摘要：

目的: 以水杨酸和硫脲为原料通过化学反应合成小分子药物碳纳米点（carbon nanodots, CDots），探讨其对脂多糖（lipopolysaccharide, LPS）诱导的细胞炎症模型的抗炎作用，并结合化学表征分析其机制。方法: 通过微波加热的方法一步合成水杨酸和硫脲碳纳米点，并通过透射电子显微镜（transmission electron microscopy, TEM）、傅里叶变换红外光谱（Fourier transform infrared, FTIR）对合成碳纳米点进行表征，通过细胞计数试剂盒-8（cell counting kit-8，CCK-8）实验检测碳纳米点的生物安全性，通过实时荧光定量聚合酶链反应（real-time quantitative polymerase chain reaction，RT-qPCR）法和蛋白质免疫印迹法（Western blotting）检测经碳纳米点处理后，小鼠巨噬细胞RAW264.7中炎症细胞因子肿瘤坏死因子-α（tumor necrosis factor-α，TNF-α）和白细胞介素-1β（interleukin-1β，IL-1β）的表达。结果: TEM显示，碳纳米点平均直径为3.5 nm，分布均匀，晶格结构明显。FTIR结果证明，碳纳米点中有酰胺键存在。在CCK-8实验中，与对照组相比较，浓度为10、20、50、100、200 μg/mL的碳纳米点处理细胞后，细胞存活率均保持在85%以上（P>0.05）。RT-qPCR和Western blotting检测LPS诱导的炎症模型中炎症因子的表达，结果显示，相对于对照组（经过LPS处理，未经碳纳米点处理），碳纳米点处理组炎症细胞因子TNF-α和IL-1β的表达明显降低（P<0.05）。结论: 以水杨酸和硫脲为原料合成的小分子药物碳纳米点由于其独特的小尺寸、良好的生物相容性和明显的抗炎特性，可以较好地应用于生物抗炎。

关键词: 碳纳米点, 水杨酸, 酰胺键, 抗炎作用

Abstract:

Objective: The present study describes using salicylic acid and thiourea as raw materials to synthesize small molecular pharmaceutical carbon nanodots(CDots) through chemical reactions. Explore its anti-inflammatory effect on lipopolysaccharide(LPS)-induced cell inflammation model, and clarify the mechanism by chemical characterization. Methods: The salicylic acid and thiourea-based CDots were synthesized in one step by microwave heating, and the synthesized CDots were characterized by transmission electron microscopy (TEM) and Fourier transform infrared (FTIR) characterizations. The biocompatibility of CDots was detected by cell counting kit-8 (CCK-8) test. And the expression of inflammation cytokines tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) in mouse macrophage RAW264.7 cells after CDots treatment was detected by real-time quantitative polymerase chain reaction (RT-qPCR) and Western blotting assays. Results: TEM image showed that the CDots are well dispersed with an average diameter of 3.5 nm and well-resolved lattice spacing. FTIR characterizations proved that there are amide bonds in the CDots. In the CCK-8 assay, compared with the control group, the viabilities of cells treated with CDots of 10, 20, 50, 100 and 200 μg/mL remained above 85% (P>0.05); RT-qPCR and Western blotting were used to detect the expression of inflammation cytokines in the LPS-induced inflammation model. Compared with the control group, the expression of inflammation cytokines TNF-α and IL-1β in the CDots treatment group reduced significantly(P<0.05). Conclusion: It is concluded that the synthesized small-molecule pharmaceutical CDots synthesized with salicylic acid and thiourea owing to its unique small size and excellent biocompatibility demonstrated anti-inflammation potentials in vitro.

Key words: carbon nanodots, salicylic acid, amide bond, anti-inflammation effects