Nutraceutical formulation based on a synergic combination of melatonin and palmitoylethanolamide for the management of allergic events

Abstract

The management of allergic events is a growing global health issue, especially in industrialized countries. This disease is an immune-mediated process, regulated by the interaction of IgE with an allergen, resulting in mast cell activation, which concerns the release of several immune-inflammatory modulators, i.e., histamine, β-hexosaminidase, COX-2, IL-6, and TNF-α, responsible for the main allergic-reaction associated symptoms. The aim of the present study was the efficacy evaluation of an alternative remedy, an innovative nutraceutical formulation (NF) based on the synergic combination of melatonin (MEL) and palmitoylethanolamide (PEA) for the prevention and treatment of immune disease. At first, the intestinal bioaccessibility of PEA and MEL in NF was assessed at 1.6 and 36%, respectively. Then the MEL and PEA ability to modulate the release of immune-inflammatory modulators in the human mast cell line (HMC-1.2) at their bioaccessible concentration was investigated. Our results underline that NF treatment was able to reduce COX-2 mRNA transcription levels (-30% vs. STIM, p < 0.0001) in stimulated HMC-1.2 and to contract COX-2 enzymatic activity directly (IC₅₀: 152 μg/mL). Additionally, NF showed valuable ability in reducing histamine and β-hexosaminidase release in stimulated HMC-1.2, as well as in decreasing TNF-α and IL-6 mRNA transcription levels and protein production.

Keywords: human mast cell; immune response; melatonin; nutraceutical formulation; palmitoylethanolamide; synergic combination.

Figure 1

Chromatographic profile and chemical structures of PEA and MEL standards by HPLC-DAD-FLD analysis. PEA (500 ppm) was analyzed by HPLC-DAD at the wavelength of 205 nm. MEL (25 ppm) was analyzed by HPLC-FLD.

Figure 2

Cytotoxic effect of PEA (A), melatonin (B), and NF (C) on HMC-1.2 cells after 72 h evaluated by the MTT assay. Values are expressed as mean ± SEM from three independent experiments.

Figure 3

Histamine (A) and β-hexosaminidase (B) release in HMC-1.2 cells following pretreatment with PEA, MEL, and NF and stimulation with PMA 50 nM and ionophore 1 μM. Values are expressed as mean ± SEM from three independent experiments. °°°°p < 0.0001 indicates a significant effect of PMA/Ionophore compared to unstimulated cells (CTR); ****p < 0.0001 indicates significant effect of PEA, MEL, and NF compared to stimulated cells.

Figure 4

Expression of IL-6 (A) and TNF-α (B) assessed with qPCR analysis in HMC-1.2 cells pretreated for 1 h with PEA 16 μM, melatonin 80 nM and NF 16 μM + 80 nM and stimulated with PMA 50 nM and ionophore 1 μM for 6 h. (C) Levels of IL-6 and TNF-α in supernatants assessed by multiplex assay. Values are expressed as mean ± SEM from three independent experiments. °°°°p < 0.0001 indicates a significant effect of PMA/Ionophore compared to unstimulated cells (CTR); *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001 indicate a significant effect of PEA, MEL or NF compared to stimulated cells.

Figure 5

(A) Expression of PTGS2 assessed with qPCR analysis in HMC-1.2 cells treated with PEA 16 μM, melatonin 80 nM, and NF 16 μM + 80 nM for 1 h following stimulation with PMA 50 nM and ionophore 1 μM and treatment for 6 h. Inhibition of COX-2 activity (expressed in %) of PEA (B) MEL (C) and NF (D). Values are expressed as mean ± SEM from three independent experiments. °°°°p < 0.0001 indicates a significant effect of PMA/Ionophore compared to unstimulated cells (CTR); **p < 0.01 and ****p < 0.0001 indicate a significant effect of PEA, MEL or NF compared to stimulated cells.