OBJECTIVES: To explore the mechanism of Shenqi Buzhong (SQBZ) Formula for alleviating mitochondrial dysfunction in a rat model of chronic obstructive pulmonary disease (COPD) in light of the AMPK/SIRT1/PGC-1α pathway. METHODS: Fifty male SD rat models of COPD, established by intratracheal lipopolysaccharide (LPS) instillation, exposure to cigarette smoke, and gavage of Senna leaf infusion, were randomized into 5 groups (n=10) for treatment with saline (model group), SQBZ Formula at low, moderate and high doses (3.08, 6.16 and 12.32 g/kg, respectively), or aminophylline (0.024 g/kg) by gavage for 4 weeks, with another 10 untreated rats as the control group. Pulmonary function of the rats were tested, and pathologies and ultrastructural changes of the lung tissues were examined using HE staining and transmission electron microscopy. The levels of SOD, ATP, MDA, and mitochondrial membrane potential in the lungs were detected using WST-1, colorimetric assay, TBA, and JC-1 methods. Flow cytometry was used to analyze ROS level in the lung tissues, and the protein expression levels of P-AMPKα, AMPKα, SIRTI, and PGC-1α were detected using Western blotting. RESULTS: The rat models of COPD showed significantly decreased lung function, severe histopathological injuries of the lungs, decreased pulmonary levels of SOD activity, ATP and mitochondrial membrane potential, increased levels of MDA and ROS, and decreased pulmonary expressions of P-AMPKα, SIRTI, and PGC-1α proteins. All these changes were significantly alleviated by treatment with SQBZ Formula and aminophylline, and the efficacy was comparable between high-dose SQBZ Formula group and aminophylline group. CONCLUSIONS SQBZ Formula ameliorates mitochondrial dysfunction in COPD rats possibly by activating the AMPK/SIRT1/PGC-1α pathway.
Animals ; Pulmonary Disease, Chronic Obstructive/drug therapy* ; Drugs, Chinese Herbal/therapeutic use* ; Sirtuin 1/metabolism* ; Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha ; Rats, Sprague-Dawley ; Male ; Rats ; AMP-Activated Protein Kinases/metabolism* ; Mitochondria/metabolism* ; Disease Models, Animal ; Signal Transduction/drug effects*

Objective To investigate the effects of Shenqi Tiaoshen Formula(SQTSF)for alleviating airway inflammation in rats with both chronic obstructive pulmonary disease(COPD)and lung-kidney qi deficiency syndrome and explore its therapeutic mechanism.Methods Forty-eight SD rats were randomly divided into control group,model group,low-,medium-,and high-dose SQTSF groups,and aminophylline(APL)group.In all but the control group,rat models of COPD with lung-kidney qi deficiency syndrome were established and treated with saline,SQTSF or APL via daily gavage as indicated(starting from day 30).The rats were observed for changes in body weight,grip strength,lung function,lung pathology,inflammatory cytokines in bronchoalveolar lavage fluid(BALF),oxidative stress levels,iron ion metabolism,cellular and mitochondrial ultrastructural changes in the lung tissue,and expressions of Nrf2/SLC7A11/GPX4 signaling pathway and ferroptosis-related proteins.Results The rats in the model group exhibited obvious symptoms of lung-kidney qi deficiency syndrome with significantly decreased body weight,grip strength,and lung function parameters.Examination of the lung tissue revealed showed significant inflammatory cell infiltration and emphysema with obvious bronchial,perivascular,and alveolar inflammation and alveolar destruction,significantly increased IL-1β,TNF-α,IL-6,and IL-13 levels in BALF,and elevated pulmonary oxidative stress levels and Fe2+and total iron ion concentrations.The rat models also showed characteristic ultrastructural changes of ferroptosis in the lung tissue cells under transmission electron microscope and significantly decreased Nrf2,GPX4,and SLC7A11 and increased ACSL4 expressions in the lung tissue.Treatment with SQTSF significantly improved these pathological changes in the rat models with a better effect than APL.Conclusion SQTSF can effectively improve airway inflammation and oxidative stress in COPD rats with lung-kidney qi deficiency possibly by inhibiting ferroptosis via regulating the Nrf2/SLC7A11/GPX4 signaling pathway.

