| Chronic Obstructive Pulmonary Disease(COPD)is characterized by incomepletely reversible airflow limitation.It is often progressively worsened.COPD is related to the harmful particles,toxic gas and abnormal inflammatory response.COPD is mainly caused by genetic and external factors such as smoking,environmental pollution,occupational dust and infection,etc.Smoking is recognized as one of the most important risk factors.COPD is a global health problem seriously affecting the quality of patient’s life.With the growth of the global population,the economic burden caused by COPD will continue to increase.So far,effective drugs for preventing and treating COPD have not yet been developed.The purpose of this study was to investigate the pharmacodynamic effects and explore the action mechanism of Qizhi Yifei Granules on the COPD model rats.1 The preliminary pharmacodynamic comparative study of Qizhi Yifei granules in different pharmaceutical processes on COPD model ratsObjective:To compare the preliminary pharmacodynamics of Qizhi Yifei granules with different pharmaceutical processes and to screen out the preparation techniques with better pharmacodynamics effects.Method:A rat model of COPD was established by infusion of LPS with cigarette smoke exposure.10 rats were put into a self-made box with the atmosphere of 8 ignited cigarettes and then kept for 30 minutes every morning for 28 days.The LPS powder was prepared into a solution of 1 mg/mL with normal saline solution and then injected into the airway of rats in a dose of 200 μL on the 1st,11st and 21st days,respectively.On these injection days,smoking was suspended.On the 21st day of experiment,7 male rats were randomly taken from vehicle control group rats and model rats for pre-dose model identification,respectively.The rest of rats were randomly divided into 7 groups:vehicle control group,model control group,roxithromycin group,two groups of Qizhi Yifei granules process 1 with high(9.10 g/kg)and small(4.55 g/kg)dosage,and two groups of Qizhi Yifei Granules process 2 with high(9.10 g/kg)and small(4.55 g/kg)dosage.18 rats per group,half male and half female.On the 21st day,the model control group and the vehicle control group were fed once daily with 1 mL/100 g of normal water,while other groups were given the corresponding drugs once a day at 1mL/100g for 40 days.On the 61st day,all rats were sacrificed in order to determine 0.3%forced expiratory volume(FEV0.3),FEV0.3/FVC%,forced expiratory flow rate(FEV25-75%),maximum forced expiratory flow rate(PEF),maximum ventilation(MVV)and forced vital capacity(RL),total exhalation resistance(Re),dynamic compliance(Cdyn),lung index and psychrometric ratio.The lungs of rats in each group were subjected to HE and AB-PAS staining.Results:Compared with the model control group,FEV0.3/FVC%and MVV were significantly increased and RL was significantly decreased in the Qizhi Yifei Granules process 1 groups with high and small dose(P<0.01).Compared with the model control group,FEV0.3/FVC%,MVV was significantly increased in the Qizhi Yifei Granules process 2 groups with high and small dose(P<0.01 or 0.05),Cdyn was significantly increased in the Qizhi Yifei Granules process 2 groups with small dose(P<0.01).RL and Re were significantly decreased in the Qizhi Yifei Granules process 2 groups with small dose.The results of HE staining showed that compared with the model control group,the pathological changes of lung tissue in both two processes were significantly improved.The results of AB-PAS staining showed that the number of goblet cells was significantly decreased in the Qizhi Yifei Granules of two processes.According to the results of Cdyn,Re,and lung index,the process 2 with small dosage was more effective than process 1 with small dosage.Compared with the process 1 with high dosage,the number of goblet cells in the airway cilia was more significantly reduced in the process 2 with high dosage.Conclusion:Process 2(water extraction and alcohol precipitation)showed a better result for Qizhi Yifei granules than process 1(full water extraction).Therefore,pharmacodynamics experiments