宣肺化瘀方经激活泛素-蛋白酶体途径抗肺纤维化的相关机制研究  

Mechanisms of Xuanfei Huayu Decoction on pulmonary fibrosis via activating ubiquitin-proteasome pathway

作　　者：覃惠 胡静[1] 温昊天 顾凯娟 石克华[4] 熊必丹[4] QIN Hui;HU Jing;WEN Haotian;GU Kaijuan;SHI Kehua;XIONG Bidan(School of Basic Medical Science,Shanghai University of Traditional Chinese Medicine,Shanghai 201203,China;Seventh People＇s Hospital of Shanghai University of Traditional Chinese Medicine,Shanghai 200137,China;Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine,Shanghai 201203,China;Shanghai Municipal Hospital of Traditional Chinese Medicine,Shanghai University of Traditional Chinese Medicine,Shanghai 200071,China)

机构地区：[1]上海中医药大学基础医学院,上海201203 [2]上海中医药大学附属第七人民医院,上海200137 [3]上海中医药大学附属曙光医院,上海201203 [4]上海中医药大学附属市中医医院,上海200071

出　　处：《上海中医药大学学报》2018年第4期75-81,共7页Academic Journal of Shanghai University of Traditional Chinese Medicine

基　　金：国家自然科学基金资助项目(81001491);上海市自然科学基金资助项目(15ZR1441100);上海中医药大学经方理论应用研究中心项目(A1-Z183020110);上海中医药大学院院合作项目;上海市"中医学"一流学科科研创新基金项目

摘　　要：目的:基于泛素-蛋白酶体通路探讨宣肺化瘀方(XFHY)防治大鼠肺纤维化的作用机制。方法:将60只雄性SPF级Wistar大鼠随机分为6组,即阴性对照组、模型组、醋酸泼尼松组、XFHY高剂量组、XFHY中剂量组、XFHY低剂量组。除阴性对照组外,均经鼻滴入博莱霉素建立大鼠肺纤维化模型。XFHY高、中、低剂量组大鼠分别每日灌胃XFHY14.38 g/kg、7.19 g/kg、3.60 g/kg,醋酸泼尼松组大鼠每日剂量为0.167 mg/kg,模型组和阴性对照组给予等体积的0.9%Na Cl溶液灌胃。各组均灌胃28 d。处死大鼠,通过肺组织HE染色和天狼猩红染色鉴定模型。采用碱性水解法测定肺组织中羟脯氨酸(HYP)含量;采用Power Lab测定肺功能;应用ELISA法检测支气管肺泡灌洗液(BALF)和肺组织中TGF-β1含量;应用Western blot检测肺组织Ub、PSMC2、Smurf1、Smurf2蛋白表达。结果:HE染色和天狼猩红染色结果显示,肺纤维化模型组肺组织可见大量胶原纤维沉积,宣肺化瘀方各剂量组肺泡间隔胶原纤维表达明显减少。与阴性对照组比较,模型组HYP含量及BALF和肺组织中TGF-β1含量显著升高,而平均呼气峰流量(PEF)和每分钟通气量(MV)显著降低,Ub、PSMC2、Smurf1、Smurf2蛋白表达明显降低(P<0.01,P<0.05);与模型组比较,XFHY各剂量组胶原纤维表达和HYP含量均显著下降,其中XFHY高剂量组比XFHY低剂量组和中剂量组胶原纤维表达更少(P<0.05),HYP含量更低(P<0.01,P<0.05);而XFHY各剂量组PEF和MV均有所升高,其中,XFHY高剂量组比XFHY低剂量组和中剂量组更高(P<0.01,P<0.05);XFHY高剂量组BALF和肺组织中TGF-β1含量显著降低,而Ub、PSMC2、Smurf1、Smurf2蛋白表达明显升高(P<0.01)。结论:宣肺化瘀方能有效防治肺纤维化,其机制可能是通过激活泛素-蛋白酶体通路、降低BALF和肺组织中TGF-β1的蛋白含量,从而减少胶原纤维形成。Objective： To explore the mechanisms of Xuanfei Huayu Decoction（ XFHY） in the prevention and treatment of pulmonary fibrosis in rats based on the ubiquitin-proteasome pathway. Methods： 60 male Wistar rats of SPF grade were randomly divided into 6 groups,including negative control group,model group,prednisone acetate group,XFHY groups with low-,medium-and high-dose. Except the negative control group,the pulmonary fibrosis model was established in other rats by bleomycin with nasal instillation. The XFHY-treated groups were treated with Xuanfei Huayu Decoction at daily dose of 3. 60 g/kg,7. 19 g/kg and 14. 38 g/kg by intragastric administration,respectively. The prednisone acetate group was treated with prednisone acetate at daily dose of 0.167 mg/kg. The model group and negative control group were treated with 0.9% Na Cl solution at equal volume. All rats were sacrificed after intragastric administration for 28 days. The lung tissue was taken and HE staining and sirius red staining were performed to identify the model. The content of hydroxyproline（ HYP） in lung tissue was detected by alkali hydrolysis. The pulmonary function indexes were detected by Power Lab. The level of TGF-β1 in the bronchoalveolar lavage fluid（ BALF） and lung tissue were determined by ELISA. The proteins expressions of Ub,PSMC2,Smurf1 and Smurf2 in lung tissue were tested by Western blot. Results： The results of HE staining and sirius red staining showed that a large number of collagen fibrous depositions in the lung tissue were observed in the model group. The expression of collagen fibers in the alveolar septum was significantly reduced in XFHY-treated groups. Compared with the negative control group,the HYP content and the level of TGF-β1 in the BALF and lung tissue were significantly increased in the model group,while the mean peak expiratory flow（ PEF） and ventilation per minute（ MV） were decreased significantly,and the protein expressions of Ub,PSMC2,Smurf1 and Smurf2 were obviously decreased（ P〈0.05,P�

关 键 词：肺纤维化 泛素-蛋白酶体通路 宣肺化瘀方 Smad泛素调节因子 

分 类 号：R285.5[医药卫生—中药学]