刺五加皂苷VI通过抑制PI3K/AKT/NF-κB信号通路减少肠上皮细胞凋亡，从而减轻三硝基苯磺酸诱导的小鼠克罗恩病样结肠炎
[Asperosaponin VI alleviates TNBS-induced Crohn's disease-like colitis in mice by reducing intestinal epithelial cell apoptosis via inhibiting the PI3K/AKT/NF-κB signaling pathway].
作者信息
Niu Minzhu, Yin Lixia, Duan Ting, Huang Ju, Li Jing, Geng Zhijun, Hu Jianguo, Song Chuanwang
机构信息
Department of Immunology, School of Laboratory Medicine, Bengbu Medical University, Bengbu 233030, China.
Clinical Laboratory.
出版信息
Nan Fang Yi Ke Da Xue Xue Bao. 2024 Dec 20;44(12):2335-2346. doi: 10.12122/j.issn.1673-4254.2024.12.09.
OBJECTIVES
To investigate the effects of asperosaponin VI (AVI) on intestinal epithelial cell apoptosis and intestinal barrier function in a mouse model of Crohn's disease (CD)-like colitis and explore its mechanisms.
METHODS
Male C57BL/6 mice with TNBS-induced CD-like colitis were treated with saline or AVI (daily dose 150 mg/kg) by gavage for 6 days. The changes in body weight, colon length, DAI scores, and colon pathologies of the mice were observed, and the expressions of inflammatory factors and tight injunction proteins were detected using ELISA and RT-qPCR. The effects of AVI on barrier function and apoptosis of mouse intestinal epithelial cells and TNF‑α‑treated Caco-2 cells were analyzed using immunofluorescence staining, TUNEL assay, and Western blotting. Network pharmacology, TUNEL assay, and Western blotting were performed to explore and validate the therapeutic mechanisms of AVI for CD.
RESULTS
In the mouse models of CD-like colitis, AVI significantly improved body weight loss, colon shortening and DAI and tissue inflammation scores, alleviated intestinal villi and goblet cell injuries, and lowered the expressions of inflammatory factors. AVI treatment significantly reduced the loss of tight junction proteins and apoptosis in both mouse intestinal epithelial cells and TNF‑α-stimulated Caco-2 cells. KEGG enrichment pathway analysis suggested that the therapeutic effect of AVI on CD was associated with inhibition of PI3K/AKT/NF-κB pathway activation, which was confirmed by lowered expressions of p-PI3K, p-AKT, and p-p65 in AVI-treated mouse models and Caco-2 cells. In Caco-2 cells, Recilisib significantly blocked the inhibitory effect of AVI on the PI3K/AKT/NF-κB pathway and TNF-α-induced apoptosis, and AKT1 knockdown experiment confirmed the role of the PI3K/AKT pathway for mediating the activation of downstream NF-κB signaling.
CONCLUSIONS
AVI can improve TNBS-induced CD-like colitis in mice by reducing intestinal epithelial cell apoptosis and intestinal barrier damage via inhibiting the PI3K/AKT/NF-κB signaling pathway.
目的
在克罗恩病（CD）样结肠炎小鼠模型中研究远志皂苷VI（AVI）对肠上皮细胞凋亡和肠道屏障功能的影响，并探讨其作用机制。
方法
用三硝基苯磺酸（TNBS）诱导建立CD样结肠炎雄性C57BL/6小鼠模型，通过灌胃给予生理盐水或AVI（每日剂量150mg/kg），连续6天。观察小鼠体重、结肠长度、疾病活动指数（DAI）评分及结肠病理变化，采用酶联免疫吸附测定（ELISA）和逆转录-定量聚合酶链反应（RT-qPCR）检测炎症因子和紧密连接蛋白的表达。采用免疫荧光染色、TUNEL检测和蛋白质免疫印迹法分析AVI对小鼠肠上皮细胞及肿瘤坏死因子-α（TNF-α）处理的人结肠腺癌细胞（Caco-2细胞）屏障功能和凋亡的影响。运用网络药理学、TUNEL检测和蛋白质免疫印迹法探索并验证AVI治疗CD的机制。
结果
在CD样结肠炎小鼠模型中，AVI显著改善体重减轻、结肠缩短以及DAI和组织炎症评分，减轻肠绒毛和杯状细胞损伤，并降低炎症因子表达。AVI处理显著减少小鼠肠上皮细胞和TNF-α刺激的Caco-2细胞中紧密连接蛋白的丢失和细胞凋亡。京都基因与基因组百科全书（KEGG）富集通路分析表明，AVI对CD的治疗作用与抑制磷脂酰肌醇-3激酶（PI3K）/蛋白激酶B（AKT）/核因子κB（NF-κB）信号通路激活有关，这在AVI处理的小鼠模型和Caco-2细胞中通过p-PI3K、p-AKT和p-p65表达降低得到证实。在Caco-2细胞中，瑞西利西显著阻断AVI对PI3K/AKT/NF-κB信号通路的抑制作用以及TNF-α诱导的细胞凋亡，AKT1基因敲低实验证实PI3K/AKT信号通路在介导下游NF-κB信号激活中的作用。
结论
AVI可通过抑制PI3K/AKT/NF-κB信号通路减少肠上皮细胞凋亡和肠道屏障损伤，从而改善TNBS诱导的小鼠CD样结肠炎。
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