Objective To explore the therapeutic mechanism of Liuwei Buqi(LWBQ)Formula for chronic obstructive pulmonary disease(COPD)in rat models.Methods SD rat models of COPD established by cigarette smoking combined with intratracheal lipopolysaccharide(LPS)instillation and hormone injection were treated with LWBQ Formula by gavage with or without intraperitoneal injection of MCC950 for 3 weeks,starting at the 5th week of modeling.After the treatments,the rats were examined for lung pathologies,lung function,total cell count and white blood cell count in bronchoalveolar lavage fluid(BALF),and serum levels of IL-6,TNF-α,IL-18 and NO.The mRNA expressions of NLRP3,ASC,caspase-1,GSDMD-N,IL-1β,and IL-18 in the lung tissue were detected with qRT-PCR.Results Compared with the normal control rats,the COPD rat models had severe lung pathologies and showed significantly decreased lung function,increased total cell and leukocyte subset counts in BALF,and increased serum levels of IL-6,TNF-α,IL-18 and NO and mRNA expressions of pyroptosis-related proteins in the lung tissue.Treatment of the rat models with LWBQ Formula significantly improved lung pathology and lung function,reduced total cell and leukocyte counts in BALF,and decreased serum levels of the inflammatory factors and expressions of pyroptosis-related proteins in the lung tissue.The combined treatment with MCC950 further improved lung pathology and function in spite of a significant difference,but BALF cell counts,serum inflammatory factor levels and pulmonary expressions of pyroptosis-related proteins were all significantly reduced following the treatment.Conclusion LWBQ Formula can delay the progression of COPD in rats possibly by inhibiting lung tissue pyroptosis via regulating the NLRP3/caspase-1/GSDMD pathway to reduce inflammatory response and lung damage.

Objective To investigate the effects of Shenqi Tiaoshen Formula(SQTSF)for alleviating airway inflammation in rats with both chronic obstructive pulmonary disease(COPD)and lung-kidney qi deficiency syndrome and explore its therapeutic mechanism.Methods Forty-eight SD rats were randomly divided into control group,model group,low-,medium-,and high-dose SQTSF groups,and aminophylline(APL)group.In all but the control group,rat models of COPD with lung-kidney qi deficiency syndrome were established and treated with saline,SQTSF or APL via daily gavage as indicated(starting from day 30).The rats were observed for changes in body weight,grip strength,lung function,lung pathology,inflammatory cytokines in bronchoalveolar lavage fluid(BALF),oxidative stress levels,iron ion metabolism,cellular and mitochondrial ultrastructural changes in the lung tissue,and expressions of Nrf2/SLC7A11/GPX4 signaling pathway and ferroptosis-related proteins.Results The rats in the model group exhibited obvious symptoms of lung-kidney qi deficiency syndrome with significantly decreased body weight,grip strength,and lung function parameters.Examination of the lung tissue revealed showed significant inflammatory cell infiltration and emphysema with obvious bronchial,perivascular,and alveolar inflammation and alveolar destruction,significantly increased IL-1β,TNF-α,IL-6,and IL-13 levels in BALF,and elevated pulmonary oxidative stress levels and Fe2+and total iron ion concentrations.The rat models also showed characteristic ultrastructural changes of ferroptosis in the lung tissue cells under transmission electron microscope and significantly decreased Nrf2,GPX4,and SLC7A11 and increased ACSL4 expressions in the lung tissue.Treatment with SQTSF significantly improved these pathological changes in the rat models with a better effect than APL.Conclusion SQTSF can effectively improve airway inflammation and oxidative stress in COPD rats with lung-kidney qi deficiency possibly by inhibiting ferroptosis via regulating the Nrf2/SLC7A11/GPX4 signaling pathway.