will be further conducted with process 2 in next section.2 The rapeutic effects and preliminary mechanism of Qizhi Yifei granules on COPD rats2.1 Effects of Qizhi Yifei granules on pulmonary function and related inflammatory factors in COPD ratsObjective:To investigate the effects and mechanism of Qizhi Yifei Granules on COPD rats.Methods:Wistar rats were randomly divided into 6 groups:vehicle control group,model control group,roxithromycin group,and three dosage groups of Qizhi Yifei Granules with high(18.20 g/kg),medium 9.10 g/kg)and low(4.55 g/kg),18 rats per group,half male and half female.A rat model of COPD was established by infusion of LPS with cigarette smoke exposure.At the end of the 61 days,all rats were sacrificed in order to determine 0.3%forced expiratory volume(FEV0.3),FEV0.3/FVC%,maximum ventilation(MVV)and forced vital capacity(RL).The expression levels of IL-8 and IL-13 in rat serum and lung tissue homogenate and Serum hydroxyproline(HYP)levels in rat serum were measured.The lungs of rats in each group were subjected to HE and Masson staining.Results:Compared with the vehicle control group,FEV0.3、FEV0.3/FVC%、FEF25-75%、MVV、PEF and Cdyn were significantly decreased(P<0.01 or 0.05)、RL significantly increased in the model control group(P<0.05),IL-8 and IL-13 expression levels in the serum and the lung homogenate were significantly increased(P<0.05 or 0.01).The expression level of HYP in the serum was significantly increased in the model control group(P<0.05).The lung tissue collagen fiber area divided by the total area of lungs was significantly increased in the model control group(P<0.05),FVC,FEV0.3,FEV0.3/FVC%,MVV,FEF25-75%and PEF were significantly increased(P<0.01),and the RL was significantly decreased in the Qizhi Yifei Granules groups with high,middle and low dose(P<0.01 or 0.05).The Cdyn were significantly increased and the Re were significantly decreased in the Qizhi Yifei Granules groups with low dose(P<0.01).The results of Elisa assay showed that the serum levels of IL-8 in high and middle dose of Qizhi Yifei Granules group were significantly decreased(P<0.01 or 0.05),Serum levels of IL-13 in middle and low dose groups were significantly decreased(P<0.01 or 0.05).The contents of IL-8 and IL-13 in the lung homogenate of the low dose group were significantly decreased(P<0.05).Biochemical tests showed that,the serum levels of HYP in the high and low dose groups were significantly decreased(P<0.0 or 0.05).The results of HE staining showed that compared with the model control group,the pathological changes of lung tissue in high,middle and low dose groups of Qizhi Yifei Granules were significantly improved.The results of Masson staining showed that lung tissue collagen fiber area divided by the total area of lungs in high,medium and low doses of Qizhi Yifei Granules decreased significantly(P<0.05).Conclusion:Qizhi Yifei Granules could reduce the infiltration of inflammatory cells in the lung tissue,improve the lung function of COPD model rats and reduce the deposition of collagen fibers in the lung tissue,ameliorate airway remodeling in lung tissue of COPD model rats by regulating the expression levels of inflammatory factors such as IL-8 and IL-13,and thereby protected the lung tissue of model rats finally.2.2 Effects of Qizhi Yifei granules on expression level of airway remodeling associated proteins in lung tissue of COPD ratsObjective:The experiment was proceeded on a rat model of COPD based on infusion of LPS with cigarette smoke exposure.Several parameters of lung tissue of rats were determined,including metolloproteinase-9(MMP-9),metolloproteinase-12(MMP-12),anti-metolloproteinase-9(TIMP-1),epidemal growth factor respector(EGFR),transforming growth factor(TGF-β1),and expression levels of type III and type I collagen in airway.The protection effects of Qizhi Yifei granules on lung tissue of COPD rats were observed.To investigate the effects of Qizhi Yifei granules on the lung tissues cathepsin-antiprotease balance in lung tissues of COPD rats.To clarify the inhibitory effects of Qizhi Yifei granules on airway remodeling in