Li Kexin, Gong Qianwen, Lu Bin, Huang Kaiyan, Tong Yixuan, Mutsvene Tinashe Emmanuel, Lin Meng, Xu Zhiqiang, Lu Fan, Li Xingyi, Hu Liang
National Clinical Research Center for Ocular Diseases, Eye Hospital, Wenzhou Medical University, Wenzhou, 325027, People's Republic of China.
National Engineering Research Center of Ophthalmology and Optometry, Eye Hospital, Wenzhou Medical University, Wenzhou, 325027, People's Republic of China.
Eye Vis (Lond). 2023 May 1;10(1):17. doi: 10.1186/s40662-023-00334-5.
To investigate the anti-inflammatory and antioxidative effects of gallic acid (GA) on human corneal epithelial cells (HCECs) and RAW264.7 macrophages as well as its therapeutic effects in an experimental dry eye (EDE) mouse model.
A cell counting kit-8 (CCK-8) assay was used to test the cytotoxicity of GA. The effect of GA on cell migration was evaluated using a scratch wound healing assay. The anti-inflammatory and antioxidative effects of GA in vitro were tested using a hypertonic model (HCECs) and an inflammatory model (RAW264.7 cells). The in vivo biocompatibility of GA was detected by irritation tests in rabbits, whereas the preventive and therapeutic effect of GA in vivo was evaluated using a mouse model of EDE.
In the range of 0-100 μM, GA showed no cytotoxicity in RAW264.7 cells or HCECs and did not delay the HCECs monolayer wound healing within 24 h. Ocular tolerance to GA in the in vivo irritation test was good after seven days. In terms of antioxidative activity, GA significantly reduced the intracellular reactive oxygen species (ROS) in lipopolysaccharide (LPS) activated RAW264.7 macrophages and HCECs exposed to hyperosmotic stress. Furthermore, after pre-treatment with GA, the expression levels of nuclear factor E2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), and NADPH quinone oxidoreductase-1 (NQO-1) were significantly upregulated in RAW264.7 macrophages. GA also exhibits excellent anti-inflammatory properties. This is mainly demonstrated by the ability of GA to effectively downregulate the nuclear transcription factor-κB (NF-κB) pathway in LPS-activated RAW264.7 macrophages and to reduce inflammatory factors, such as nitric oxide (NO), interleukin 6 (IL-6), and tumor necrosis factor alpha (TNF-α). In vivo efficacy testing results in a mouse model of EDE showed that GA can effectively prevent and inhibit the apoptosis of corneal epithelial cells (CECs), reduce inflammatory factors in the cornea and conjunctiva as well as protect goblet cells.
In vitro and in vivo results indicate that GA possesses potent anti-inflammatory and antioxidative properties with no apparent cytotoxicity within the range of 0-100 μM. It is a promising eye drop formulation for the effective prevention and treatment of dry eye disease (DED).
研究没食子酸（GA）对人角膜上皮细胞（HCECs）和RAW264.7巨噬细胞的抗炎和抗氧化作用及其在实验性干眼（EDE）小鼠模型中的治疗效果。
使用细胞计数试剂盒-8（CCK-8）检测GA的细胞毒性。采用划痕伤口愈合试验评估GA对细胞迁移的影响。使用高渗模型（HCECs）和炎症模型（RAW264.7细胞）检测GA在体外的抗炎和抗氧化作用。通过兔眼刺激试验检测GA的体内生物相容性，而使用EDE小鼠模型评估GA在体内的预防和治疗效果。
在0-100μM范围内，GA对RAW264.7细胞或HCECs无细胞毒性，且在24小时内不延迟HCECs单层伤口愈合。体内刺激试验中，7天后GA的眼耐受性良好。在抗氧化活性方面，GA显著降低脂多糖（LPS）激活的RAW264.7巨噬细胞和暴露于高渗应激的HCECs中的细胞内活性氧（ROS）。此外，用GA预处理后，RAW264.7巨噬细胞中核因子E2相关因子2（Nrf2）、血红素加氧酶-1（HO-1）和NADPH醌氧化还原酶-1（NQO-1）的表达水平显著上调。GA还表现出优异的抗炎特性。这主要体现在GA能够有效下调LPS激活的RAW264.7巨噬细胞中的核转录因子-κB（NF-κB）途径，并减少炎症因子，如一氧化氮（NO）、白细胞介素6（IL-6）和肿瘤坏死因子α（TNF-α）。EDE小鼠模型的体内疗效测试结果表明，GA可有效预防和抑制角膜上皮细胞（CECs）的凋亡，减少角膜和结膜中的炎症因子，并保护杯状细胞。
体外和体内结果表明，GA具有强大的抗炎和抗氧化特性，在0-100μM范围内无明显细胞毒性。它是一种有前景的滴眼剂配方，可有效预防和治疗干眼病（DED）。
