Effect Of Buqi Yifei Recipe On Airway Remodeling And TGF-?/Smads Signal Pathway In Copd Rats

1.Objective To study the effect of "Liuwei Buqi recipe" on airway remodeling and TGF-?/Smads signal pathway of COPD and its possible mechanism of action,in order to reveal the possible target,link and way of action of Buqi Yifei recipe in treating COPD model rats,and to obtain evidence for further optimizing the treatment of COPD.2.Methods 100 male SPF SD rats were selected and 15 were selected as the normal control group according to the random number table method.The other rats were used to establish COPD experimental animal model on the 1st and 14 th day of the experiment by intratracheal instillation of lipopolysaccharide(LPS)and smoke.The rats were randomly divided into five groups: model group,Liuwei Buqi recipe high dose group,Liuwei Buqi recipe middle dose group,Liuwei Buqi recipe low dose group and Guilong Kechuanning group(positive control group).The normal control group and the model group were given normal saline at a dose of 10ml/kg·d,each dose group of liuweibuqi recipe was given liuweibuqi recipe suspension,the high,middle and low dose groups were given 1.06g/kg·d,0.53g/kg·d,0.265g/kg·d,respectively(equivalent to 20,10 and 5 times of the adult clinical routine dose),and the Guilong Kechuanning group was given Guilong Kechuanning tablet suspension at a dose of 1.06g/kg·d,0.53g/kg·d,0.265g/kg·d,respectively 0.82mg/kg·d(equivalent to 10 times of the adult clinical routine dosage),each group started the drug intervention on the first day of the 9th week,twice a day,and the course of treatment was 12 weeks.The biological characteristics of rats were observed every day;the lung function was measured after the drug intervention;the lung histopathological changes were detected by HE staining;the levels of TNF-?,IL-1?,IL-8 and TGF-?1 in serum and BALF were detected by ELISA;the expression of MMP-9 and TIMP-1 in lung tissue was detected by immunohistochemistry;western The expression of TGF-?1,Smad2/3,Muc5 ac,Col-I and a-SMA in lung tissue was detected by blot,and the m RNA levels of TGF-?1,Smad2/3,Muc5 ac,Col-I and a-SMA in lung tissue were detected by RT-PCR.3.Results 3.1 Changes in biological characteristics of rats in each group In the normal control group,the rats breathed evenly and stably,without cough,cough and asthma,clean mouth and nose,no secretion,glossy hair,normal activity,sensitive response,stable weight growth,normal water intake,normal stool,ruddy tongue.In the model group,the rats were short of breath,even with obvious mouth opening,coughing,coughing and panting occasionally,secretions in the mouth and nose,tarnished fur,partial depilation,sluggish activity,sluggish crouching,slow response,perspiration,slower weight growth than other groups,less water intake,loose stools,dark tongue,ecchymosis and ecchymosis.The general characteristics and respiratory symptoms of rats in Liuwei Buqi recipe group and Guilong Kechuanning group were worse than those in the normal control group,better than those in the model group,and the growth trend of body weight since the 9th week was faster than that in the model group.There was no significant difference in general characteristics and respiratory system symptoms between each dose group of Liuwei Buqi recipe and Guilong Kechuanning group.3.2 Changes of lung function in each group Compared with the normal control group,the FVC,FEV0.3,FEV0.3/FVC(%)and PEF in the model group decreased significantly(P<0.01).Compared with the model group,the FVC,FEV0.3,FEV0.3/FVC(%)and PEF in the high dose group of Liuwei Buqi recipe and Guilong Kechuanning group were significantly higher(P<0.01);the FVC,FEV0.3/FVC(%)and PEF in the middle and low dose groups of Liuwei Buqi recipe were significantly higher(P<0.01 or P<0.05),FEV0.3 was significantly higher(P>0.05).Compared with Guilong Kechuanning group,the FVC,FEV0.3 of Liuwei Buqi recipe high dose group were significantly higher(P<0.01 or P<0.05),FEV0.3/FVC(%)and PEF were significantly higher(P>0.05);FVC,FEV0.3,FEV0.3/FVC(%)and PEF of Liuwei Buqi recipe middle dose group were significantly lower(P>0.05);FEV0.3 and PEF of Liuwei Buqi recipe low dose group were significantly lower(P>0.05)There was no