Fang S, Yuan R, Sun R, Ma T
School of Anesthesiology, Wannan Medical College, Wuhu 241002, China.
Anesthesia Laboratory and Training Center, Wannan Medical College, Wuhu 241002, China.
Nan Fang Yi Ke Da Xue Xue Bao. 2022 Dec 20;42(12):1815-1821. doi: 10.12122/j.issn.1673-4254.2022.12.09.
To investigate whether knockout of S1PR3 improves lipopolysaccharide (LPS)-induced acute lung injury in mice by inhibiting mitogen activated protein kinases (MAPKs).
Male C57BL/6J and S1PR3 knockout (S1PR3) mice were both randomized into two groups (=8) for intratracheal instillation of normal saline or LPS to induce acute lung injury. The expressions of S1PR3, IL-1β and IL-18 in the lung tissues were detected using RT-qPCR, lung tissue injury was observed with HE staining, and cell apoptosis was detected using flow cytometry. Western blotting was performed to detect the expression levels of caspase-1, GSDMD, p-JNK, p-ERK and p-p38 proteins. In the cell experiment, type II alveolar epithelial cells (MLE-12 cells) were treated with PBS, LPS, CYM5541 (a S1PR3 agonist), or CYM5541 + LPS, and the cell apoptosis and expression levels of MAPK signal pathway molecules were detected.
The expression of S1PR3 was up-regulated and serum IL-1β and IL-18 levels were elevated significantly in the nontransgenic mice with acute lung injury ( < 0.001). By comparison, the elevation of IL-1β and IL-18 levels was obviously reduced in S1PR3 knockout mice with acute lung injury, which also showed significant improvement of pulmonary hemorrhage, inflammation and exudation, lowered wet-to-dry ratio of the lungs, and decreased cell apoptosis and expressions of cleaved caspase-1 and GSDMD ( < 0.05). In MLE-12 cells, treatment with the S1PR3 agonist significantly increased the expression of pyroptosis-associated proteins ( < 0.05). S1PR3 knockout strongly inhibited the activation of MAPKs family (JNK and ERK p38; < 0.05), but their expressions were significantly increased following treatment with the S1PR3 agonist ( < 0.05).
Inhibition of S1PR3 can improve LPSinduced acute lung injury in mice by inhibiting the activation of MAPK signaling.
研究敲除S1PR3是否通过抑制丝裂原活化蛋白激酶（MAPKs）来改善脂多糖（LPS）诱导的小鼠急性肺损伤。
将雄性C57BL/6J小鼠和S1PR3基因敲除（S1PR3 -/-）小鼠均随机分为两组（n = 8），通过气管内滴注生理盐水或LPS来诱导急性肺损伤。采用RT-qPCR检测肺组织中S1PR3、IL-1β和IL-18的表达，用HE染色观察肺组织损伤情况，通过流式细胞术检测细胞凋亡。进行蛋白质免疫印迹法检测caspase-1、GSDMD、p-JNK、p-ERK和p-p38蛋白的表达水平。在细胞实验中，用PBS、LPS、CYM5541（一种S1PR3激动剂）或CYM5541 + LPS处理II型肺泡上皮细胞（MLE-12细胞），并检测细胞凋亡和MAPK信号通路分子的表达水平。
急性肺损伤的非转基因小鼠中S1PR3的表达上调，血清IL-1β和IL-18水平显著升高（P < 0.001）。相比之下，急性肺损伤的S1PR3基因敲除小鼠中IL-1β和IL-18水平的升高明显降低，同时肺出血、炎症和渗出也有显著改善，肺组织湿干比降低，细胞凋亡以及裂解的caspase-1和GSDMD的表达减少（P < 0.05）。在MLE-12细胞中，用S1PR3激动剂处理显著增加了焦亡相关蛋白的表达（P < 0.05）。S1PR3基因敲除强烈抑制了MAPKs家族（JNK、ERK和p38）的激活（P < 0.05），但在用S1PR3激动剂处理后它们的表达显著增加（P < 0.05）。
抑制S1PR3可通过抑制MAPK信号的激活来改善LPS诱导的小鼠急性肺损伤。
