Lipopolysaccharide induces calcitonin gene-related peptide in the RAW264.7 macrophage cell line

Calcitonin gene-related peptide (CGRP) is widely distributed and plays important roles in a wide array of biological functions. It is enriched in primary sensory neurons and hence involved in nociception and neurogenic inflammation. Recent studies have shown that CGRP can be produced by immune cells such as monocytes/macrophages following inflammatory stimulation, suggesting a role in innate immunity. However, it is unclear how CGRP is up-regulated in macrophages and if it plays a role in macrophage functions such as the production of cytokines and chemokines. Using enzyme-linked immunosorbent assay (ELISA) and multiplex ELISA, lipopolysaccharide (LPS) was found to induce CGRP in the RAW 264.7 macrophage cell line. LPS-induced inflammatory mediators such as nerve growth factor (NGF), interleukin-1beta (IL-1beta), IL-6, prostaglandin E(2) (PGE(2)) and nuclear factor-kappaB (NF-kappaB) signalling are involved in inducing CGRP, whereas the NGF receptor trkA and CGRP receptor signalling pathways are unexpectedly involved in suppressing LPS-induced CGRP, which leads to the fine-tune regulation of CGRP release. Exogenous CGRP and CGRP receptor antagonists, in a concentration-dependent manner, stimulated, inhibited or had no effect on basal or LPS-induced release of monocyte chemoattractant protein-1, IL-1beta, IL-6, tumour necrosis factor-alpha and IL-10 in RAW macrophages. The ligand-concentration-dependent regulation of the production of inflammatory mediators by CGRP receptor signalling is a novel mechanism underlying the stimulating and suppressing role of CGRP in immune and inflammatory responses. Together, our data suggest that monocytes/macrophages are an important source of CGRP. Inflammation-induced CGRP has a positive or negative reciprocal effect on the production of other pro- and anti-inflammatory mediators. Thereby CGRP plays both facilitating and suppressing roles in immune and inflammatory responses.