Liu Shasha, Dong Shu'an, Shi Jia, Liu Huayang, Gao Qiaoying, Yu Jianbo
Department of Anesthesiology and Critical Care Medicine, Tianjin NanKai Hospital, Tianjin Medical University, Institute of Integrative Medicine for Acute Abdominal Diseases, Tianjin Key Laboratory of Acute Abdomen Disease Associated Organ Injury and ITCWM Repair, Tianjin 300100, China.
Department of Laboratory, Tianjin NanKai Hospital, Tianjin Medical University, Institute of Integrative Medicine for Acute Abdominal Diseases, Tianjin Key Laboratory of Acute Abdomen Disease Associated Organ Injury and ITCWM Repair, Tianjin 300100, China. Corresponding author: Yu Jianbo, Email:
Zhonghua Wei Zhong Bing Ji Jiu Yi Xue. 2024 Apr;36(4):377-380. doi: 10.3760/cma.j.cn121430-20231204-01036.
To investigate the effect of nuclear factor E2-related factor 2 (Nrf2) on the cellular tight junction protein Claudin-18 in endotoxin-induced acute lung injury (ALI).
Eighteen healthy male C57BL/6 mice were divided into control group, endotoxin-induced ALI model group (ALI group) and Nrf2 activator tert-butylhydroquinone (tBHQ) pretreatment group (tBHQ+ALI group) according to random number table method, with 6 mice in each group. Mice endotoxin-induced ALI model was reproduced by intraperitoneal injection of lipopolysaccharide (LPS, 15 mg/kg), and the mice in the control group was injected with an equal amount of phosphate buffer solution (PBS). The mice in the tBHQ+ALI group received three intraperitoneal injections of tBHQ (a total of 50 mg/kg) at an interval of 1 hour before molding. The last injection of tBHQ was accompanied by LPS of 15 mg/kg. The mice in the control group and model group were given equal amounts of PBS, and PBS or LPS was given at the last injection. The mice were sacrificed at 12 hours after LPS injection to take lung tissues. After the lung tissue was stained with hematoxylin-eosin (HE) staining, the pathological changes were observed under light microscopy, and the lung injury score was calculated. The lung wet/dry ratio (W/D) was determined. Nrf2 protein expression in the lung tissue was detected by Western blotting. Positive expression of Claudin-18 in the lung tissue was determined by immunohistochemistry.
The lung tissue showed normal structure, without significant pathological change in the control group. Compared with the control group, the alveolar septum widened accompanied by inflammatory cell infiltration, capillary hyperemia and tissue edema in the ALI group, the lung injury score and lung W/D ratio were significantly increased (lung injury score: 6.50±1.05 vs. 1.83±0.75, lung W/D ratio: 3.79±0.22 vs. 3.20±0.14, both P < 0.01), and the Nrf2 protein expression and Claudin-18 positive expression in the lung tissue were significantly lowered [Nrf2 protein (Nrf2/β-actin): 0.41±0.33 vs. 1.22±0.33, Claudin-18 (A value): 0.28±0.07 vs. 0.44±0.10, both P < 0.05]. After tBHQ pretreatment, the degree of lung histopathological injury was significantly reduced compared with the ALI group, the alveolar space slightly abnormal, inflammatory cell infiltration and tissue edema reduced, the lung injury score and lung W/D ratio were significantly decreased (lung injury score: 3.00±0.89 vs. 6.50±1.05, lung W/D ratio: 3.28±0.19 vs. 3.79±0.22, both P < 0.01), and Nrf2 protein expression and Claudin-18 positive expression in the lung tissue were significantly increased [Nrf2 protein (Nrf2/β-actin): 1.26±0.09 vs. 0.41±0.33, Claudin-18 (A valure): 0.45±0.04 vs. 0.28±0.07, both P < 0.05].
Nrf2 alleviated pulmonary edema and improved endotoxin-induced ALI by up-regulating Claudin-18 expression.
探讨核因子E2相关因子2（Nrf2）对内毒素诱导的急性肺损伤（ALI）中细胞紧密连接蛋白Claudin-18的影响。
将18只健康雄性C57BL/6小鼠按随机数字表法分为对照组、内毒素诱导的ALI模型组（ALI组）和Nrf2激活剂叔丁基对苯二酚（tBHQ）预处理组（tBHQ+ALI组），每组6只。通过腹腔注射脂多糖（LPS，15 mg/kg）复制小鼠内毒素诱导的ALI模型，对照组小鼠注射等量的磷酸盐缓冲液（PBS）。tBHQ+ALI组小鼠在造模前1小时每隔1小时腹腔注射3次tBHQ（共50 mg/kg）。最后一次注射tBHQ时同时注射15 mg/kg的LPS。对照组和模型组小鼠给予等量的PBS，最后一次注射时给予PBS或LPS。LPS注射后12小时处死小鼠取肺组织。肺组织经苏木精-伊红（HE）染色后，在光学显微镜下观察病理变化，并计算肺损伤评分。测定肺湿/干比（W/D）。采用蛋白质免疫印迹法检测肺组织中Nrf2蛋白表达。采用免疫组织化学法测定肺组织中Claudin-18的阳性表达。
对照组肺组织结构正常，无明显病理变化。与对照组相比，ALI组肺泡间隔增宽，伴有炎性细胞浸润、毛细血管充血和组织水肿，肺损伤评分和肺W/D比值显著升高（肺损伤评分：6.50±1.05比1.83±0.75，肺W/D比值：3.79±0.22比3.20±0.14，均P<0.01），肺组织中Nrf2蛋白表达和Claudin-18阳性表达显著降低[Nrf2蛋白（Nrf2/β-肌动蛋白）：0.41±0.33比1.22±0.33，Claudin-18（A值）：0.28±0.07比0.44±0.10，均P<0.05]。tBHQ预处理后，与ALI组相比，肺组织病理损伤程度明显减轻，肺泡腔轻度异常，炎性细胞浸润和组织水肿减轻，肺损伤评分和肺W/D比值显著降低（肺损伤评分：3.00±0.89比6.50±1.05，肺W/D比值：3.28±0.19比3.79±0.22，均P<0.01），肺组织中Nrf2蛋白表达和Claudin-18阳性表达显著增加[Nrf2蛋白（Nrf