Objective To explore the therapeutic mechanism of Liuwei Buqi(LWBQ)Formula for chronic obstructive pulmonary disease(COPD)in rat models.Methods SD rat models of COPD established by cigarette smoking combined with intratracheal lipopolysaccharide(LPS)instillation and hormone injection were treated with LWBQ Formula by gavage with or without intraperitoneal injection of MCC950 for 3 weeks,starting at the 5th week of modeling.After the treatments,the rats were examined for lung pathologies,lung function,total cell count and white blood cell count in bronchoalveolar lavage fluid(BALF),and serum levels of IL-6,TNF-α,IL-18 and NO.The mRNA expressions of NLRP3,ASC,caspase-1,GSDMD-N,IL-1β,and IL-18 in the lung tissue were detected with qRT-PCR.Results Compared with the normal control rats,the COPD rat models had severe lung pathologies and showed significantly decreased lung function,increased total cell and leukocyte subset counts in BALF,and increased serum levels of IL-6,TNF-α,IL-18 and NO and mRNA expressions of pyroptosis-related proteins in the lung tissue.Treatment of the rat models with LWBQ Formula significantly improved lung pathology and lung function,reduced total cell and leukocyte counts in BALF,and decreased serum levels of the inflammatory factors and expressions of pyroptosis-related proteins in the lung tissue.The combined treatment with MCC950 further improved lung pathology and function in spite of a significant difference,but BALF cell counts,serum inflammatory factor levels and pulmonary expressions of pyroptosis-related proteins were all significantly reduced following the treatment.Conclusion LWBQ Formula can delay the progression of COPD in rats possibly by inhibiting lung tissue pyroptosis via regulating the NLRP3/caspase-1/GSDMD pathway to reduce inflammatory response and lung damage.

ObjectiveTo investigate the effect of Mashao Pingchuan decoction （MSPC） on lipopolysaccharides （LPS）-induced autophagy in human bronchial airway epithelial cells （16HBE） via the phosphatidylinositol 3-kinase （PI3K）/protein kinase B （Akt）/mammalian target of rapamycin （mTOR） signaling pathway. Method16HBE cells were selected for the study， and cell counting kit-8 （CCK-8） was used to detect the activity of of LPS-induced 16HBE cells and the effect of MSPC-containing serum on the cells. Suitable LPS-induced 16HBE cells were screened by the CCK-8 method， and the content of tumor necrosis factor-α （TNF-α） was measured to identify the established model. And MSPC-containing serum was prepared. The cells were divided into normal group， LPS group， LPS+MSPC group， LY294002+LPS group and LY294002+LPS+MSPC group. Transmission electron microscopy was performed to observe the changes in autophagic vesicles and ultrastructure of the cells. Western blot was performed to detect the protein expressions of PI3K， phosphorylated PI3K （p-PI3K）， Akt， phosphorylated Akt （p-Akt）， mTOR， phosphorylated mTOR （p-mTOR） and microtubule-associated protein 1 light chain 3B （LC3B）， and enzyme-linked immunosorbent assay （ELISA） was used to detect the expressions of inflammatory factors interleukin-5 （IL-5）， IL-6， TNF-α and IL-10 in the five groups. ResultLPS inhibited the 16HBE cells in a dose-dependent manner. Compared with the normal group， the LPS group （150 mg·L^-1 of LPS） increased the expression of pro-inflammatory factor TNF-α after 24 h of treatment （P<0.05） and facilitated the autophagosome formation， and MSPC-containing serum exerted a concentration-dependent promotion effect on the 16HBE cells， inhibited the autophagy to a certain degree and enhanced the cell status. Western blot revealed that the protein expressions of p-PI3K， p-Akt and p-mTOR in the model group were lower （P<0.05） and the protein expression of LC3B was higher （P<0.01） than those in the normal group. Compared with the conditions in the LPS group， the protein expressions of p-PI3K， p-Akt and p-mTOR in the LPS+MSPC group were elevated （P<0.05） and that of LC3B was reduced （P<0.05）. Compared with the LPS+LY294002 group， the LY294002+LPS+MSCP group had up-regulated protein expressions of p-PI3K， p-Akt and p-mTOR （P<0.05） and down-regulated protein expression of LC3B （P<0.05）. ELISA showed that the LPS group had higher levels of IL-5， IL-6， TNF-α and IL-10 than the normal group， while the levels of TNF-α， IL-6 and IL-8 were decreased （P<0.01） and the level of IL-10 was increased （P<0.01） after treatment with MSCP. ConclusionMSCP may lower the LPS-induced autophagy in 16HBE cells and improve the inflammatory response through activating the PI3K/Akt/mTOR signaling pathway.

