Lucchesi Daniela, Russo Rossella, Gabriele Morena, Longo Vincenzo, Del Prato Stefano, Penno Giuseppe, Pucci Laura
Department of Clinical and Experimental Medicine, Section of Metabolic Diseases, University of Pisa, Pisa, Italy.
Institute of Agricultural Biology and Biotechnology, National Research Council, CNR, Pisa, Italy.
PLoS One. 2014 Oct 17;9(10):e109298. doi: 10.1371/journal.pone.0109298. eCollection 2014.
Increased oxidative stress contributes to the functional impairment of endothelial progenitor cells (EPCs), the pivotal players in the servicing of the endothelial cell lining. Several evidences suggest that decreasing oxidative stress by natural compounds with antioxidant properties may improve EPCs bioactivity. Here, we investigated the effects of Lisosan G (LG), a Triticum Sativum grain powder, and Lady Joy (LJ), a bean lysate, on function of EPCs exposed to oxidative stress. Peripheral blood mononuclear cells were isolated and plated on fibronectin-coated culture dishes; adherent cells, identified as early EPCs, were pre-treated with different concentrations of LG and LJ and incubated with hydrogen peroxide (H2O2). Viability, senescence, adhesion, ROS production and antioxidant enzymes gene expression were evaluated. Lysate-mediated Nrf-2 (nuclear factor (erythroid-derived 2)-like 2)/ARE (antioxidant response element) activation, a modulator of oxidative stress, was assessed by immunocytochemistry. Lady Joy 0.35-0.7 mg/ml increases EPCs viability; pre-treatment with either LG 0.7 mg/ml and LJ 0.35-0.7 mg/ml protect EPCs viability against H2O2-induced injury. LG 0.7 and LJ 0.35-0.7 mg/ml improve EPCs adhesion; pre-treatment with either LG 0.35 and 0.7 mg/ml or LJ 0.35, 0.7 and 1.4 mg/ml preserve adhesiveness of EPCs exposed to H2O2. Senescence is attenuated in EPCs incubated with lysates 0.35 mg/ml. After exposure to H2O2, LG pre-treated cells show a lower senescence than untreated EPCs. Lysates significantly decrease H2O2-induced ROS generation. Both lysates increase glutathione peroxidase-1 and superoxide dismutase-2 (SOD-2) expression; upon H2O2 exposure, pre-treatment with LJ allows higher SOD-2 expression. Heme oxigenase-1 increases in EPCs pre-treated with LG even upon H2O2 exposure. Finally, incubation with LG 0.7 mg/ml results in Nrf-2 translocation into the nucleus both at baseline and after the oxidative challenge. Our data suggest a protective effect of lysates on EPCs exposed to oxidative stress through the involvement of antioxidant systems. Lisosan G seems to activate the Nrf-2/ARE pathways.
氧化应激增加会导致内皮祖细胞（EPCs）功能受损，而内皮祖细胞是维持内皮细胞内衬的关键细胞。多项证据表明，具有抗氧化特性的天然化合物降低氧化应激可能会改善内皮祖细胞的生物活性。在此，我们研究了小麦籽粒粉Lisosan G（LG）和豆类裂解物Lady Joy（LJ）对暴露于氧化应激的内皮祖细胞功能的影响。分离外周血单核细胞并接种于纤连蛋白包被的培养皿上；将鉴定为早期内皮祖细胞的贴壁细胞用不同浓度的LG和LJ预处理，并与过氧化氢（H2O2）孵育。评估细胞活力、衰老、黏附、活性氧产生和抗氧化酶基因表达。通过免疫细胞化学评估裂解物介导的氧化应激调节剂核因子（红系衍生2）样2（Nrf-2）/抗氧化反应元件（ARE）的激活。0.35 - 0.7 mg/ml的Lady Joy可提高内皮祖细胞活力；用0.7 mg/ml的LG或0.35 - 0.7 mg/ml的LJ预处理可保护内皮祖细胞活力免受H2O2诱导的损伤。0.7 mg/ml的LG和0.35 - 0.7 mg/ml 的LJ可改善内皮祖细胞黏附；用0.35和0.7 mg/ml的LG或0.35、0.7和1.4 mg/ml的LJ预处理可保持暴露于H2O2的内皮祖细胞的黏附性。用0.35 mg/ml的裂解物孵育的内皮祖细胞衰老减弱。暴露于H2O2后，经LG预处理的细胞衰老程度低于未处理的内皮祖细胞。裂解物可显著降低H2O2诱导的活性氧生成。两种裂解物均增加谷胱甘肽过氧化物酶-1和超氧化物歧化酶-2（SOD-2）的表达；暴露于H2O2时，用LJ预处理可使SOD-2表达更高。即使在暴露于H2O2后，经LG预处理的内皮祖细胞中血红素加氧酶-1也会增加。最后，用0.7 mg/ml的LG孵育在基线和氧化应激刺激后均导致Nrf-2易位至细胞核。我们的数据表明，裂解物通过抗氧化系统参与对暴露于氧化应激的内皮祖细胞具有保护作用。Lisosan G似乎激活了Nrf-2/ARE途径。
