Department of Cell Biology and Anatomy, University of South Carolina, Columbia, SC 29208, USA.
J Ethnopharmacol. 2010 Jul 20;130(2):222-30. doi: 10.1016/j.jep.2010.03.040. Epub 2010 May 4.
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.
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.
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.
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.
人参在亚洲作为民间药物已使用了数千年，并已成为全球流行的草药。最近的研究表明，人参（包括西洋参）在心血管系统中具有抗氧化作用；然而，其潜在机制尚不完全清楚。因此，我们研究了 Nrf2（内源性抗氧化防御系统的主要转录因子）在调节西洋参介导的心肌细胞抗氧化作用中的作用。
由加拿大国家研究理事会国家计量标准研究所提供西洋参的标准化粗提取物。H9C2 细胞是大鼠心肌细胞系，用血管紧张素 II（Ang II）或肿瘤坏死因子 alpha（TNFalpha）孵育以诱导氧化应激，通过测量活性氧和氮物种的形成来检测。通过外源性添加过氧化氢（H2O2）诱导氧化应激诱导的细胞死亡。通过 Western blot 测量蛋白质，通过定量实时 PCR 测定 mRNA 表达。通过抗氧化反应元件（ARE）-荧光素酶报告基因测定评估 Nrf2 驱动的转录活性。通过染色质免疫沉淀测定确定 Nrf2 与其靶基因启动子的直接结合。利用腺病毒过表达 Nrf2 shRNA 敲低 H9C2 细胞中的 Nrf2。免疫化学染色用于检测心脏中的 Nrf2 表达。
西洋参诱导 H9C2 细胞中 Nrf2 蛋白表达、Nrf2 核易位、Nrf2 转录活性、Nrf2 直接与靶基因启动子结合以及一组由 Nrf2 驱动的抗氧化基因表达的显著增加。此外，西洋参抑制了在转染对照 shRNA 的 H9C2 细胞中 Ang II 或 TNFalpha 诱导的自由基形成和 H2O2 诱导的细胞死亡，但在转染 Nrf2 shRNA 的细胞中则没有。最后，西洋参的口服给药显著增加了小鼠心脏中的 Nrf2 活性。
这些结果表明，西洋参通过激活 Nrf2 途径抑制心肌细胞中的氧化应激和氧化应激诱导的细胞死亡，从而提供对病理性心脏重塑的心脏保护。
