N-benzyl-N-methyldecan-1-amine, derived from garlic, and its derivative alleviate 2,4-dinitrochlorobenzene-induced atopic dermatitis-like skin lesions in mice

Abstract

Given the intricate etiology and pathogenesis of atopic dermatitis (AD), the complete cure of AD remains challenging. This study aimed to investigate if topically applying N-benzyl-N-methyldecan-1-amine (BMDA), derived from garlic, and its derivative [decyl-(4-methoxy-benzyl)-methyl-1-amine] (DMMA) could effectively alleviate AD-like skin lesions in 2,4-dinitrochlorobenzene (DNCB)-treated mice. Administering these compounds to the irritated skin of DNCB-treated mice significantly reduced swelling, rash, and excoriation severity, alongside a corresponding decrease in inflamed epidermis and dermis. Moreover, they inhibited spleen and lymph node enlargement and showed fewer infiltrated mast cells in the epidermis and dermis through toluidine-blue staining. Additionally, they led to a lower IgE titer in mouse sera as determined by ELISA, compared to vehicle treatment. Analyzing skin tissue from the mice revealed decreased transcript levels of inflammatory cytokines (TNF-α, IL-1β, and IL-6), IL-4, iNOS, and COX-2, compared to control mice. Simultaneously, the compounds impeded the activation of inflammation-related signaling molecules such as JNK, p38 MAPK, and NF-κB in the mouse skin. In summary, these findings suggest that BMDA and DMMA hold the potential to be developed as a novel treatment for healing inflammatory AD.

Keywords: 2,4-Dinitrochlorobenzene; Atopic dermatitis; IgE titer; Inflammation; Mast cells; N-benzyl-N-methyldecan-1-amine.

Figure 1

Topical application of BMDA or DMMA reduces the severity of AD-like skin lesion in DNCB-treated mice (A) The schedule of BMDA or DMMA treatment in DNCB-treated mice (B-D) Photographs of skin lesions from DNCB-induced Balb/C mice (n = 5 mice per group) treated with BMDA (0.3%), DMMA (0.3%), steroid (0.15%) or vehicle (a base cream) for 14 days. Representative images are shown. Skin thickness from untreated mice was measured using a caliper and served as a basis for comparison with other mouse groups. (***p < 0.001, against vehicle).

Figure 2

Effect of BMDA or DMMA treatment on epidermis and dermis thickness in DNCB-induced AD skin lesions (A-C) Skin lesions from DNCB-treated Balb/C mice, applied with BMDA, DMMA, steroid or vehicle for 14 days, were stained with hematoxylin–eosin solution after fixation with 10% formalin and paraffin section. The thickness of the epidermis and dermis from the mice were measured after hematoxylin–eosin staining. (**p < 0.01, ***p < 0.001, against vehicle).

Figure 3

Effect of BMDA or DMMA administration on lymphoid organs in DNCB-treated mice (A-C) Spleen and inguinal lymph node sizes and weights were measured in DNCB-treated Balb/C mice, applied with BMDA, DMMA, steroid, or vehicle for 14 days. The spleen and inguinal lymph nodes were demounted for the measurements. (*p < 0.05, ***p < 0.001, against vehicle).

Figure 4

Effect of BMDA or DMMA application on allergic reaction in DNCB-treated mice (A–D) Skin lesions from DNCB-treated Balb/C mice, applied with BMDA, DMMA, steroid, or vehicle for 14 days, were stained with toluidine-blue solution after fixation with 10% formalin and paraffin section. The number of infiltrating mast cells into the epidermis and dermis was counted from five randomly selected fields under microscopy. The scale bars indicate 100 µm. Additionally, blood samples were taken from the DNCB-treated Balb/C mice, applied with BMDA, DMMA, steroid, or vehicle for 14 days, and IgE titers in the sera were thereafter measured using its specific ELISA. (***p < 0.001, against vehicle).

Figure 5

Effect of BMDA or DMMA regimen on transcript levels of inflammatory cytokines and mediator proteins in the skin from DNCB-treated mice (A–F) Transcript levels of inflammatory cytokines (TNF-1α, IL-1β and IL-6), IL-4, and mediator proteins (iNOS and COX2) were measured by qRT-PCR from the skin tissues of the mice. (***p < 0.001, against vehicle).

Figure 6

Effect of BMDA or DMMA treatment on signaling molecules involved in inflammation in the skin from DNCB-treated mice (A, B) Protein lysates from the skin tissues of DNCB-treated Balb/C mice, applied with BMDA DMMA, steroid, or vehicle for 14 days, were prepared. The phosphorylation levels of signaling molecules involved in inflammation were analyzed with immunoblotting using their specific antibodies. Phosphorylation sites of JNK indicate Thr183 and Tyr185 residues in JNK. Phosphorylation sites of p38MAPK denote Thr180 and Tyr182 residues in p38MAPK. Phosphorylation site of NF-κB (p65; RelA) designates Ser536 residue in NF-κB. The relative expression of the proteins was analyzed after scanning using AlphaView, version 3.2.2 (Cell Biosciences Inc., Santa Clara, CA, USA). (*p < 0.05, **p < 0.01 and ***p < 0.001, against vehicle).

Figure 7

Efficacy of BMDA or DMMA on DNCB-induced inflammation in the skin of mice DNCB sensitization on the skin induces the activation of JNK, p38 MAPK, and NF-κB, resulting in an increase in the expression of inflammatory cytokines (TNF-α, IL-1β, and IL-6) and mediator proteins (iNOS and COX2). However, treatment with BMDA or DMMA inhibits the phosphorylation of JNK, p38 MAPK, and NF-κB, leading to a downregulation in the transcription of inflammatory cytokines (TNF-α, IL-1β, and IL-6), IL-4, and mediator proteins (iNOS and COX2). Consequently, this diminishes allergic reaction such as an increase of the infiltration of mast cells and IgE titer, ultimately alleviating the severity of swelling, rash, and excoriation induced by DNCB in the skin.