Ann Dermatol.  2016 Feb;28(1):22-29. 10.5021/ad.2016.28.1.22.

Selective Cannabinoid Receptor-1 Agonists Regulate Mast Cell Activation in an Oxazolone-Induced Atopic Dermatitis Model

• Nam G
• Jeong SK
• Park BM
• Lee SH
• Kim HJ
• Hong SP
• Kim B
• Kim BW

• KMID: 2429478
• DOI: http://doi.org/10.5021/ad.2016.28.1.22

Abstract

BACKGROUND
Many inflammatory mediators, including various cytokines (e.g. interleukins and tumor necrosis factor [TNF]), inflammatory proteases, and histamine are released following mast cell activation. However, the endogenous modulators for mast cell activation and the underlying mechanism have yet to be elucidated. Endogenous cannabinoids such as palmitoylethanolamide (PEA) and N-arachidonoylethanolamine (anandamide or AEA), were found in peripheral tissues and have been proposed to possess autacoid activity, implying that cannabinoids may downregulate mast cell activation and local inflammation.
OBJECTIVE
In order to investigate the effect of cannabinoid receptor-1 (CB1R) agonists on mast cell activation, AEA-derived compounds were newly synthesized and evaluated for their effect on mast cell activation.
METHODS
The effects of selected compounds on FcepsilonRI-induced histamine and beta-hexosaminidase release were evaluated in a rat basophilic leukemia cell line (RBL-2H3). To further investigate the inhibitory effects of CB1R agonist in vivo, an oxazolone-induced atopic dermatitis mouse model was exploited.
RESULTS
We found that CB1R inhibited the release of inflammatory mediators without causing cytotoxicity in RBL-2H3 cells and that CB1R agonists markedly and dose-dependently suppressed mast cell proliferation indicating that CB1R plays an important role in modulating antigen-dependent immunoglobulin E (IgE)-mediated mast cell activation. We also found that topical application of CB1R agonists suppressed the recruitment of mast cells into the skin and reduced the level of blood histamine.
CONCLUSION
Our results indicate that CB1R agonists down-regulate mast cell activation and may be used for relieving inflammatory symptoms mediated by mast cell activation, such as atopic dermatitis, psoriasis, and contact dermatitis.

Keyword

Atopic dermatitis; Cannabinoid receptor agonists; Histamine; Mast cells

MeSH Terms

Animals
Basophils
beta-N-Acetylhexosaminidases
Cannabinoid Receptor Agonists
Cannabinoids
Cell Line
Cytokines
Dermatitis, Atopic*
Dermatitis, Contact
Histamine
Immunoglobulin E
Immunoglobulins
Inflammation
Interleukins
Leukemia
Mast Cells*
Mice
Peptide Hydrolases
Psoriasis
Rats
Skin
Tumor Necrosis Factor-alpha
Cannabinoid Receptor Agonists
Cannabinoids
Cytokines
Histamine
Immunoglobulin E
Immunoglobulins
Interleukins
Peptide Hydrolases
Tumor Necrosis Factor-alpha
beta-N-Acetylhexosaminidases

Figure

• 

  Fig. 1 Newly synthesized cannabinoids and the cAMP assay screening for cannabinoid receptor modulators. (A) The structure of α-oleoyl oleoylamine ethanolamine (α-OOE) and α-oleoyl oleoylamine serinol (α-OOS). (B, C) Synthetic compounds were screened in stable CHO cell lines overexpressing cannabinoid receptor-1 (CB1R) or CB2R. Pre-treatment with CBR agonists significantly diminished the cytosolic levels of cAMP induced by 10 µM forskolin (B). N-arachidonoylethanolamine (AEA) and Hu308 were used as controls for CB1R and CB2R, respectively. All data are presented as the mean±standard error, t-test and analysis of variance. *p<0.01.

• 

  Fig. 2 Effects of cannabinoid receptor-1 (CB1R) agonists on the degranulation of rat basophilic leukemia cell line (RBL-2H3) mast cells. (A, B) The inhibitory effects of α-oleoyl oleoylamine ethanolamine (α-OOE), α-oleoyl oleoylamine serinol (α-OOS), and N-arachidonoylethanolamine (AEA) were determined on ionomycin (2 µM)-induced histamine (A) or β-hexosaminidase release (B) from rat mast cells. (C, D) RBL-2H3 cells sensitized with anti-dinitropheno-immunoglobulin E (anti-DNP-IgE) (0.5 µg/ml) for 12 hr were pretreated with the CB1R agonists. Histamine (C) or β-hexosaminidase release (D) from mast cells was measured. (E) Bone marrow derived-cultured mast cells from mouse bone marrow was sensitized with anti-DNP-IgE for 12 hr and histamine release from cells was measured. All data are presented as the mean±standard error, t-test and analysis of variance. NON: non-stimulated, CON: control (stimulated), IgE: Immunoglobulin E, DNP: dinitrophenol, BSA: bovine serum albumin. *p<0.01 and **p<0.05.

• 

  Fig. 3 Inhibition of proliferation of rat basophilic leukemia cell line (RBL-2H3) cells by cannabinoid receptor-1 (CB1R) agonists. RBL-2H3 cells were untreated or treated with the indicated concentrations of CB1R agonists for 24 hr (A) or with 25 µM agonists for the indicated time periods (B). (C) RBL-2H3 cell proliferation was analyzed by counting cell numbers following CB1R agonist treatment for 48 hr. α-OOE: α-oleoyl oleoylamine ethanolamine, α-OOS: α-oleoyl oleoylamine serinol, AEA: N-arachidonoylethanolamine. Significant difference (*p<0.01 in Student's t-test) as compared with control samples.

• 

  Fig. 4 Effects of cannabinoid receptor-1 agonists in oxazolone-induced atopic dermatitis mice models. (A) Histology of dorsal skin lesions. The dorsal skin of each mouse was removed and fixed for toluidine blue staining (×400). Images are representative of five mice. (B) Change of skin fold thickness in oxazolone model. α-oleoyl oleoylamine ethanolamine (α-OOE), α-oleoyl oleoylamine serinol (α-OOS), and N-arachidonoylethanolamine (AEA) treatment prevented the increase of skin fold thickness. (C) Quantitative histomorphometry of the number of mast cells. Mast cells were counted in the toluidine blue-stained slide. (D) Total blood histamine levels in serum. The histamine concentration of each mouse was measured according to a previously described method. Each point represents the mean±standard error. Data are from five to six mice. VEH: vehicle, DEXA: dexamethasone. *p<0.01, **p<0.05 significantly different from the vehicle group (Student's t-test).

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