ObjectiveTo establish and evaluate a chronic obstructive pulmonary disease （COPD） model with lung-spleen qi deficiency. MethodA rat model mimicking COPD with lung-spleen qi deficiency was established by the combination of cigarette smoking and intratracheal instillation of lipopolysaccharide （LPS） along with gavage of Sennae Folium infusion. Forty male SPF-grade SD rats were randomly assigned to blank， model， and low- （L-FXY）， medium- （M-FXY）， and high-dose （H-FXY） Sennae Folium infusion groups. Other groups except the blank group were exposed to daily cigarette smoke， with LPS administrated via intratracheal instillation on the 1st and 14th days. On the 28th day of modeling， the L-FXY， M-FXY， and H-FXY groups were administrated with Sennae Folium infusion at 5， 10， and 20 g·kg^-1， respectively， and at 4 ℃ for three weeks. The modeling lasted for 49 days. The general conditions （body mass， food intake， fecal water content， and anal temperature） and behaviors （grip strength test and tail suspension test） of rats in different groups were examined. The lung function， lung histopathology， D-xylose， amylase， and gastrin levels in the serum， interleukin（IL）-1β and IL-6 levels in the alveolar lavage fluid， levels of T-lymphocyte subsets （CD4⁺， CD8⁺， and CD4⁺/CD8⁺） in the peripheral blood， and thymus and spleen indices were measured. ResultTwo rats died in the H-FXY group. Compared with the blank group， both the M-FXY and H-FXY groups exhibited reduced body mass and food intake （P<0.01） and increased fecal water content （P<0.01）. The anal temperature in the H-FXY group was lower than that in the blank group （P<0.01）. The grip strength decreased in the modeling groups compared with the blank group （P<0.01）， and the duration of immobility in the tail suspension test increased in the M-FXY and H-FXY groups （P<0.05， P<0.01）. Compared with the blank group， the modeling groups showed reduced 0.3 second forced expiratory volume （FEV_0.3）， FEV_0.3/forced vital capacity （FVC）（P<0.01）， thickening of bronchial walls， proliferation of goblet cells， and the presence of emphysematous changes. In terms of gastrointestinal function， the M-FXY and H-FXY groups had lower levels of D-xylose， gastrin， and α-amylase than the blank group （P<0.01）. Regarding the immune and inflammatory indices， the M-FXY and H-FXY groups showed lower thymus and spleen indices than the blank group （P<0.01）. Compared with the blank group， the modeling groups presented lowered CD4⁺ level （P<0.01） and CD4⁺/CD8⁺ ratio （P<0.05， P<0.01） in the peripheral blood and elevated levels of IL-1β and IL-6 in the alveolar lavage fluid （P<0.01） than the blank group. ConclusionA model of COPD with lung-spleen Qi deficiency was established through the combination of daily cigarette smoke， intratracheal instillation with LPS， and gavage of Sennae Folium infusion. The comprehensive evaluation results suggested medium-dose （10 g·kg^-1） Sennae Folium infusion for gavage during the modeling of COPD with lung-spleen Qi deficiency.

Qi-Yu-San-Long decoction(QYSLD)is a traditional Chinese medicine that has been clinically used in the treatment of non-small-cell lung cancer(NSCLC)for more than 20 years.However,to date,metabolic-related studies on QYSLD have not been performed.In this study,a post-targeted screening strategy based on ultra-performance liquid chromatography coupled with quadrupole time-of-flight full infor-mation tandem mass spectrometry(UPLC-QTOF-MSE)was developed to identify QYSLD-related xeno-biotics in rat urine.The chemical compound database of QYSLD constituents was established from previous research,and metabolites related to these compounds were predicted in combination with their possible metabolic pathways.The metabolites were identified by extracted ion chromatograms using predicted m/z values as well as retention time,excimer ions,and fragmentation behavior.Overall,85 QYSLD-related xenobiotics(20 prototype compounds and 65 metabolites)were characterized from rat urine.The main metabolic reactions and elimination features of QYSLD included oxidation,reduction,decarboxylation,hydrolysis,demethylation,glucuronidation,sulfation,methylation,deglycosylation,acetylation,and associated combination reactions.Of the identified molecules,14 prototype compounds and 58 metabolites were slowly eliminated,thus accumulating in vivo over an extended period,while five prototypes and two metabolites were present in vivo for a short duration.Furthermore,one pro-totype and five metabolites underwent the process of"appearing-disappearing-reappearing"in vivo.Overall,the metabolic profile and characteristics of QYSLD in rat urine were determined,which is useful in elucidating the active components of the decoction in vivo,thus providing the basis for studying its mechanism of action.

