Abstract

Objective

Alcohol abuse can cause severe injury to human brain. Astrocytes are the most abundant nonneuronal cells that function to maintain the brain homeostasis. In present study, we aimed to investigate the role of ROCK2 in astrocytes exposed to alcohol.

Methods

Astrocytes were transfected with lentivirus (LV)-anti-ROCK2 vector to downregulate the expression of ROCK2. The ROCK2 expression in mRNA and protein level was analyzed by real-time PCR and Western blotting, respectively. Cytokines or indicators involved in inflammation and oxidative stress were determined by assay kits. Proteins involved in nuclear factor kappa B (NF-κB) signaling pathway and NOD-like receptor protein 3 (NLRP3) inflammasome were analyzed by Western blotting.

Results

Alcohol exposure dramatically upregulated ROCK2 expression and lactate dehydrogenase (LDH) activity in astrocytes. On the contrary, transfecting with LV-anti-ROCK2 vector downregulated ROCK2 expression and LDH activity in astrocytes, demonstrating that downregulation of ROCK2 alleviated alcohol-induced astrocytic injury. Furthermore, downregulation of ROCK2 attenuated alcohol-induced inflammation by reducing the levels of pro-inflammatory cytokines (tumor necrosis factor (TNF)-α and interleukin (IL)-6) and enhanced the level of anti-inflammatory IL-10. Downregulation of ROCK2 also attenuated alcohol-induced oxidative stress by reducing the reactive oxygen species (ROS) production, as well as enhancing the activity of anti-oxidative superoxide dismutase (SOD) and glutathione (GSH). More importantly, downregulation of ROCK2 inhibited the activation of NF-κB signaling pathway and NLRP3 inflammasome.

Conclusion

Therefore, ROCK2 could be a potential target to treat alcohol-induced astrocytic injury and the downregulation of ROCK2 might be a promising approach to protect against alcohol-induced astrocytic injury.

中文翻译：

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ROCK2的下调减弱了星形胶质细胞中酒精诱导的炎症和氧化应激

摘要

客观的

酗酒会对人的大脑造成严重伤害。星形胶质细胞是最丰富的非神经元细胞，具有维持大脑稳态的功能。在本研究中，我们旨在研究 ROCK2 在暴露于酒精的星形胶质细胞中的作用。

方法

用慢病毒 (LV)-抗 ROCK2 载体转染星形胶质细胞以下调 ROCK2 的表达。分别通过实时PCR和蛋白质印迹分析ROCK2在mRNA和蛋白质水平的表达。与炎症和氧化应激有关的细胞因子或指标由检测试剂盒确定。通过蛋白质印迹分析参与核因子 kappa B (NF-κB) 信号通路和 NOD 样受体蛋白 3 (NLRP3) 炎性体的蛋白质。

结果

酒精暴露显着上调了星形胶质细胞中的 ROCK2 表达和乳酸脱氢酶 (LDH) 活性。相反，转染 LV-anti-ROCK2 载体可下调星形胶质细胞中 ROCK2 的表达和 LDH 活性，表明 ROCK2 的下调减轻了酒精诱导的星形胶质细胞损伤。此外，ROCK2 的下调通过降低促炎细胞因子（肿瘤坏死因子 (TNF)-α 和白细胞介素 (IL)-6）的水平和提高抗炎 IL-10 的水平来减轻酒精诱导的炎症。ROCK2 的下调还通过减少活性氧 (ROS) 的产生以及增强抗氧化超氧化物歧化酶 (SOD) 和谷胱甘肽 (GSH) 的活性来减弱酒精诱导的氧化应激。更重要的是，

结论

因此，ROCK2 可能是治疗酒精诱导的星形胶质细胞损伤的潜在靶点，而 ROCK2 的下调可能是一种有希望的方法来防止酒精诱导的星形胶质细胞损伤。