摘要
观察和评价分析土木香内酯对肺纤维化小鼠中介导细胞自噬的mTOR蛋白的调控机制。将60只昆明小鼠随机分为对照组、模型组、土木香内酯高浓度组、土木香内酯中浓度组、土木香内酯低浓度组和强的松组,每组10只。通过采用博来霉素(BLM)来建立模型,对照组小鼠注射0.9%的生理盐水,造模成功后,对相应的分组小鼠进行不同的处理,对照组和模型组给予0.9%的生理盐水[0.01 g/(kg·d)],土木香内酯高浓度组、土木香内酯中浓度组、土木香内酯低浓度组分别给予不同浓度的土木香内酯,强的松组给予强的松[0.455 g/(kg·d)]灌胃,于造模后d 7,14,28分批提取样本。采用苏木素-伊红(HE)及Masson染色观察小鼠肺泡炎和纤维化程度;蛋白免疫印迹法(Western blot)检测小鼠肺组织mTOR,核糖体S6,微管相关蛋白1轻链3Ⅱ(MAP1 LC3-Ⅱ)的表达;电镜检测观察小鼠肺组织自噬情况。与对照组比较,模型组小鼠在7,14,28 d肺泡炎及肺纤维化显著(P<0.01);相对于模型组,土木香内酯高浓度组、土木香内酯中浓度组、土木香内酯低浓度组和强的松组对小鼠肺泡炎和纤维化程度的改善程度有差异,差异有统计学意义(P<0.05)。相对于对照组,其它各组肺组织mTOR,S6蛋白表达水平均升高,小鼠肺组织LC3-Ⅱ表达水平降低。但相对于模型组,土木香内酯高浓度组、土木香内酯中浓度组、土木香内酯低浓度组肺组织mTOR,S6蛋白表达水平降低,差异有统计学意义(P<0.05)。土木香内酯高浓度组、土木香内酯中浓度组、土木香内酯低浓度组肺组织LC3-Ⅱ表达水平升高,差异有统计学意义(P<0.05)。电镜检测观察小鼠肺组织显示,对照组未见自噬,而模型组细胞形态最差,存在个别自噬。通过本研究表明,土木香内酯对博来霉素所致小鼠肺纤维化能够起到治疗的效果,可能与其降低介导细胞自噬的mTOR蛋白表达有关。
To observe and evaluate the regulation mechanism of Alantolactone on mTOR protein mediating autophagy in pulmonary fibrosis mice, 60 Kunming mice were randomly divided into control group, model group, Alantolactone high concentration group, Alantolactone medium concentration group, Alantolactone low concentration group and predtisone group, with 10 mice in each group.The model was established by using bleomycin(BLM),and mice in the control group were injected with 0.9% normal saline.After successful modeling, mice in the corresponding groups were treated with different treatments.The control group and model group were given 0.9% normal saline [0.01 g/(kg/d)],high concentration group, medium concentration group, low concentration group were given different concentrations of Alantolactone, prednisone group was given prednisone [0.455 g/(kg/d)] to fill the stomach, samples were collected in batches at 7,14,28 days after moulding.The degree of alveolitis and fibrosis in mice was observed by Hematoxylin-Eosin(HE) and Masson staining.Protein immunoblot method(Western blot) was used to detect lung tissue of mice mTOR,ribosomal S6,light chain 3 Ⅱ microtubule associated protein 1(MAP1 LC3-Ⅱ) expression.The autophagy of lung tissue in mice was observed by electron microscopy.Results showed compared with the control group, alveolitis and pulmonary fibrosis in model group were significant on d 7,14 and 28(P<0.01).Compared with the model group, the improvement degree of alveolar inflammation and fibrosis in mice in the high concentration group, medium concentration group, low concentration group and prednisone group were different, with statistical significance(P<0.05).Compared with the control group, other groups of lung tissue mTOR,S6 protein expression levels were increased, lung tissue LC3-Ⅱ expression levels were decreased.However, compared with the model group, the expression levels of mTOR and S6 protein in the lung tissues of the high concentration group, medium concentration group and low concentration group were decreased, and the difference was statistically significant(P<0.05).LC3-lung tissue Ⅱ expression levels in the high concentration group, medium concenoation group and low concentration group were increased, the difference was statistically significant(P < 0.05).Electron microscopic examination of the lung tissue of mice showed that no autophagy was observed in the control group, while the cell morphology in the model group was the worst, with individual autophagy.This study indicates that Alantolactone has a therapeutic effect on bleomycin-induced pulmonary fibrosis in mice, which may be related to the decrease of the expression of mediating autophagy mTOR protein.
作者
宋倩男
李淼
李贵星
邓琳千
张明浩
冯大庆
李可心
陈宏
SONG Qian-nan;LI Miao;LI Gui-xing;DENG Lin-qian;ZHANG Ming-hao;FENG Da-qing;LI Ke-xin;CHEN Hong(Department of Pediarics,First Afiliated Hospial of Heilongiang Universily of TCM,Harbin 150040,China)
出处
《药物生物技术》
CAS
2022年第3期245-249,共5页
Pharmaceutical Biotechnology
基金
中国博士后科学基金第13批特别资助(No.2020T130178)
黑龙江省博士后科研启动金资助项目(No.LBH-Q20178)
黑龙江省自然科学基金项目(No.H2018052)
黑龙江省中医药科研项目(No.ZHY19-004)。
