Abstract
Melittin, the major active peptide of honeybee venom (BV), has potential for use in adjuvant immunotherapy. The immune system response to different stimuli depends on the secretion of different metabolites from macrophages. One potent stimulus is lipopolysaccharide (LPS), a component isolated from gram-negative bacteria, which induces the secretion of pro-inflammatory cytokines in macrophage cell cultures. This secretion is amplified when LPS is combined with melittin. In the present study, pure melittin was isolated from whole BV by flash chromatography to obtain pure melittin. The ability of melittin to enhance the release of tumour necrosis factor-α (TNF-α), Interleukin (IL-1β, IL-6, and IL-10) cytokines from a macrophage cell line (THP-1) was then assessed. The response to melittin and LPS, applied alone or in combination, was characterised by metabolic profiling, and the metabolomics results were used to evaluate the potential of melittin as an immune adjuvant therapy. The addition of melittin enhanced the release of inflammatory cytokines induced by LPS. Effective chromatographic separation of metabolites was obtained by liquid chromatography-mass spectrometry (LC-MS) using a ZIC-pHILIC column and an ACE C4 column. The levels of 108 polar and non-polar metabolites were significantly changed (p ˂ 0.05) following cell activation by the combination of LPS and melittin when compared to untreated control cells. Overall, the findings of this study suggested that melittin might have a potential application as a vaccine adjuvant.
Highlights
Correct recognition of inflammatory signals in the body is vital to avoid the risk of autoimmune and chronic inflammatory diseases
The studies were performed on THP-1 cell lines before and after differentiation with phorbol 12-myristate 13-acetate (PMA) to evaluate the differences in cell lysis due to melittin (Figure 1)
The cytokine production was analysed by enzyme-linked immunosorbent assay (ELISA) following melittin administration at 0.5 and 1 μg/mL doses, whereas melittin was added at 1 μg/mL for the metabolomics study
Summary
Correct recognition of inflammatory signals in the body is vital to avoid the risk of autoimmune and chronic inflammatory diseases. Many of these signals arise from macrophages, and determination and investigation of the composition of local tissue microenvironments (e.g., metabolites, cytokines, and inflammatory signals) can be performed in order to understand the metabolic behaviour of macrophages following their activation [1]. Inflammation is a primary process that is triggered as a protective response to any stimulus of the innate immune system, such as injuries or infections, which pose a real threat to normal cell biology [2].
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