Insight into the molecular mechanism of a herbal injection by integrating network pharmacology and in vitro

Chinese medical herbs could treat complex diseases through the synergistic effect of multi-components, multi-targets and multi-channels. However, it was difficult to systematically investigate the pharmacological mechanisms of action due to the complex chemical composition and the lack of an effective research approach. Fortunately, network pharmacology as an integrated approach was proposed to systematically investigate and explain the underlying molecular mechanisms of Chinese medical herbs. Reduning injection (RDN) is one of the herbal injections for treatment of upper respiratory tract infections (URTIs). Previous studies revealed the molecular mechanism of RDN on URTIs through network pharmacology. In this work, the mechanism of RDN was verified by enzyme linked immunosorbent assay (ELISA), Western Blot, immunofluorescence assay and electrophoretic mobility shift assay (EMSA) in lipopolysaccharide (LPS)-induced RAW264.7 cells and enzyme assay. RDN dose-dependently suppressed the production of nitric oxide (NO), prostaglandin E2 (PGE2), interleukin-6 (IL-6) and interleukin-1β (IL-1β), and reduced the protein expression of inducible NO synthetase (iNOS) and cyclooxygenase-2 (COX-2), which could be related to its suppression on the phosphorylations of mitogen-activated protein (MAP) kinases, including extracellular signal-regulated kinase (ERK), c-jun NH2-terminal kinase(JNK) and p38, as well as the activation and translocation of nuclear factor-κB (NF-κB). In addition, the activity of RDN on PGE2 was also partly attributed to the inhibition of COX-2 enzyme. Therefore, it can be concluded that RDN inhibited the production of inflammatory mediators and the macrophage activation to treat URTIs via down-regulating the activation of MAPK and NF-κB signaling pathways, which might pave a way to illustrate the molecular mechanism of herbs.