Abstract:

Objective: To analyze the protective effect and mechanism of nuclear factor-erythroid 2-related factor 2 (Nrf2) signal pathway agonist tertiary butylhydroquinone (tBHQ) on astrocytes under oxygen glucose deprivation (OGD). Methods: Astrocytes were divided into 3 groups: the control group, the OGD group, and the tBHQ group. The cell proliferation activity after OGD and tBHQ intervention was assessed using the CCK -8 assay. Oxidative stress levels were evaluated by measuring superoxide dismutase (SOD) activity and malondialdehyde (MDA) content. The relative expression levels of pyroptosis-related genes (Caspase-1, NLRP3, IL-1β, IL 18) and antioxidant-related genes (HO-1, NQO1) were detected using real time fluorescent quantitative polymerase chain reaction (PCR). Results: Significant differences were observed among different treatment groups in cell proliferation activity (F = 8.676, P = 0.003), SOD activity (F = 5.818, P = 0.013), MDA content (F = 9.049, P = 0.004), relative expression of pyroptosis-related genes Caspase-1 (F = 17.926, P = 0.003), NLRP3 (F = 10.164, P = 0.012), IL-1β (F = 13.472, P = 0.006), IL-18 (F = 8.292, P = 0.019), and antioxidant -related genes HO-1 (F = 30.468, P = 0.001), NQO1 (F = 29.621, P = 0.001). Compared with the control group, the OGD group exhibited reduced cell proliferation activity (t = 4.114, P = 0.001) and SOD activity (t = 2.149, P = 0.029), increased MDA content (t = -2.852, P = 0.015), upregulated expression of pyroptosis-related genes Caspase-1 (t = -3.759, P = 0.009), NLRP3 (t = -4.119, P = 0.006), IL-1β (t = -4.747, P = 0.003) and IL 18 (t = -3.122, P = 0.021), and downregulated expression of antioxidant-related genes HO-1 (t = 3.816, P = 0.009) and NQO1 (t = 5.303, P = 0.002). Following tBHQ intervention, cell proliferation activity increased (t = 2.621, P = 0.019), SOD activity increased (t = 3.292, P = 0.005), MDA content decreased (t = -4.160, P = 0.001), expression of Caspase-1 (t = -5.916, P = 0.001), NLRP3 (t = -3.647, P = 0.011), IL-1β (t = -4.193, P = 0.006) and IL-18 (t = -3.825, P = 0.009) decreased, and expression of HO-1 (t = 7.805, P = 0.000) and NQO1 (t = 7.483, P = 0.000) increased. Conclusions: OGD can suppress the expression of antioxidant-related genes HO-1 and NQO1, promote astrocytes pyroptosis and oxidative stress, and inhibit cell proliferation activity. Nrf2 signal pathway agonist tBHQ can enhance the expression of HO-1 and NQO1, reduce oxidative stress in OGD-exposed astrocytes, reverse pyroptosis, and exert protective effects on the cells.

Key words: Ischemic stroke, Cell hypoxia, Glucose, Astrocytes, NF-E2-related factor 2, Cell proliferation, Pyroptosis, Cells, cultured

摘要：

目的: 探讨核因子E2相关因子2（Nrf2）信号转导通路激动剂特丁基对苯二酚（tBHQ）对氧糖剥夺星形胶质细胞的保护作用及其作用机制。方法: 将常规培养的CTX-TNA2大鼠脑Ⅰ型星形胶质细胞系分为对照组、氧糖剥夺组和tBHQ组，采用CCK-8法检测细胞增殖活性，酶标仪检测细胞氧化水平[超氧化物歧化酶（SOD）活性及丙二醛（MDA）含量]，实时荧光定量聚合酶链反应测定细胞焦亡相关基因Caspase-1、NLRP3、IL-1β、IL-18及抗氧化相关基因HO-1、NQO1的相对表达量。结果: 上述不同处理组星形胶质细胞增殖活性（F = 8.676，P = 0.003），SOD活性（F = 5.818，P = 0.013）和MDA含量（F = 9.049，P= 0.004），细胞焦亡相关基因Caspase-1（F = 17.926，P = 0.003）、NLRP3（F = 10.164，P = 0.012）、IL-1β（F = 13.472，P = 0.006）、IL-18（F = 8.292，P = 0.019）及抗氧化相关基因HO-1（F = 30.468，P = 0.001）、NQO1（F = 29.621，P = 0.001）相对表达量差异具有统计学意义，两两比较发现，氧糖剥夺后星形胶质细胞增殖活性降低（t = 4.114，P = 0.001），SOD活性降低（t = 2.149，P = 0.029），MDA含量升高（t =-2.852，P = 0.015），细胞焦亡相关基因Caspase-1（t =-3.759，P = 0.009）、NLRP3（t =-4.119，P = 0.006）、IL-1β（t =-4.747，P = 0.003）、IL-18（t =-3.122，P = 0.021）相对表达量升高，抗氧化相关基因HO-1（t = 3.816，P = 0.009）、NQO1（t = 5.303，P = 0.002）相对表达量降低；经tBHQ干预后，星形胶质细胞增殖活性升高（t = 2.621，P = 0.019），SOD活性增加（t = 3.292，P = 0.005），MDA含量降低（t =-4.160，P = 0.001），Caspase-1（t =-5.916，P= 0.001）、NLRP3（t =-3.647，P = 0.011）、IL-1β（t =-4.193，P = 0.006）、IL-18（t =-3.825，P = 0.009）相对表达量降低，HO-1（t = 7.805，P = 0.000）、NQO1（t = 7.483，P = 0.000）相对表达量升高。结论: 氧糖剥夺可抑制抗氧化相关基因HO-1和NQO1表达，促进星形胶质细胞焦亡及氧化水平，进而抑制细胞增殖活性；Nrf2通路激动剂tBHQ则可促进HO-1和NQO1基因表达，提高氧糖剥夺星形胶质细胞抗氧化水平，逆转星形胶质细胞焦亡，对细胞具有保护作用。

关键词: 缺血性卒中, 细胞低氧, 葡萄糖, 星形细胞, NF-E2相关因子2, 细胞增殖, 细胞焦亡, 细胞, 培养的