COPD rats lung tissues.To provide experimental basis for clinical treatment and development of new drugs in this research.Method:Wistar rats were randomly divided into 6 groups:vehicle control group,model control group,roxithromycin group,and three dosage groups of Qizhi Yifei granules with high(18.20 g/kg),medium(9.10 g/kg)and low(4.55 g/kg).18 rats per group,half male and half female.A rat model of COPD was established by infusion of LPS with cigarette smoke exposure.At the end of the 61 days,all rats were sacrificed to determine the expression levels of MMP-9,MMP-12,TIMP-1,EGFR,TGF-β1,and type Ⅰ collagen in lung tissues with immunohistochemistry.Results:Compared with the vehicle control group,the expression levels of MMP-9,MMP-12 and TIMP of the lung tissue and the airway type Ⅲ collagen in the model control group were significantly increased(P<0.01).Compared with the model control group,the expression levels of MMP-9,MMP-12 and TIMP were significantly decreased(P<0.01 or 0.05)in the roxithromycin group,and three dosage groups of Qizhi Yifei granules with high,medium and low dose,the expression level of EGFR were significantly decreased(P<0.01 or 0.05)in the roxithromycin group,and the low dose,the expression level of TGF-β1 were significantly decreased(P<0.01 or 0.05)in the roxithromycin group,and two dosage groups of Qizhi Yifei granules with high and medium dose.the expression level of airway type Ⅲcollagen were significantly decreased(P<0.05)in the roxithromycin group,and two dosage groups of Qizhi Yifei Granules with high and low dose.the expression level of airway type Ⅰcollagen was significantly decreased(P<0.05)in the Qizhi Yifei granules low dose.Conclusion:Qizhi Yifei granules could modulate the balance of metalloproteinase-antimetalloprotease system by regulating the expressions of EGFR,TGF-β1 and other factors,which could help to inhibit airway remodeling and improve airway blockage.2.3 The effects of Qizhi Yifei granules on mucus hypersecretion in model rats with COPDObjective:In this study,a rat model of COPD was established by infusion of LPS with cigarette smoke exposure.By measuring the lung tissue expressions of NE,MUC5B and conducting AB-PAS staining in the rats’ lung,the specific role of neutrophils and other secretory NE in the lung of COPD rat model was observed.the effects of Qizhi Yifei granules on goblet cell metaplasia,mucus secretion in the airway was investigated.Their benefits on the improvement of airway obstruction was futher clarified.This research was provided experimental basis for clinical treatment and development of new drugs.Method:Wistar rats were randomly divided into 6 groups:vehicle control group,model control group,roxithromycin group,and three dosage groups of Qizhi Yifei Granules with high(18.20 g/kg),medium(9.10 g/kg)and low(4.55 g/kg).18 rats per group,half male and half female.A rat model of COPD was established by infusion of LPS with cigarette smoke exposure.At the end of the 61 days,all rats were sacrificed to determine the expressions of NE and MUC5B in lung tissues with immunohistochemistry.The lungs of rats in each group were subjected to AB-PAS staining.Result:Compared with the vihicle control group,the expression levels of NE and MUC5B of the lung tissue were significantly incresead in the model control group(P<0.05),the number of goblet cells on the tracheal ciliary of lung tissue in the model control group were significantly decreased(P<0.01).Compared with the model control group,the expression levels of NE and the number of goblet cells on the tracheal ciliary of lung tissue were significantly decreased(P<0.01 or 0.05)in the roxithromycin group,and three dosage groups of Qizhi Yifei granules with high,medium and low dose.The expression level of MUC5B were significantly decreased(P<0.05).Conclusion:Qizhi Yifei granules could regulate the synthesis and secretion of goblet cells in the lung of COPD model rats to adjust the metaplasia and expression of goblet cells and reduce the secretion of mucus,which had a certain effect on airway blockage. |
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