Objective : To investigate the effect of Qiyu Sanlong Decoction ( QYSLD) on inhibiting lung cancer by regulating tumor immune microenvironment through PD-1 signaling pathway and on Th1 immune response in lung cancer bearing mice． Methods : Lewis lung cancer cell line ( LLC) was used to model subcutaneously implanted tumor in mice．72 mice were randomly divided into Control group ( n = 18 ) ，cisplatin group ( n = 18 ) ，QYSLD group (n = 18) and combined group (n = 18) ．The general survival of mice was observedand recorded ; the specific Th1 cell response to tumor antigen was detected by ELISA method ; the proportion of PD-1 positive cells and the ap- optosis of memory CD4 + T cells in spleen T cells of mice were detected by flow cytometry ; and the number and vol- ume of pulmonary metastatic nodules were counted under an anatomical microscope after 21 days of continuous treatment. Results : QYSLD could improve the general survival of mice bearing lung cancer，up-regulate the im- mune response of CD4 + T cells and the specific killing effect of CD8 + T cells．The proportion of apoptotic cells in PD-1 positive cells and CD4 + memory T cells in spleen T cells of mice was down-regulated，and lung metastasis of mouse tumor cells was inhibited． Conclusion QYSLD can improve the survival status of mice bearing lung cancer and enhance Th1 immune response by mediating PD-1 signaling pathway，improve immune function and inhibit im- mune escape of tumor cells，thus inhibiting the metastasis of lung cancer.

Objective To observe the clinical effect of traditional Chinese medicine (TCM) characteristic lung rehabilitation in treatment of patients with chronic obstructive pulmonary disease (COPD) and TCM syndrome of lung and kidney qi deficiency at stable period. Methods Sixty patients with stable COPD and lung and kidney qi deficiency syndrome admitted to the First Affiliated Hospital of Anhui University of Chinese Medicine from June to August 2017 were enrolled, and they were divided into routine treatment group and lung rehabilitation treatment group according to the random number table method, each group 30 cases. The routine treatment group was given Seretide (serevent/futicasone) dry powderi nhalation therapy; on the basis of therapy in the routine treatment group, the lung rehabilitation treatment group was treated with TCM characteristic lung rehabilitation technology (acupoint application + Chinese medicine ionic induction + oral administration of Chinese medicine Liuweibuqi granules, delivery at appropriate intervals); both groups were treated for 2 months. The changes of TCM syndrome score, western medicine symptom score, the times of acute exacerbation of COPD, COPD assessment test (CAT) score, lung function indexes: forced expiratory volume in one second (FEV1), FEV1/forced vital capacity (FVC) were observed before and after treatment in two groups. Results After treatment, TCM syndrome score, western medicine symptom score, CAT score, and after treatment the times of acute exacerbation of COPD in both groups were significantly lower than those before treatment, and the above indexes in the lung rehabilitation treatment group were markedly lower than those in routine treatment group [TCM syndrome score:11.93±1.80 vs. 14.27±2.88, western medicine symptom score: 14.20±2.75 vs. 11.93±4.23, CAT score: 14.87±2.60 vs. 16.23±4.39, the times of acute exacerbation of COPD (times): 0.63±0.49 vs. 0.95±0.83, all P < 0.05]. The improvement of FEV1 in the two groups was not significant; but FEV1/FVC in lung rehabilitation treatment group was obviously higher than that before treatment, FEV1/FVC in lung rehabilitation treatment group was significantly higher than that in the routine treatment group [(57.93±7.27)% vs. (52.49±6.61)%, P < 0.05]. Conclusion The application of TCM characteristic lung rehabilitation in the treatment of COPD patients with stable lung and kidney qi deficiency syndrome based on bronchodilators and glucocorticoids can reduce the number of acute exacerbation, improve the patients' clinical symptoms and living quality, but the improvement of lung function is not significant.