Bioactivity-guided identification and cell signaling analysis to investigate the immunomodulatory effects of ginseng on U937 cells

Abstract

Panax ginseng (Ginseng) is one of the most commonly used medicinal herbs worldwide. It is believed to have beneficial effects against human diseases, and its active components, ginsenosides, may play critical roles in its diverse physiological actions. However, the mechanisms underlying ginseng’s effects remain to be investigated. We hypothesize that some biological effects of ginseng are due to its anti-inflammatory activities. To investigate the effects of ginseng on inflammation, human monocytic cells (U937) were treated sequentially with 70% ethanol–water extracts of Panax ginseng (PGSE) and tumor necrosis factor-alpha (TNF-α). Gene expression profiles of the cells were examined by genechip analysis (Human Genome U133 Plus 2.0 arrays, Affymetrix) and validated by Taqman quantitative RT-PCR, and protein expressions were assayed by ELISA. The composition of ginsenosides in 70% ethanol–water extracts of ginseng was analyzed by HPLC. Activation of signaling kinases was examined by Western blot analysis and the respective anti-phospho-protein kinase antibodies. PGSE significantly inhibited the transcription and secretion of CXCL-10 following TNF-α stimulation. Nine ginsenosides including Rb1, Rb2, Rc, Rd, Re, Rf, Rg1, Rg3 and Rh1 were identified in the PGSE by HPLC. Seven out of nine ginsenosides could significantly inhibit TNF-α-induced CXCL-10 expression in U937 cells but the CXCL-10 suppressive effects of individual ginsenosides was less than that of the crude extract or the mixture of reconstituted ginsenosides. The suppressive effect of the reconstituted mixture of nine ginsenosides at the corresponding doses was comparable to the PGSE treatment in a dose-dependent manner. Furthermore, the CXCL-10 suppression can be correlated with the inactivation of ERK1/2 pathways by the PGSE. In summary, our results provide evidence that ginseng can suppress TNF-α-inducible cytokines and signaling proteins in promonocytic cells.