OSI-027抑制mTOR信号通路减轻高氧诱导的幼鼠肺损伤

OSI-027 alleviates hyperoxia-induced lung injury in juvenile rats by inhibiting the mTOR signaling pathway

目的:探讨哺乳动物雷帕霉素靶蛋白(mammalian target of rapamycin,mTOR)1/2 双重抑制剂 OSI-027 对高体积分数氧(高氧)致 sprague-dawley(SD)幼鼠肺损伤及纤维化的抑制作用。方法:72 只 3 周龄 SD 幼鼠随机分为空气+生理盐水、高氧+生理盐水、高氧+雷帕霉素和高氧+OSI-027 组,分别建立动物模型(各组 n=18)。高氧干预采用 90%氧气持续处理,生理盐水、雷帕霉素、OSI-027 干预分别于观察期第 1、3、6、8、10、13 天经腹腔注射给药,在造模第 3、7、14 天取各组幼鼠测量体质量变化、肺湿干重比(wet/dry ratio,W/D)、肺组织病理学检查、肺损伤评分、肺泡间隔厚度测定、肺组织免疫组化和蛋白印迹检测 mTOR 及磷酸化核糖体 S6 蛋白激酶(pS6K1)蛋白在肺组织的分布和表达。结果:从时间因素看,各组幼鼠体质量(F 时间=297.098,P=0.000)、mTOR 免疫组化(F 时间=379.978,P=0.000)、mTOR(F 时间=166.991,P=0.000)和 pS6K1(F 时间=122.676,P=0.000)蛋白水平都随时间延长而增加。除空气组外,其余各组肺损伤评分(F 时间=1410.362,P=0.000)、肺泡间隔厚度(F 时间=356.312,P=0.000)、pS6K1 免疫组化(F 时间=57.992,P=0.000)都随时间延长而升高,肺 W/D(F 时间=28.915,P=0.000)第 3、7 天时升高,第 14 天时下降。从分组因素看,体质量(F 分组=176.597,P=0.000)空气组明显高于其他组,肺 W/D(F 分组=28.484,P=0.000)和肺泡间隔厚度(F 分组=296.223, P=0.000)空气组明显低于其他组,除第 3 天外,mTOR 免疫组化(F 分组=134.100,P=0.000)高氧组明显高于其他组,PS6K1 免疫组化(F 分组=234.697,P=0.000)、mTOR(F 分组=59.377,P=0.000)和 PS6K1(F 分组=101.837,P=0.000)蛋白印迹高氧组明显高于其他组,肺损伤评分(F 分组=2 420.076,P=0.000)高氧雷帕组明显高于其他组,高氧 OSI 组明显低于高氧组和高氧雷帕组。结论:高浓度氧可激活肺组织 mTOR 信号途径;mTOR 可能促进了高氧肺损伤纤维化的发生发展,其调控机制可能与抑制 mTOR 信号通路的活化有关。 mTOR 复合物 1/2(mTORC1/2)双重抑制剂 OSI-027 能减轻高氧致 SD 幼鼠肺损伤及纤维化。

Objective:To investigate the inhibitory effect of the mammalian target of rapamycin(mTOR) 1/2 dual inhibitor OSI-027 on hyperoxia-induced lung injury and fibrosis in juvenile Sprague-Dawley (SD) rats. Methods:A total of 72 juvenile SD rats aged 3 weeks were randomly divided into air+normal saline group,hyperoxia+normal saline group,hyperoxia+rapamycin group,and hyperoxia+ OSI-027 group,with 18 rats in each group. An animal model was established. Hyperoxia intervention was performed with 90% oxy gen,and normal saline,rapamycin,and OSI-027 interventions were performed via intraperitoneal injection on days 1,3,6,8,10,and 13 of observation,respectively. On days 3,7,and 14,the change in body weight,lung wet/dry(W/D) ratio,lung injury scores,and alve olar septal thickness were measured;lung histopathological examination was performed;immunohistochemistry and Western blot were used to evaluate the distribution and expression of mTOR and phosphorylated ribosomal S6 kinase(pS6K1) in lung tissue. Results:As for the factor of time,there were significant increases over time in body weight (Ftime=297.098,P=0.000), immunohistochemistry of mTOR(Ftime=379.978,P=0.000),mTOR(Ftime=166.991,P=0.000),and pS6K1(Ftime=122.676,P=0.000). All groups except the air+normal saline group had significant increases in lung injury scores(Ftime=1 410.362,P=0.000),alveolar septum thickness(Ftime=356.312,P=0.000),and pS6K1 immunohistochemistry(Ftime=57.992,P=0.000) over time,as well as an increase in lung W/D ratio on days 3 and 7(Ftime=28.915,P=0.000) and a reduction in lung W/D ratio on day 14. As for the factor of grouping,the air+normal saline group had a significantly higher body weight(Fgroup= 176.597,P=0.000) and significantly lower lung W/D ratio(Fgroup=28.484,P=0.000) and alveolar septum thickness(Fgroup=296.223,P= 0.000) than the other groups. At all time points except day 3,the hyperoxia+normal saline group had significantly higher mTOR immunohistochemistry(Fgroup=134.100,P=0.000),pS6K1 immunohistochemistry(Fgroup=234.697,P=0.000),mTOR(Fgroup=59.377,P=0.000), and pS6K1(Fgroup=101.837,P=0.000) than the other groups;the hyperoxia+rapamycin group had significantly higher lung injury scores than the other groups(Fgroup=2 420.076,P=0.000),and the hyperoxia+OSI-027 group had significantly lower scores than the hyperox- ia+normal saline group and the hyperoxia+rapamycin group. Conclusion:A high concentration of oxygen can activate the mTOR sig- naling pathway in lung tissue;mTOR may promote the development and progression of hyperoxia-induced pulmonary fibrosis,possibly by inhibiting activation of the mTOR signaling pathway. OSI-027 can alleviate hyperoxia-induced lung injury and fibrosis in juvenile SD rats.