Effects of Angelica gigas Nakai as an Anti-Inflammatory Agent in In Vitro and In Vivo Atopic Dermatitis Models

Abstract

We investigated the cellular and molecular mechanisms mediating the effects of Angelica gigas Nakai extract (AGNE) through the mitogen-activated protein kinases (MAPKs)/NF-κB pathway using in vitro and in vivo atopic dermatitis (AD) models. We examined the effects of AGNE on the expression of proinflammatory cytokines and chemokines in human mast cell line-1 (HMC-1) cells. Compound 48/80-induced pruritus and 2,4-dinitrochlorobenzene- (DNCB-) induced AD-like skin lesion mouse models were also used to investigate the antiallergic effects of AGNE. AGNE reduced histamine secretion, production of proinflammatory cytokines including interleukin- (IL-) 1β, IL-4, IL-6, IL-8, and IL-10, and expression of cyclooxygenase- (COX-) 2 in HMC-1 cells. Scratching behavior and DNCB-induced AD-like skin lesions were also attenuated by AGNE administration through the reduction of serum IgE, histamine, tumor necrosis factor-α (TNF-α), IL-6 levels, and COX-2 expression in skin tissue from mouse models. Furthermore, these inhibitory effects were mediated by the blockade of the MAPKs and NF-κB pathway. The findings of this study proved that AGNE improves the scratching behavior and atopy symptoms and reduces the activity of various atopy-related mediators in HMC-1 cells and mice model. These results suggest the AGNE has a therapeutic potential in anti-AD.

Figure 1

Effect of AGNE on histamine, IL-1β, IL-4, IL-6, IL-8, IL-10, and TNF-α level in PMACI-stimulated HMC-1 cells. (a) Cells (6 × 10⁵ cells/well) were pretreated with AGNE (0.1, 1, and 10 μg/mL) for 1 h and then PMACI (50 μM PMA + 1 μg/mL CI A23187) for 4 h. Histamine content was measured using a histamine assay kit. (b–g) Cells were pretreated with AGNE for 1 h and then PMACI for 12 h. Levels of inflammatory cytokines were measured using ELISA. Data is mean ± SEM of triplicate determinations from separate triplicate experiments (^∗P < 0.05 versus control and ^#P < 0.05 versus PMACI alone).

Figure 2

Effect of AGNE on serum and tissue levels of IgE, histamine, IL-6, and TNF-α in DNCB-induced mice. Blood and skin tissue samples were collected, and serum levels of (a) IgE, (b) histamine, (c) IL-6, and (d) TNF-α and tissue levels of (e) IL-6 and (f) TNF-α in indicated group were measured using ELISA and assay kit. Data is means ± SEM of triplicate determinations from separate triplicate experiments (^∗P < 0.05 versus control group and ^#P < 0.05 versus DNCB-treated group).

Figure 3

Effect of AGNE on dermatohistopathological changes in mice model. (a) AGNE (10, 20, and 40 mg/kg) was orally administered 1 h before intradermal injection of compound 48/80 (50 μg/kg). Scratching behaviors were counted in ICR mice. (b) BALB/c mice were sensitized with DNCB and induced atopic dermatitis. Terfenadine was used as a positive control. (c) DNCB-induced dorsal skin was stained with H&E. Data is means ± SEM of triplicate determinations from separate triplicate experiments (^∗P < 0.05 versus control group and ^#P < 0.05 versus compound 48/80 or DNCB-treated group).

Figure 4

Effect of AGNE on protein expression in PMACI-induced HMC-1 cells. Cellular proteins were used to detect (a) I-κBα phosphorylation (cytosol) and (b) DNA-binding activity of NF-κB p65 (nuclear), (c) COX-2 activation, and (d) ERK, (e) JNK, and (f) p38 phosphorylation (cellular hole protein) in HMC-1 cells. Data is means ± SEM of triplicate determinations from separate triplicate experiments (^∗P < 0.05 versus control and ^#P < 0.05 versus PMACI alone or DNCB-treated group).

Figure 5

Effect of AGNE on protein expression in DNCB-induced BALB/c mice. Total protein tissue samples were collected to detect (a) COX-2 activation and (b) ERK, (c) JNK, and (d) p38 phosphorylation in mice model. Data is means ± SEM of triplicate determinations from separate triplicate experiments (^∗P < 0.05 versus control and ^#P < 0.05 versus PMACI alone or DNCB-treated group).