An essential role of Nrf2 in American ginseng-mediated anti-oxidative actions in cardiomyocytes.

AIM OF THE STUDY: Ginseng has been used as a folk medicine for thousands of years in Asia, and has become a popular herbal medicine world-wide. Recent studies have revealed that ginseng, including American ginseng, exerts antioxidant effects in the cardiovascular system; however, the underlying mechanisms are not fully understood. Thus, we investigated role of Nrf2, a master transcription factor of endogenous anti-oxidative defense systems, in the regulation of American ginseng-mediated anti-oxidative actions in cardiomyocytes. MATERIALS AND METHODS: A standardized crude extract of American ginseng was supplied by the National Research Council of Canada, Institute for National Measurement Standards. H9C2 cells, a rat cardiomyocyte cell line, were exposed to angiotensin II (Ang II) or tumor necrosis factor alpha (TNFalpha) to induce oxidative stress that was examined by measuring formation of reactive oxygen and nitrogen species. Oxidative stress-induced cell death was induced by exogenous addition of hydrogen peroxide (H(2)O(2)). Proteins were measured by Western blot and mRNA expression was determined by quantitative real time PCR. Nrf2-driven transcriptional activity was assessed by antioxidant response element (ARE)-luciferase reporter assay. Direct Nrf2 binding to its target gene promoters was determined by chromatin immunoprecipitation assay. Adenoviral over-expression of Nrf2 shRNA was utilized to knock down Nrf2 in H9C2 cells. Immunochemical staining was applied for Nrf2 expression in the heart. RESULTS: American ginseng induced dramatic increases in Nrf2 protein expression, Nrf2 nuclear translocation, Nrf2 transcriptional activity, direct Nrf2 binding to its target gene promoters, and expression of a group of anti-oxidative genes driven by Nrf2 in H9C2 cells. In addition, American ginseng inhibited Ang II- or TNFalpha-induced free radical formation and H(2)O(2)-induced cell death in H9C2 cells over-expressed with control shRNA but not in the cells over-expressed with Nrf2 shRNA. Finally, oral administration of American ginseng markedly increased Nrf2 activity in murine hearts. CONCLUSION: These results demonstrate that American ginseng suppresses oxidative stress and oxidative stress-induced cell death in cardiomyocytes through activating the Nrf2 pathway, thereby providing cardioprotection against pathological cardiac remodeling.

American ginseng preferentially suppresses STAT/iNOS signaling in activated macrophages.

AIM OF THE STUDY: Ginseng has been used as general tonic for thousands of years in Asia and becomes a popular herbal medicine all over the world. However, the cellular and molecular mechanisms underlying its benefit effects are less explored. Thus, we investigated the effect of a crude extract from Panax quinquefolius (American ginseng) on suppression of pro-inflammatory responses in macrophages with a focus on signal transducer and activator of transcription (STAT) signaling. MATERIALS AND METHODS: The crude extract of American ginseng that was supplied by the National Research Council of Canada, Institute for National Measurement Standards (NRCC-INMS) was freshly solvated in Dulbecco's Modified Eagle Medium (DMEM) prior to each experiment. RAW264.7 cells, a murine macrophage cell line, were exposed to lipopolysaccharide (LPS) to induce inflammatory responses such as expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase 2 (COX2). Proteins were measured by Western blot and mRNA expression was determined by quantitative real-time PCR (Q-PCR). Activator protein 1 (AP-1)-, nuclear factor-kappaB (NF-kappaB)- and STAT-mediated transcriptional activities were investigated using luciferase reporter constructs. RESULTS: American ginseng inhibited LPS-induced iNOS expression; however, it did not affect LPS-induced COX2 expression. While American ginseng had no impact on LPS-induced activation of AP-1 or NF-kappaB pathways, it dramatically inhibited LPS-induced activation of STAT signaling. Moreover, American ginseng and AG490, an inhibitor of STAT cascade, synergistically suppressed the LPS-induced iNOS expression. CONCLUSION: American ginseng selectively inhibits the expression of iNOS via suppression of STAT cascade but not NF-kappaB and AP-1 pathways in inflamed macrophages. Such a preferential suppression of STAT/iNOS cascade by American ginseng might have therapeutic potential for inflammatory diseases with over-activation of iNOS.