significant difference in FVC,FEV0.3/FVC(%)and PEF(P>0.05).3.3 Pathological changes of lung tissue in each group In the normal control group,the ciliated columnar epithelium of bronchial mucosa at all levels was complete,the structure of respiratory tract and alveolar epithelium was complete,the cilia were arranged orderly,smooth muscle was complete,no fibrous tissue hyperplasia was found,and the morphology and structure of alveolar tissue were complete.In the model group,the ciliated columnar epithelium was incomplete,the ciliated epithelium was partly exfoliated,the goblet cells were increased,the mucinous glands were proliferated and hypertrophied;the inflammatory cells mainly composed of lymphocytes and macrophages were seen in each layer of the bronchial wall;the bronchial wall was significantly thicker than that of the normal control group,and the mucinous thrombus was seen in the bronchiole;the alveolar cavity was irregularly enlarged,and part of the alveoli were enlarged Rupture and fusion into pulmonary bullae.The above pathology of Liuwei Buqi recipe group and Guilong Kechuanning group were improved in varying degrees,especially in the high dose group.3.4 Expression of MMP-9 and TIMP-1 in lung tissue of rats in each group The number of MMP-9 and TIMP-1 positive cells in lung tissue of rats in normal control group was less,showing negative or weak positive expression;the number of MMP-9 and TIMP-1 positive cells in model group was more,showing strong positive expression;the number of MMP-9 and TIMP-1 positive cells in lung tissue of rats in each dose group of Liuwei Buqi recipe and Guilong Kechuanning group decreased,showing weak positive expression,among which,the high dose group of Liuwei Buqi recipe showed weak positive expression Expression.The expression of MMP-9 and TIMP-1 was moderate or weak in the middle dose group and Guilong Kechuanning group.3.5 Changes of TNF-?,IL-1?,IL-8 and TGF-?1 levels in serum and BALF of rats in each group Compared with the normal control group,the levels of TNF-?,IL-1?,IL-8 and TGF-?1 in serum and BALF of the model group were significantly higher(P<0.01).Compared with the model group,the levels of TNF-?,IL-1?,IL-8 and TGF-?1 in serum and BALF in the high dose group of Liuwei Buqi recipe and Guilong Kechuanning group decreased significantly(P<0.01 or P<0.05),especially in the high dose group of Liuwei Buqi recipe.Compared with Guilong Kechuanning group,the levels of TNF-?,IL-1?,IL-8 and TGF-?1 in serum and BALF of Liuwei Buqi recipe high dose group decreased,but the difference was not statistically significant(P>0.05);TNF-?,IL-1?,IL-8 and TGF-?1 in serum of Liuwei Buqi recipe middle and low dose group The levels of TNF-?,IL-1?,IL-8 in serum and BALF of the low dose group of Liuwei Buqi recipe increased significantly(P<0.01 or P<0.05).3.6 Changes of TGF-?1,Smad2/3,Muc5 ac,Col-I and a-SMA protein expression and m RNA level in lung tissue of rats in each group Compared with the normal control group,TGF-?1,Smad2/3,Muc5 ac,Col-I and a-SMA protein expression and m RNA level in the model group increased significantly(P<0.01).Compared with the model group,the expression of TGF-?1,Smad2/3,Muc5 ac,Col-I and a-SMA protein and m RNA level in Liuwei Buqi recipe high dose group and Guilong Kechuanning group decreased significantly(P<0.01 or P<0.05),especially in Liuwei Buqi recipe high dose group.Compared with Guilong Kechuanning group,Liuwei Buqi recipe high dose group decreased,but the difference was not statistically significant(P>0.05);Liuwei Buqi recipe middle and low dose group increased,Liuwei Buqi recipe low dose group increased significantly(P<0.01).4.Conclusion 4.1 Liuwei Buqi recipe can reduce the pathological changes of lung tissue in COPD model rats and increase the pulmonary ventilation function.4.2 Liuwei Buqi recipe can alleviate airway remodeling by reducing the relative expression of MMP-9 and TIMP-1 in lung tissue of model animals.4.3 Liuwei Buqi recipe can improve airway remodeling by inhibiting the release of related inflammatory cytokines.4.4 By regulating the expression of TGF-?/Smads cell signal pathway related proteins and gene level,it may be one of the targets of Liuwei Buqi recipe to intervene the airway remodeling of COPD.