Department of Cosmetic Plastic and Burn Surgery, West China Hospital Sichuan University, Chengdu, China.
Eur Rev Med Pharmacol Sci. 2020 Feb;24(4):2077-2086. doi: 10.26355/eurrev_202002_20387.
Previous studies have shown that Quinazoline (QNZ) plays extremely important roles in the cellular physiological activity, but it has been rarely examined on cell behavior following intervertebral disc degeneration (IVDD). The aim of this study was to investigate whether QNZ mediates oxidative stress and inflammation contributed to IL-1β-induced nucleus pulposus (NP) cells degeneration in vitro.
NP were isolated cells from human disc samples collected from patients and the IL-1β-induced NP cells degenerated model was constructed. The cells were randomly divided into 3 groups, namely, Control group, IL-1β group (10 µM), QNZ + IL-1β group (containing 10 nM QNZ and 10 µM IL-1β). Then, the cell viability was determined by CCK-8 assay, and the levels of collagen I, collagen II, aggrecan, p16, p53, β-galactosidase (β-gal), antioxidant enzymes, 8-hydroxy-2-deoxyguanosine (8-OHdG), NF-κB/MAPKs signaling-related proteins and inflammatory factors were examined using Western blot and reverse transcription-quantitative polymerase chain reaction (RT-qPCR) in NP cells. Finally, the expressions of IL-1β, IL-6, and TNF-α in the cell supernatants were also determined by enzyme-linked immunosorbent assay (ELISA).
This study showed that IL-1β promoted the progress of IDD, with markedly increased expressions of collagen I, p16, p53, and β-gal, as well as decreased expressions of collagen II and aggrecan. However, QNZ treatment could reverse the effects of IL-1β. It was found that cell proliferation was increased, ROS level was decreased, antioxidant enzymes were upregulated, and inflammatory factors were reduced after QNZ stimulation. Moreover, NF-κB/MAPKs signaling proteins IKKβ, IκBα, p65, ERK, JNK, and p38 were significantly dephosphorylated by QNZ.
These results indicated that QNZ prevented NP degradation via restraining oxidative stress and inflammation through inhibition of the NF-κB/MAPKs signaling pathway. QNZ may become a novel insight into the therapy of IVDD in the future.
已有研究表明，喹唑啉（QNZ）在细胞生理活动中发挥着极其重要的作用，但在椎间盘退变（IVDD）后细胞行为方面的研究却很少。本研究旨在探讨 QNZ 是否通过调节氧化应激和炎症来介导白细胞介素 1β（IL-1β）诱导的体外髓核（NP）细胞退变。
从患者椎间盘样本中分离 NP 细胞，并构建 IL-1β诱导的 NP 细胞退变模型。细胞随机分为 3 组，分别为对照组、IL-1β 组（10 μM）、QNZ+IL-1β 组（含 10 nM QNZ 和 10 μM IL-1β）。采用 CCK-8 法检测细胞活力，采用 Western blot 和逆转录定量聚合酶链反应（RT-qPCR）检测 NP 细胞中Ⅰ型胶原、Ⅱ型胶原、聚集蛋白聚糖、p16、p53、β-半乳糖苷酶（β-gal）、抗氧化酶、8-羟基-2-脱氧鸟苷（8-OHdG）、核因子-κB/丝裂原活化蛋白激酶（NF-κB/MAPKs）信号相关蛋白及炎症因子水平。最后，采用酶联免疫吸附试验（ELISA）检测细胞上清液中白细胞介素 1β（IL-1β）、白细胞介素 6（IL-6）和肿瘤坏死因子-α（TNF-α）的表达。
本研究表明，IL-1β 可促进 IDD 进展，使Ⅰ型胶原、p16、p53 和β-gal 表达增加，而使Ⅱ型胶原和聚集蛋白聚糖表达减少。然而，QNZ 处理可逆转 IL-1β 的作用。结果发现，QNZ 刺激后细胞增殖增加，ROS 水平降低，抗氧化酶上调，炎症因子减少。此外，QNZ 还可显著磷酸化 NF-κB/MAPKs 信号通路蛋白 IKKβ、IκBα、p65、ERK、JNK 和 p38。
这些结果表明，QNZ 通过抑制 NF-κB/MAPKs 信号通路抑制氧化应激和炎症，防止 NP 降解。QNZ 可能成为未来治疗 IVDD 的新方法。
