银杏黄酮通过抑制核因子κB途径减轻硫酸葡聚糖钠诱导的结肠炎

摘要(中文译文)

银杏双黄酮是从银杏叶中分离出的一种双黄酮类化合物，在结肠炎中已显示出抗炎作用。然而，其分子机制仍不清楚。本研究探讨了银杏双黄酮对葡聚糖硫酸钠（DSS）诱导的小鼠结肠炎及脂多糖（LPS）处理的Caco-2细胞的治疗作用。通过免疫荧光、逆转录定量聚合酶链反应和Western blotting评估炎症、结肠上皮屏障完整性及细胞损伤。此外，通过过表达或敲低p65来检验其在这些效应中的参与作用。银杏双黄酮治疗显著减轻了DSS诱导小鼠的体重下降、疾病活动指数和组织病理学损伤。它在体内和体外均下调了促炎细胞因子，如白细胞介素-6（IL-6）、肿瘤坏死因子-α（TNF-α）和白细胞介素-1β（IL-1β），并上调了紧密连接蛋白，如闭锁小带蛋白-1（ZO-1）和Occludin。值得注意的是，银杏双黄酮抑制了结肠组织和Caco-2细胞中的p-p65和C-X-C基序趋化因子配体2。分子对接、生物素下拉实验和细胞热位移实验提示银杏双黄酮与p65之间存在相互作用。p65过表达逆转了银杏双黄酮对LPS诱导的炎症和屏障破坏的抑制作用。p65敲低使LPS诱导的改变正常化，下调了促炎细胞因子（IL-6、TNF-α和IL-1β），并上调了紧密连接蛋白（ZO-1和Occludin）。值得注意的是，加入银杏双黄酮并未进一步改善单独敲低p65的有益效应。这些发现表明，银杏双黄酮通过核因子κB通路调节炎症和上皮屏障功能障碍，从而减轻结肠炎。本研究证明了银杏双黄酮作为结肠炎治疗药物的潜力。

Abstract

Ginkgetin is a biflavone isolated from Ginkgo biloba leaves. It has shown anti-inflammatory effects in colitis. However, its molecular mechanisms remain unclear. This study investigated the therapeutic effects of ginkgetin on dextran sulfate sodium (DSS)-induced colitis in mice and lipopolysaccharide (LPS)-treated Caco-2 cells. Inflammation, the integrity of the colonic epithelial barrier, and cellular damage were assessed via immunofluorescence, reverse transcription quantitative polymerase chain reaction, and Western blotting. Furthermore, p65 was overexpressed or knocked down to examine its involvement in the observed effects. Treatment with ginkgetin significantly attenuated weight loss, disease activity index, and histopathological damage in DSS-induced mice. It also downregulated pro-inflammatory cytokines, such as interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), and interleukin-1β (IL-1β), and upregulated tight junction proteins, such as zonula occludens-1 (ZO-1) and Occludin, in vivo and in vitro. Notably, ginkgetin inhibited p-p65 and C-X-C motif chemokine ligand 2 in colonic tissues and Caco-2 cells. Molecular docking, biotin pull-down, and cellular thermal shift assays suggested an interaction between ginkgetin and p65. p65 overexpression reversed the inhibitory effects of ginkgetin on LPS-induced inflammation and barrier disruption. p65 knockdown normalized LPS-induced alterations, downregulated pro-inflammatory cytokines (IL-6, TNF-α, and IL-1β), and upregulated tight junction proteins (ZO-1 and Occludin). Notably, the addition of ginkgetin did not further improve the beneficial effects of p65 knockdown alone. These findings suggest that ginkgetin alleviated colitis by modulating inflammation and epithelial barrier dysfunction through the nuclear factor kappa B pathway. This study demonstrates the potential of ginkgetin as a therapeutic agent for colitis.

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