Centella asiatica_ restored the delayed wound closure by DEX application. The efficacy of _Centella asiatica_ to improve wound healing is related to promoting collagen synthesis, angiogenesis, and keratinocyte migration [17; 28]. Therefore, we investigated whether Puerrin might be related to these mechanisms. Collagen is the essential component of the dermal extracellular matrix (ECM), and collagen deposition in the dermis plays an important role in the wound healing process. Collagen is involved in the hemostasis phase by interacting with platelets in the wound areas [29]. The recruited platelets can release various growth factors, including the transforming growth factor-beta (TGF-\(\beta\)), platelet-derived growth factor (PDGF), and vascular endothelial growth factor (VEGF) [30]. TGF-\(\beta\) can induce fibroblast differentiation and collagen synthesis to promote wound closure and scarring [31]. VEGF is a crucial factor in the wound angiogenesis process as it promotes the proliferation and migration of endothelial cells [32]. Previous studies reported that glucocorticoids impaired wound healing by inhibiting collagen synthesis and angiogenesis in vivo [33; 34].