Chen Shanshan, He Ruonan, Li Ying, Zhang Shuo
The First Affiliated Hospital of Zhejiang Chinese Medical University (Zhejiang Provincial Hospital of Traditional Chinese Medicine), Hangzhou, 310053, Zhejiang, China.
The First Clinical Medical College, Zhejiang Chinese Medical University, Hangzhou, 310053, Zhejiang, China.
Heliyon. 2024 Apr 26;10(9):e29797. doi: 10.1016/j.heliyon.2024.e29797. eCollection 2024 May 15.
Non-steroid anti-inflammatory drugs (NSAIDs) are a class of prescription drugs with antipyretic, analgesic, anti-inflammatory, and antiplatelet effects. However, long-term use of NSAIDs will disrupt the intestinal mucosal barrier, causing erosion, ulcers, bleeding, and even perforation. Pure total flavonoids from Citrus (PTFC) is extracted from the dried peel of Citrus, showing a protective effect on intestinal mucosal barrier with unclear mechanisms.
In the present study, we used diclofenac (7.5 mg kg, i.g.) to induce a rat model of NSAIDs-related intestinal lesions. PTFC (50, 75, 100 mg·kg-1 d-1, i.g.) was administered 9 days before the initial diclofenac administration, followed by co-administration on the last 5 days. Exosomes were identified by western blotting and transmission electron microscopy (TEM), and then co-cultured with IEC-6 cells. The expression of long non-coding RNA (lncRNA) H19, autophagy-related 5 (Atg5), ZO-1, Occludin, and Claudin-1 were detected by quantitative real-time PCR (qRT-PCR). The expression of light chain 3 (LC3)-I, LC3-II, ZO-1, Occludin and Claudin-1 proteins was tested by western blotting. The localization of both exosomes and autophagosomes was examined by immunofluorescent technique.
The treatment of PTFC attenuated intestinal mucosal mechanical barrier function disturbance in diclofenac-induced NSAIDs rats. IEC-6 cells co-cultured with NSAIDs rats-derived exosomes possessed the lowest levels of protective autophagy, and severe intestinal barrier injuries. Cells co-cultured with the exosomes extracted from rats administrated PTFC exhibited an improvement of autophagy and intestinal mucosal mechanical barrier function. The prevention effect was proportional to the concentration of PTFC administered.
PTFC ameliorated NSAIDs-induced intestinal mucosal injury by down-regulating exosomal lncRNA H19 and promoting autophagy.
非甾体抗炎药（NSAIDs）是一类具有解热、镇痛、抗炎和抗血小板作用的处方药。然而，长期使用NSAIDs会破坏肠道黏膜屏障，导致糜烂、溃疡、出血甚至穿孔。柑橘纯总黄酮（PTFC）是从柑橘干燥果皮中提取的，对肠道黏膜屏障具有保护作用，但其机制尚不清楚。
在本研究中，我们使用双氯芬酸（7.5毫克/千克，腹腔注射）诱导大鼠NSAIDs相关肠道损伤模型。在首次给予双氯芬酸前9天给予PTFC（50、75、100毫克·千克-1·天-1，腹腔注射），随后在最后5天联合给药。通过蛋白质免疫印迹法和透射电子显微镜（TEM）鉴定外泌体，然后与IEC-6细胞共培养。通过定量实时PCR（qRT-PCR）检测长链非编码RNA（lncRNA）H19、自噬相关蛋白5（Atg5）、紧密连接蛋白1（ZO-1）、闭合蛋白（Occludin）和Claudin-1的表达。通过蛋白质免疫印迹法检测微管相关蛋白1轻链3（LC3）-I、LC3-II、ZO-1、Occludin和Claudin-1蛋白的表达。通过免疫荧光技术检测外泌体和自噬体的定位。
PTFC治疗减轻了双氯芬酸诱导的NSAIDs大鼠肠道黏膜机械屏障功能障碍。与NSAIDs大鼠来源的外泌体共培养的IEC-6细胞具有最低水平的保护性自噬和严重的肠道屏障损伤。与从给予PTFC的大鼠中提取的外泌体共培养的细胞表现出自噬和肠道黏膜机械屏障功能的改善。预防效果与PTFC给药浓度成正比。
PTFC通过下调外泌体lncRNA H19和促进自噬改善NSAIDs诱导的肠道黏膜损伤。
