# Improvement Effect of Membrane-Free Stem Cell Extract on
## 4 Discussion  
MFSCE derived from ADSCs was found to have more advantages than ADSCs as a novel functional natural product for various biological applications. [PERSON] et al. [11] reported the protein profiling of MFSCE through nanoscale liquid chromatography coupled to tandem mass spectrometry analysis, demonstrating that MFSCE regulated the integrin pathway proteins, inflammation regulatory proteins, and wound healing proteins. In particular, MFSCE-related proteins have been shown to have anti-inflammatory effects. In this regard, our previous study reported that MFSCE attenuated the production of inflammatory mediators including nitric oxide, cyclooxygenase-2, and prostaglandin E\({}_{2}\) by modulating the nuclear factor-kappa B (NF-kB) and mitogen-activated protein kinases (MAPKs) pathway in LPS/IFN-\(\gamma\)-induced RAW264.7 macrophages [16]. Moreover, [PERSON] et al. [18] indicated that MFSCE suppressed the production of IL-1\(\alpha\) stimulated inflammatory factors, such as inducible nitric oxide synthase (iNOS), cyclooxygenase-2, nitric oxide, and prostaglandin E\({}_{2}\), and downregulated the NF-kB and MAPKs signaling pathways in rat chondrocytes. [PERSON] et al. [19] demonstrated that MFSCE-treated keratinocytes and fibroblasts inhibited inflammation and wrinkle formation. AD is a chronic and serious  
Figure 7: Effect of MFSCE on IL-4 (**A**), IL-10 (**B**), IFN-\(\gamma\) (**C**), TNF-\(\alpha\) (**D**), and TARC (**E**) expression in AD-like dorsal skin of HDM-sensitized NC/Nga mice. Values are mean \(\pm\) SD. \({}^{\text{a--e}}\) Means with different letters are significantly different (\(p<0.05\)) as determined by [PERSON]’s multiple range test. The mice were grouped and treated as described in Figure 2.  
inflammatory skin disease; therefore, in the present study, the beneficial effects of MFSCE with excellent anti-inflammatory effects were investigated in HDM-induced, AD-like NC/Nga mice. To explore the protective and improvement effects on AD, an animal model of human symptom-like AD is required. [PERSON] et al. [20] demonstrated that the clinical symptoms and histopathological research data in NC/Nga mice were similar to those in human AD. In addition, NC/Nga mice spontaneously hyperproduce serum IgE influenced by environmental factors, particularly HDM allergens, which are highly sensitized as an environmental factor to human AD [21]. These observations are associated with the clinical symptoms of human AD [22]. Here, NC/Nga mice are a relevant model for human AD, as numerous studies have used them to investigate the pathophysiology and treatment of AD using natural products. In addition, a well-known contributor among HDM species is _Dermatophagoides farinae_ (Df). For several decades, it is considered to be a common cause of AD by their body and feces as environmental air-born allergens [23]. Therefore, Dfe used in present study played a role as a representative material of HDM, triggering skin barrier dysfunction in NC/Nga mice. [PERSON] et al. [24] demonstrated the ameliorative effects of _Gardenia jasminoides_ extract, a natural herbal medicine, in an Dfe-induced AD model in NC/Nga mice. [PERSON] et al. [25] confirmed that cacao beans from _Thebrooma cacao_, a source of flavonoids, showed strong anti-AD effects on Dfe-stimulated, AD-like symptoms in NC/Nga mice. Therefore, this study used HDM-sensitized NC/Nga mice to demonstrate a clinical, systemic, and topical improvement effect of MFSCE on AD-like skin inflammation. Clinically, patients with AD experience skin inflammation characterized by erythema, dry skin, edema, excoriation, erosion, lichenification, and scratching [26]. Thus, the alleviation of AD-like symptoms is especially important in improving the quality of life for patients with AD. Our results demonstrate that repeated dermal application of MFSCE ameliorated dermatitis scores and scratching frequency in HDM-stimulated NC/Nga mice.