摘要
目的:研究参芪调肾方对香烟烟雾(CS)暴露法建立的慢性阻塞性肺疾病(COPD)模型小鼠的炎症抑制、肺组织病理及骨组织内破骨细胞介导的骨吸收的改善作用机制。方法:将40只小鼠随机分为正常对照组、模型对照组、参芪调肾方8.53、17.06、34.11 g/kg组,每组8只,除正常对照组外,其余4组采用CS暴露法造模24 w,第12 w-24 w各组灌胃给予相应药物或生理盐水,连续12 w;ELISA法检测肺组织、外周血、骨组织内炎症因子白细胞介素(IL)-1、IL-6、肿瘤坏死因子-α(TNF-α)含量;HE染色观察肺组织病理学改变;抗酒石酸酸性磷酸酶(TRAP)染色观察骨组织内破骨细胞;免疫组化法观察骨组织内骨保护素(OPG)、核因子κB受体活化因子配体(RANKL)、核因子κB p65(NF-κB p65)、基质金属蛋白酶-9(MMP-9)蛋白在小鼠胫骨组织中的定位表达;Western blot法检测肺组织NF-κB p65、p-NF-κB p65、MMP-9及胫骨OPG、RANKL、NF-κB p65、p-NF-κB p65、MMP-9、组织蛋白酶K(CTSK)蛋白表达。结果:与正常对照组比较,模型对照组肺组织损伤明显,管腔内、管壁及周围见大量炎症细胞聚集;肺组织、外周血及骨组织内炎症因子IL-1β、IL-6、TNF-α含量显著升高(P<0.01);骨组织内破骨细胞数量、RANKL、NF-κB p65、MMP-9、CTSK、TRAP蛋白表达显著升高(P<0.01),OPG蛋白表达显著降低(P<0.01);与模型对照组比较,参芪调肾方各剂量组可不同程度改善小鼠支气管壁增厚、肺气肿形成等病理改变,减轻炎性细胞浸润,参芪调肾方34.11 g/kg组可显著下调肺组织内NF-κB p65、p-NF-κB p65、MMP-9蛋白表达(P<0.01)、降低炎症因子IL-1β、IL-6、TNF-α含量(P<0.01);明显降低骨组织内破骨细胞数量以及下调RANKL、NF-κB p65、p-NF-κB p65、MMP-9、CTSK、TRAP蛋白表达(P<0.05或P<0.01),显著升高OPG蛋白阳性表达(P<0.01)。结论:参芪调肾方对COPD模型小鼠合并骨质疏松(OP)有一定防治作用,可能是通过调控OPG/RANKL/NF-κB/MMP-9信号通路,改善COPD模型小鼠肺组织病理,降低肺、外周血、骨组织炎症反应,抑制破骨细胞过度活化。
Objective:To investigate the effects of Shenqi Tiaoshen Formula(参芪调肾方) on inflammation inhibition, lung pathology and osteoclasts-mediated bone resorption in chronic obstructive pulmonary disease(COPD) model mice established by cigarette smoke(CS) exposure. Methods:Forty mice were randomly divided into normal control group, model group, Shenqi Tiaoshen Formula 8.53 g/kg, 17.06 g/kg and 34.11 g/kg groups, with 8 mice in each group. Except for the normal control group, the mice in the other four groups were exposed to CS for 24 weeks, and the mice in each group were given corresponding drugs or normal saline by intragastric administration from the 12th to the 24th week continuously. The contents of inflammatory factors interleukin-1(IL-1),interleukin-6(IL-6) and tumor necrosis factor-α(TNF-α) in lung tissue, peripheral blood and bone tissue were detected by ELISA. HE staining was used to observe the pathological changes in lung tissue. Tartrate-resistant acid phosphatase(TRAP) staining was used to observe osteoclast in bone tissue. Immunohistochemical method was used to detect the localization and expression of osteoprotegerin(OPG),receptor activator of nuclear factor κB ligand(RANKL),nuclear factor κB p65(NF-κB p65) and matrix metalloproteinase-9(MMP-9) in tibia tissue of mice. The protein expression of NF-κB p65,p-NF-κB p65,MMP-9 in lung tissue and OPG,RANKL,NF-κB p65,p-NF-κB p65,MMP-9,cathepsin K(CTSK) in tibia were detected by Western blot. Results:Compared with the normal control group, the lung tissue of the model group was significantly damaged, and a large number of