Objective: To investigate the effect of Angelicae Pubescentis Radix (APR) on the activity of endocannabinoid hydrolase and N-acylethanolamine-hydrolyzing acid amidase (NAAA), and to demonstrate the mechanism of anti-inflammatory effect of APR by in vitro lipopolysaccharide (LPS)-induced inflammation model.
Method: APR essential oil was extracted by steam distillation, and the chemical components were identified by GC-MS. Enzymatic activity was performed by using recombinant NAAA-overexpressing protein and detected by LC-MS. Lipids were extracted by methonal/chloroform mixure and analyzed by LC-MS. mRNA and protein expression levels of proinflammatory genes were examined by Real time-PCR and ELISA assay kit, respectively. The content of nitro oxide (NO) was detected by Griess reaction.
Result: Twenty active components were identified from APR essential oil which inhibited NAAA activity in a dose-dependent manner. On the LPS-induced RAW264.7 cells, APR essential oil reversed LPS-suppressed N-palmitoylethanolamide (PEA) contents in a dose-dependent manner and reduced LPS-induced proinflammatory genes, TNF-alpha and IL-6. Moreover, APR essential oil reduced the mRNA expression of iNOS, subsequently reduced the release of NO, a classic inflammatory marker.
Conclusion: The research demonstrated that the effect of APR on inflammation is mediated by the inhibition of NAAA activity, which increase the cellular endobioactor PEA levels and decrease proinflammatory factor. The results suggest that APR can serve as a nature NAAA inhibitor.