Bee venom acupuncture alleviates trimellitic anhydride-induced atopic dermatitis-like skin lesions in mice

Abstract

Background: Bee venom acupuncture (BVA), a novel type of acupuncture therapy in which purified bee venom is injected into the specific acupuncture point on the diseased part of the body, is used primarily for relieving pain and other musculoskeletal symptoms. In the present study, therapeutic potential of BVA to improve atopic dermatitis, a representative allergic dysfunction, was evaluated in the mouse model of trimellitic anhydride (TMA)-induced skin impairment.

Methods: Mice were treated with 5% TMA on the dorsal flank for sensitization and subsequently treated with 2% TMA on the dorsum of both ears for an additional 12 days after a 3-day interval. From the 7(th) day of 2% TMA treatment, bilateral subcutaneous injection of BV (BV, 0.3 mg/kg) was performed daily at BL40 acupuncture points (located behind the knee) 1 h before 2% TMA treatment for 5 days.

Results: BVA treatment markedly inhibited the expression levels of both T helper cell type 1 (Th1) and Th2 cytokines in ear skin and lymph nodes of TMA-treated mice. Clinical features of AD-like symptoms such as ear skin symptom severity and thickness, inflammation, and lymph node weight were significantly alleviated by BV treatment. BV treatment also inhibited the proliferation and infiltration of T cells, the production of Th1 and Th2 cytokines, and the synthesis of interleukin (IL)-4 and immunoglobulin E (IgE)-typical allergic Th2 responses in blood. The inhibitory effect of BVA was more pronounced at BL40 acupoint than non-acupuncture point located at the base of the tail.

Conclusions: These results indicate that BV injection at specific acupuncture points effectively alleviates AD-like skin lesions by inhibiting inflammatory and allergic responses in a TMA-induced contact hypersensitivity mouse model.

Fig. 1

Experimental schedules for developing TMA-induced atopic dermatitis and treating with BV in mice. TMA, trimellitic anhydride; BV, bee venom

Fig. 2

Representative images (a) of mouse ears and scoring graph (b) in the NOR, AD, AD + BVA, AD + BVNA and AD + PRE groups. The graph indicates time-course severities of atopic dermatitis by scoring each image between 0 and 9 points depending on the following skin symptoms: scaling and dryness, hemorrhage and excoriation, and edema and redness. The arrow indicates the initiation of BV or prednisolon treatment. ^*** p < 0.001 vs. the NOR group; ^## p < 0.01, ^### p < 0.001 vs. the AD group

Fig. 3

Ear thickness (a) and lymph node weights (b) of the mice in the NOR, AD, AD + BVA, AD + BVNA and AD + PRE groups. ^*** p < 0.001 vs. the NOR group; ^# p < 0.05, ^### p < 0.001 vs. the AD group

Fig. 4

Serum levels of IL-4 (a) and IgE (b) in the NOR, AD, AD + BVA, AD + BVNA and AD + PRE groups using an ELISA. IL, interleukin; IgE, immunoglobulin E; ELISA, enzyme-linked immunosorbent assay. ^*** p < 0.001 vs. the NOR group; ^# p < 0.05, ^### p < 0.001 vs. the AD group

Fig. 5

Histological images and graphs indicating relative percentage of CD4- and CD8-immunopositive cells of the ear sections. Ear sections in each group were stained with hematoxylin and eosin (a1-a5), toluidine blue (b1-b5), anti-mouse CD4 IgG (c1-c5) and anti-mouse CD8 IgG (d1-d5). Black scale bar indicates 100 μm (100× magnification). Black thick lines in the images of H-E staining indicate the thickness of ear skins. Small white squares (200×) in the centers of immunohistological staining images (c2 & d2 of AD group) are magnified in the lower left corners to observe CD4- and CD8- positive cells infiltrated into the skin tissues. The representative CD4- and CD8-immunopositive cells were indicated by black arrows in indicated in small white squares in C2 and D2, respectively. The numbers of CD4- and CD8-immunopositive cells in the fixed area of the images are depicted in the bar graphs e and f, respectively, below the histological images. IgG, immunoglobulin G; CD, cluster of differentiation. ^*** p < 0.001 vs. the NOR group; ^# p < 0.05, ^## p < 0.005 and ^### p < 0.001 vs. the AD group

Fig. 6

The mRNA expression levels of IL-1β (a) and IL-4 (b) in ear tissue (E), and TNF-α (c), and IL-4 (d) in auricular lymph node (LN) in mice. IL, interleukin; TNF, tissue necrosis factor. ^** p < 0.01, ^*** p < 0.001 vs. the NOR group; ^# p < 0.05, ^## p < 0.01, ^### p < 0.001 vs. the AD group

Fig. 7

Analysis of the levels of cytokines such as IL-2(a), IL-12(b), IFN-γ(c), IL-5(d), IL-10(e), GM-CSF(f), TNF-α(g) and IL-4(h). The analysis was performed using the Bio-Plex® suspension array system, in the auricular lymph node in mice. GM-CSF, granulocyte macrophage colony-stimulating factor; IFN, interferon; IL, interleukin; TNF, tissue necrosis factor. ^* p < 0.05, ^*** p < 0.001 vs. the NOR group; ^# p < 0.05, ^## p < 0.01, ^### p < 0.001 vs. the AD group