Qin Yi, Cao Lirong, Hu Lili
ICU, Jingzhou Hospital, Tongji Medical College, Huazhong University of Science and Technology (HUST), Jingzhou, China.
Medical Department, Hubei College of Chinese Medicine, Jingzhou, China.
J Cell Biochem. 2019 Jul;120(7):11305-11317. doi: 10.1002/jcb.28407. Epub 2019 Feb 19.
Nuclear factor erythroid 2-related factor 2 (Nrf2) protects the lung from sepsis-induced injury through activating Nrf2-regulated multiple phase 2 detoxification genes, including NAD(P)H: quinine oxidoreductase-1 (NQO1) and heme oxygenase-1 (HO1). Based on the positive effect of Sirtuin 6 on Nrf2, we aim to explore the potential role of SIRT6 in the mechanism of sepsis-induced acute lung injury (ALI).
Mouse models of sepsis were constructed by instilling intratracheal of lipopolysaccharide (LPS; 4 ml/kg). After 48-hour treatment, lung tissues were collected to measure the degree of lung injury. The SIRT6, siSIRT6, and siNrf2 plasmids were cotransfected into various concentrations of LPS-treated human umbilical vein endothelial cells (HUVECs; 0, 1, 5, 10, and 50 μg/ml) using Lipofectamine 2000. Tumor necrosis factor-α (TNF-α) and interleukin (IL)-6 levels were determined by enzyme-linked immunosorbent assay. Expression levels of SIRT6, Nrf2, NQO1, and HO1 was measured by quantitative polymerase chain reaction and Western blot analysis. Cell apoptosis was determined by flow cytometry.
Lung tissues in the model group already had basic characteristics of ALI. Compared with the control model, TNF-α and IL-6 levels were much higher (P < 0.01), the levels of SIRT6, Nrf2, and Nrf2-modulated detoxification factors were downregulated (P < 0.01). SIRT6 overexpression decreased the apoptosis below to 10% (P < 0.01), significantly increased the Nrf2 expression, effectively inhibited TNF-α and IL-6 releases, and enhanced NQO1 and HO1 levels (P < 0.01). siNrf2 abolished the protective effects of SIRT6 overexpression, including increasing apoptosis and inhibiting anti-inflammatory and antioxidative genes expressions (P < 0.01).
Our study suggested SIRT6 positively regulated Nrf2 expression and activated Nrf2-regulated anti-inflammatory and antioxidative enzymes, which could effectively mitigate LPS-induced HUVECs inflammatory responses. This might reflect the mechanism of ALI induced by sepsis.
核因子红细胞2相关因子2（Nrf2）通过激活Nrf2调控的多个二期解毒基因，包括NAD（P）H：醌氧化还原酶-1（NQO1）和血红素加氧酶-1（HO1），保护肺免受脓毒症诱导的损伤。基于沉默调节蛋白6（Sirtuin 6）对Nrf2的积极作用，我们旨在探讨SIRT6在脓毒症诱导的急性肺损伤（ALI）机制中的潜在作用。
通过气管内注入脂多糖（LPS；4 ml/kg）构建脓毒症小鼠模型。治疗48小时后，收集肺组织以测量肺损伤程度。使用Lipofectamine 2000将SIRT6、siSIRT6和siNrf2质粒共转染到不同浓度LPS处理的人脐静脉内皮细胞（HUVECs；0、1、5、10和50 μg/ml）中。通过酶联免疫吸附测定法测定肿瘤坏死因子-α（TNF-α）和白细胞介素（IL）-6水平。通过定量聚合酶链反应和蛋白质免疫印迹分析测量SIRT6、Nrf2、NQO1和HO1的表达水平。通过流式细胞术测定细胞凋亡。
模型组肺组织已具有ALI的基本特征。与对照模型相比，TNF-α和IL-6水平高得多（P < 0.01），SIRT6、Nrf2和Nrf2调节的解毒因子水平下调（P < 0.01）。SIRT6过表达使细胞凋亡率降至10%以下（P < 0.01），显著增加Nrf2表达，有效抑制TNF-α和IL-6释放，并提高NQO1和HO1水平（P < 0.01）。siNrf2消除了SIRT6过表达的保护作用，包括增加细胞凋亡和抑制抗炎和抗氧化基因表达（P < 0.01）。
我们的研究表明，SIRT6正向调节Nrf2表达并激活Nrf2调节的抗炎和抗氧化酶，这可以有效减轻LPS诱导的HUVECs炎症反应。这可能反映了脓毒症诱导ALI的机制。
