School of Life Science and Technology, China Pharmaceutical University, Nanjing, China.
Department of Occupational disease, Qingdao Central Hospital, Shandong, China.
J Cell Mol Med. 2021 Jan;25(1):345-357. doi: 10.1111/jcmm.16073. Epub 2020 Dec 15.
In this study, we investigated the protective effects of gastrodin (Gas) against homocysteine-induced human umbilical vein endothelial cell (HUVEC) injury and the role of the phosphoinositide 3-kinase (PI3K)/threonine kinase 1 (Akt)/endothelial nitric oxide synthase (eNOS) and NF-E2-related factor 2 (Nrf2)/antioxidant response element (ARE) pathways. We stimulated cells with homocysteine (1 mmol/L, 24 hours) and tested the effects of gastrodin (200-800 μg/mL) on cell viability and the production of malondialdehyde (MDA), lactate dehydrogenase (LDH) and reactive oxygen species (ROS). Then, Nrf2 distribution in the cytoplasm and nucleus as well as the expression of enzymes downstream of Nrf2 was determined. Furthermore, we analysed the expression of bax, bcl-2 and cleaved caspase3, and assessed the involvement of the PI3K/Akt/eNOS pathway by Western blots. Finally, we tested the vasoactive effect of gastrodin in thoracic aortic rings. The results showed that gastrodin decreased MDA, LDH and ROS production and increased cell viability, NO production and relaxation of thoracic aortic rings. Moreover, the protective effects of Gas on NO production and relaxation of thoracic aortic rings were blocked by L-NAME but enhanced by Cav-1 knockdown, and MK-2206 treatment abolished the effect of Gas on the ROS. In addition, treatment with gastrodin increased Nrf2 nuclear translocation, thus enhancing the expression of downstream enzymes. Finally, gastrodin increased the expression of PI3K, p-Akt, and eNOS and decreased Cav-1 protein expression. In conclusion, our study suggested that gastrodin may protect HUVECs from homocysteine-induced injury, and the PI3K/Akt/eNOS and Nrf2/ARE pathways may be responsible for the efficacy of gastrodin.
在这项研究中，我们研究了天麻素（Gas）对同型半胱氨酸诱导的人脐静脉内皮细胞（HUVEC）损伤的保护作用，以及磷酸肌醇 3-激酶（PI3K）/苏氨酸激酶 1（Akt）/内皮型一氧化氮合酶（eNOS）和核因子红细胞 2 相关因子 2（Nrf2）/抗氧化反应元件（ARE）途径的作用。我们用同型半胱氨酸（1mmol/L，24 小时）刺激细胞，测试天麻素（200-800μg/mL）对细胞活力和丙二醛（MDA）、乳酸脱氢酶（LDH）和活性氧（ROS）产生的影响。然后，确定 Nrf2 在细胞质和细胞核中的分布以及 Nrf2 下游酶的表达。此外，我们分析了 bax、bcl-2 和 cleaved caspase3 的表达，并通过 Western blot 分析了 PI3K/Akt/eNOS 途径的参与。最后，我们测试了天麻素对胸主动脉环的血管活性作用。结果表明，天麻素降低 MDA、LDH 和 ROS 的产生，增加细胞活力、NO 的产生和胸主动脉环的舒张。此外，Gas 对 NO 产生和胸主动脉环舒张的保护作用被 L-NAME 阻断，但被 Cav-1 敲低增强，MK-2206 处理消除了 Gas 对 ROS 的作用。此外，天麻素治疗增加了 Nrf2 的核转位，从而增强了下游酶的表达。最后，天麻素增加了 PI3K、p-Akt 和 eNOS 的表达，降低了 Cav-1 蛋白的表达。总之，我们的研究表明，天麻素可能保护 HUVEC 免受同型半胱氨酸诱导的损伤，PI3K/Akt/eNOS 和 Nrf2/ARE 途径可能是天麻素疗效的原因。
