Anti-inflammatory Mechanism of a Diterpenoid, Ebractenoid F, Isolated from Euphorbia ebracteolata Hayata

Abstract

Euphorbia ebracteolata Hayata, belongs to Euphorbiaceae, is widely distributed through the south of China. In the traditional chinese medicine (TCM), the roots of E. ebracteolata have been prescribed for chronic inflammation-mediated diseases such as tracheitis, cutaneous tuberculosis, tumor, and psoriasis. It has been reported that E. ebracteolata is rich in bioactive components such as acetophenones, flavonoids, and diterpenes. Among the constituents of E. ebracteolata, diterpenes in E. ebracteolata have effects on anti-inflammation, anti-tumor, and anti-fungal activities. However, the anti-inflammatory mechanism of most of constituents in E. ebracteolata Radix is not yet fully discovered. For find the anti-inflammatory compound from E. ebracteolata Radix, bioassay-guided fractionation, which is a general way to isolate and characterize bioactive compounds from natural products, was conducted. It might be a worth way to find out the most potential compounds possessing biological activities. Thus, we applied this concept to process separation. In addition, a separation method, called high speed counter-current chromatography (HSCCC), was co-operated because it provides us many advantages in terms of high yield, sample loading capacity, easy scale-up, purity, and resolution. Following the bioassay-guided isolation, the compounds were elucidated and identified by various spectroscopic ways. Ebractenoid F (EF) showing the potential effects was ultimately obtained by this method, bioassay-guided isolation. Inflammation is a key factor of protection system triggered by immune cells in response to harmful stimuli. It mediates various cellular processes such as proliferation, cell cycle, and differentiation. If the stimuli remain, the acute inflammatory response is connected to the chronic inflammatory response. The nitric oxide (NO) and SEAP are the inflammatory mediators, so they could arise from the inflammation. In this vein of thought, we seek for anti-inflammatory agents which can reverse both NO and SEAP production regulated by NF-κB. EF shows the prominent inhibition of NF-κB Secretary Alkaline Phosphatase (SEAP) (IC50 = 7.71 μM), implying that it is specifically targeting on NF-κB. It decreases the downstream of NF-κB including various pro-inflammatory mediators, such as inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and interleukin-6 (IL-6), at both protein and mRNA levels. Moreover, it down-regulates phosphorylation and degradation of inhibitory κB (IκB)-α in 20 minutes. Nuclear translocation and transcriptional activity are suppressed when cells are treated with EF. In addition, we investigated whether it has an effect on LPS-induced mitogen-activated protein kinase (MAPKs, p-ERK, and p-JNK) and the upstream signaling pathways (p-AKT and p-IKK). Taken together, we tried to find the anti-inflammatory compound from E. ebracteolata using bioassay-guided fractionation in that there are not enough studies. At this research, we could discover a potential compound, ebractenoid F (EF). Through this achievement, we also expected that inflammation-mediated diseases can be managed with this compound since the NF-κB signaling pathway related to inflammation was suppressed. Although we still need to study on the effect of ebractenoid F, it is possible to be a key regulator on inflammation-related to diseases.