Ginseng is one of the most commonly used herbal medicines in China, Asia and Western countries. Studies have shown a wide range of beneficial effects of ginseng against human diseases . The potential therapeutic effects of ginseng have been attributed to its immunostimulatory, anti-oxidant and anti-inflammatory activities. In this study, we used human promonocytic U937 cells to investigate the modulatory effects of ginseng in cellular response to TNF-α-mediated inflammation. By using the genechip approach, we obtained a global gene expression profile in monocytic cell model following different experimental treatments. Our genechip results showed a potent suppressive effect of the PGSE on the expression of TNF-α-inducible genes including CXCL-10. These results have been validated by using quantitative RT-PCR and ELISA. Moreover, nine ginsenosides were identified in our ginseng extract by using HPLC analysis. Interestingly, other groups have reported the anti-inflammatory activity of these ginsenosides. Our results showed that seven out of nine ginsenosides could significantly inhibit TNF-α-induced CXCL-10 expression in U937 cells. However, the suppressive effect of individual ginsenosides on CXCL-10 induction was less than that of the mixture of ginsenosides or PGSE alone. Furthermore, we found that the CXCL-10 suppressive effect correlates with the inactivation of the ERK1/2 signalling pathways by PGSE.
The immunomodulatory effects of ginseng or ginsenosides have been reported in in vivo and in vitro studies. Kim et al. showed that Panax ginseng enhances the recovery of natural killer (NK) cell functions in cyclophosphamide-treated mice, and provides protection against infection with Listeria monocytogenes . Ginseng radix extracts induce production of TNF-α and IFN-γ in murine spleen cells and peritoneal macrophages via toll-like receptor (TLR)-4 . Additionally, Ginsenan S-IIA, a component of acidic polysaccharide of Panax ginseng, is a potent inducer of IL-8 in human monocytes and THP-1 cells . In contrast, ginseng or ginseng extract have been shown to have anti-inflammatory effects such as suppressing the expression of proinflammatory cytokines or mediators. For instance, ginsan, a polysaccharide extracted from Panax ginseng, protects mice from lethality induced by Staphylococcus aureus and such effect was associated with suppression of proinflammatory cytokines production including TNF-α, IL-1β, IL-6, IL-12, IL-18 and IFN-γ . Moreover, 20(S)-Protopanaxatriol, one of the major metabolites of ginsenosides, inhibits the increase in iNOS and COX-2 expressions following LPS stimulation through inactivation of NF-κB . The diverse immunologic effects of ginseng may be due to multiple effects of the ginsenosides or its other active components. Therefore, comprehensive studies of ginseng and its constituents are still needed to provide detailed understanding of their actions in humans.
Since our study is focused on immunomodulation, only the list of cytokines or cytokine-regulated genes is reported in Table 2. Here, the PGSE can cause a potent inhibition on the transcription of TNF-α inducible genes including CXCL-10, G protein-coupled receptor 84, TNF-α induced-protein 6, IκB-alpha, IκB-zeta and phosphodiesterase 4B (Table 2). Interestingly, those genes inhibited by PGSE have been shown to be expressed in TNF-α mediated-inflammatory diseases [15, 27–29]. Therefore, it is plausible that ginseng down regulates TNF-α mediated inflammation through suppressing the production of inflammatory mediators in monocytes or macrophages. However, it seems that this PGSE preparation did not contain potent cytokine inducing factors. As previous reports showed that the immunostimulating components such as polysaccharides of ginseng extracts come from the ethanol insoluble fraction [7, 30, 31], this component appears to have been excluded or its biological activity was attenuated by constituents in the extract we studied.
CXCL-10 is an important chemokine downstream of TNF-α signalling pathways and a well-documented mediator of inflammation. CXCL-10 initiates its biological functions through binding to its high affinity receptor CXCR-3 leading to recruitment of the activated effector lymphocytes including CD4+ and CD8+ T cells as well as NK cells to the site of infection or injury . Similar to TNF-α, the uncontrolled production of CXCL-10 also is associated with the pathogenesis of acute and chronic inflammatory diseases including intrahepatic inflammation during chronic HCV infection, atherosclerosis, inflammatory bowel disease, and multiple sclerosis as well as tumorigenesis and metastasis [33–37]. In our study, the PGSE or chemically defined mixture of its constituent ginsenosides showed potent inhibitory effects on TNF-α-stimulated CXCL-10 expression (Figure 4C) suggesting a specific anti-inflammatory property of ginseng.
Ginsenosides belong to a family of steroidal saponins that are believed to be responsible for the pharmacological effects of ginseng. About 30 different ginsenosides have been isolated and identified from Panax ginseng. The two major groups of ginsenosides are panaxadiol and panaxatriol. The panaxadiol group contains Rb1, Rb2, Rc, Rd and Rh2 whereas the panaxatriol group contains Re, Rf, Rg1, Rg2,Rg3 and Rh1. Previous studies have shown different properties of ginsenosides among each other, and differential effects of ginsenosides panaxadiol and panaxatriols have been found in inflammatory diseases . Here, we found that both of the panaxadiol and panaxatriol groups of ginsenosides showed similar inhibitory effects on TNF-α-induced CXCL-10 production. Additionally, the inhibitory effects could be due to complementary or collective effect of ginsenosides mixtures instead of a single ginsenoside. Another possible explanation is stereoisomerism of natural and synthetic compounds since the source of ginsenosides is different from the ginseng extract. Similar phenomenon has been reported by another group recently .
Following the activation of TNF-α signalling pathways, the downstream MAPK cascades and transcription factors, NF-κB and AP-1, are activated to induce gene transcription. Previous studies have shown that NF-κB and/or MAPK signalling cascades play critical roles in acute and chronic inflammatory diseases. Here our result showed that the PGSE inhibited the basal level of ERK1/2 phosphorylation at 1 or 3 mg/ml (Figure 5C). This observation is in agreement with the effect of PD98059, a known inhibitor of ERK1/2, on the suppression of TNF-α-induced CXCL-10 transcription (not shown). In contrast, the PGSE did not show any effect on TNF-α-induced activation of p38MAPK and NF-κB. These results suggest that PGSE inhibited CXCL-10 expression by perturbing MAPK signalling cascades.