inflammatory cells were gathered in and around the lumen and tube wall. The contents of inflammatory factors IL-1β,IL-6 and TNF-α in lung tissue, peripheral blood and bone tissue were significantly increased(P<0.01). The number of osteoclasts and the protein expressions of RANKL,NF-κB p65,MMP-9,CTSK and TRAP in bone tissue were significantly increased(P<0.01),while the protein expression of OPG was significantly down regulated(P<0.01). Compared with the model group, each dose group of Shenqi Tiaoshen Formula could improve the bronchial wall thickening, emphysema formation and other pathological changes in mice to different degrees, and reduce inflammatory cell infiltration. Shenqi Tiaoshen Formula 34.11 g/kg group could significantly reduce the protein expressions of NF-κB p65,P-NF-κB p65 and MMP-9(P<0.01),and reduce the contents of inflammatory factors IL-1β,IL-6 and TNF-α(P<0.01). The number of osteoclasts and protein expressions of RANKL,NF-κB p65,p-NF-κB p65,MMP-9,CTSK and TRAP in the bone tissue of Shenqi Tiaoshen Formula 34.11 g/kg group were significantly down regulated(P<0.05 or P<0.01). OPG protein was significantly increased(P<0.01). Conclusion:Shenqi Tiaoshen Formula has certain preventive and therapeutic effects on COPD model mice complicated with osteoporosis(OP),which may be mediated by regulating OPG/RANKL/NF-κB/MMP-9 signaling pathway, improving lung pathology, reducing inflammation in lung, peripheral blood and bone tissue and inhibiting osteoclast overactivation in COPD model mice.
作者
周凡超
杨勤军
吴迪
丁焕章
王卉
马啸
吴凡
李泽庚
ZHOU Fanchao;YANG Qinjun;WU Di;DING Huanzhang;WANG Hui;MA Xiao;WU Fan;LI Zegeng(The First Affiliated Hospital of Anhui University of Chinese Medicine,Hefei 230031;Anhui University of Chinese Medicine,Hefei 230031;Respiratory Disease Institute Affiliated TCM of Anhui Academy of Chinese Medicine,Hefei 230031;Key Laboratory of Education Department of Anhui Province-Key Laboratory of TCM Prevention and Treatment of Major Lung Diseases,Hefei 230031;Key Laboratory of Xin'an Medicine,Ministry of Education,Hefei 230031)
出处
《中药药理与临床》
CAS
CSCD
北大核心
2024年第4期54-60,共7页
Pharmacology and Clinics of Chinese Materia Medica
基金
国家自然科学基金区域创新发展联合基金重点支持项目(编号:U20A20398)
国家自然科学基金面上项目(编号:82374399)
安徽省科技厅临床医学研究转化专项(编号:202204295107020045)
安徽省自然科学基金(编号:2208085QH264)
安徽省高校协同创新项目(编号:GXXT-2020-025)。
关键词
参芪调肾方
慢性阻塞性肺疾病
破骨细胞
骨保护素/核因子κB受体活化因子配体/核因子κB/基质金属蛋白酶-9
炎症
骨吸收
Shenqi Tiaoshen Formula
Chronic obstructive pulmonary disease
Osteoclast
Osteoprotegerin(OPG)/receptor activator of nuclear factor kB ligand(RANKL)/nuclear factor kB(NF-kB)/matrix metallopproteinase-9(MMP-9)
lnflanmmation
Bone resoption
