Gabriele Morena, Pucci Laura, La Marca Margherita, Lucchesi Daniela, Della Croce Clara Maria, Longo Vincenzo, Lubrano Valter
National Research Council (CNR), Institute of Biology and Agricultural Biotechnology (IBBA), Pisa Unit, Research Area of Pisa, Via Moruzzi 1, 56124 Pisa, Italy.
Department of Clinical and Experimental Medicine, Section of Metabolic Diseases, University of Pisa, Via Paradisa 2, 56124 Pisa, Italy.
Cell Mol Biol Lett. 2016 Aug 12;21:10. doi: 10.1186/s11658-016-0015-z. eCollection 2016.
This study focused on an extract from fermented flour from the variety of the common bean . The extract, lysate (Lys LJ), is enriched in antioxidant compounds during the fermentation. We assessed it for its protective effect on endothelial cells treated with oxidized-LDL (ox-LDL). The oxidative stress was determined by measuring the contents of thiobarbituric acid-reactive substances and reactive oxygen metabolites. ICAM-1, ET-1 and IL-6 concentrations were assessed using ELISA. LOX-1 and CHOP expression were analyzed using both quantitative RT-PCR and ELISA or western blotting. Ox-LDL treatment induced significant oxidative stress, which was strongly reduced by pre-treatment with the extract. The ox-LDL exposure significantly enhanced ICAM-1, IL-6 and ET-1 levels over basal levels. Lys LJ pre-treatment exerted an inhibitory effect on ox-LDL-induced endothelial activation with ICAM-1 levels comparable to those for the untreated cells. IL-6 and ET-1 production, although reduced, was still significantly higher than for the control. Both LOX-1 and CHOP expression were upregulated after ox-LDL exposure, but this effect was significantly decreased after Lys LJ pre-treatment. Lys LJ alone did not alter the ICAM-1, IL-6 and ET-1 concentrations or CHOP expression, but it did significantly lower the LOX-1 protein level. Our data suggest that Lys LJ is an effective antioxidant that is able to inhibit the oxidation process, but that it is only marginally active against inflammation and ET-1 production in HMEC-1 exposed to ox-LDL.
本研究聚焦于普通豆品种发酵面粉的提取物。该提取物即裂解物（Lys LJ），在发酵过程中富含抗氧化化合物。我们评估了其对经氧化型低密度脂蛋白（ox-LDL）处理的内皮细胞的保护作用。通过测量硫代巴比妥酸反应性物质和活性氧代谢物的含量来确定氧化应激。使用酶联免疫吸附测定（ELISA）评估细胞间黏附分子-1（ICAM-1）、内皮素-1（ET-1）和白细胞介素-6（IL-6）的浓度。使用定量逆转录聚合酶链反应（RT-PCR）以及ELISA或蛋白质免疫印迹法分析凝集素样氧化型低密度脂蛋白受体-1（LOX-1）和 Chop 同源蛋白（CHOP）的表达。ox-LDL处理诱导了显著的氧化应激，而提取物预处理可显著减轻这种应激。ox-LDL暴露使ICAM-1、IL-6和ET-1水平显著高于基础水平。Lys LJ预处理对ox-LDL诱导的内皮细胞活化具有抑制作用，ICAM-1水平与未处理细胞相当。IL-6和ET-1的产生虽有所降低，但仍显著高于对照组。ox-LDL暴露后，LOX-1和CHOP的表达均上调，但Lys LJ预处理后这种作用显著减弱。单独的Lys LJ不会改变ICAM-1、IL-6和ET-1的浓度或CHOP的表达，但它确实显著降低了LOX-1蛋白水平。我们的数据表明，Lys LJ是一种有效的抗氧化剂，能够抑制氧化过程，但在暴露于ox-LDL的人微血管内皮细胞-1（HMEC-1）中，其对炎症和ET-1产生的活性仅为轻微有效。
