| Objective: By establishing a rat model of pulmonary fibrosis,ginkgo biloba extract was selected as a treatment drug to explore the effects of ginkgo biloba extract on P38 MAPK pathway and EMT,to clarify the intervention mechanism of ginkgo biloba extract on pulmonary fibrosis,and to provide theoretical basis for seeking multi-target prevention and treatment of pulmonary fibrosis from the perspective of traditional Chinese medicine.Methods: Sixty SPF male SD rats were adaptive fed for one week.Then they were randomly divided into blank control group,pulmonary fibrosis model group,pirfenidone positive group,Ginkgo biloba extract treatment group,each with 15 rats.Except blank control group,the other three groups were replicate the mouse model of pulmonary fibrosis by bleomycin(5mg/kg)tracheal injection.The day of modeling was marked 0 day.Start on the first day after molding,blank group and model group were given intragastric administration of normal saline once a day,and pirfenidone group and GEB group were given pirfenidone solution and ginkgo biloba extract solution once a day.Continuous gavage lasted for 28 days.Day 14 and day 28 after modeling,lung tissue samples were collected for experimental use.HE staining and Masson staining were used to observe the changes of lung tissue morphology and score the fibrosis,hydroxyproline was used to determine the collagen content.Western blot was used to detect the expression of P38MAPK、Collagen I、Collagen III、E-cadherin and α-SMA in lung tissues of each group.Results: 1.General conditions: In blank group,the rats were in good general state of spirit and appetite,with normal weight growth and no death during the experiment;The rats in model group had poor mental state and gradually decreased body weight,accompanied by obvious tachypnea or heavy breath sound,some of them had oral and nasal bleeding,dry and dull hair or shedding,decreased activity and poor appetite,and 3 rats died during the experiment.Overlooks the ketone group and ginkgo biloba,initial body weight decreased obviously after building,poor mental state,similar to the symptoms of model group,as the drug continued intervention,by weight,slow growth,spirit,breathing,activity and appetite,generally improved,pyrazole,ketone group of silver and apricot leaf groups all died during the experiment 1and 2.2.HE staining and Masson staining: the blank control group had intact alveolar structure,no thickening of alveolar septum,and only a small amount of blue collagen deposition around the alveolar wall or interstitium.No obvious pathological changes were observed on the 14 days and 28 days day.On day 14,the model group presented severe inflammatory response,with a large number of inflammatory cell infiltration and fibroblast aggregation,alveolar atrophy or collapse,alveolar septum significantly widened,blue collagen fiber deposition around alveoli and interstitium was observed,which was mainly characterized by inflammatory changes and showed a tendency to fibrosis.On day 28,the inflammatory response was improved and the infiltration of inflammatory cells was reduced,but the damage of lung tissue structure was more obvious than before.A large number of blue collagen fibers could be seen in the field of vision,and there were continuous fibrosis foci,presenting diffuse pulmonary fibrosis.After continuous drug intervention,compared with the model group,the above pathological changes were significantly improved or relieved in the pirfenidone group and ginkgo biloba extract group,but the pathological degree was not as serious as that in the model group,and the improvement effect of pirfenidone was better than that of ginkgo biloba extract.3.HYP content determination: compared with blank control group,HYP content in lung tissues of model group increased significantly over time(P < 0.01);After drug intervention,the HYP content in lung tissues of the pirfenidone group and ginkgo biloba group was lower than that of the model group(BOTH P < 0.01),and the HYP content in the pirfenidone group was lower than that of the ginkgo biloba extract group(P < 0.01),suggesting that ginkgo biloba extract can reduce bleomycin-induced lung collagen fiber deposition in rats.4.Western blot protein determination: Compared with blank control group,the expression of P38 MAPK protein in model group was significantly increased at14 days and 28 days,and the expression of coll-I and coll-III markers of pulmonary fibrosis and α-SMA protein marker of interstitial cells were also significantly increased.The expression of e-cadherin protein,a marker of epithelial cells,was significantly decreased.After treatment with ginkgo biloba extract and pirfenidone,the expression of P38 MAPK,Coll-I,Coll–III and α-SMA in lung tissues of the two groups were significantly lower than that of the model group,while the expression of E-cadherin was significantly higher than that of the model group.Compared with the pirfenidone group,the ginkgo biloba extract group had no statistical significance with P38 MAPK in14days and 28days(P > 0.05),the expression of coll-I,Coll-III,α-SMA were statistically significant(P < 0.01).On day 14,e-cadherin protein expression in ginkgo biloba group was higher than that in pirfenidone group(P < 0.01),but lower than that in pirfenidone group on day 28(P < 0.01).Conclusion(s):Both ginkgo biloba extract and pirfenidone can effectively improve the rats early alveolar inflammation and reduce collagen fiber deposition thus playing an anti-pulmonary fibrosis effect,but cannot completely reverse the occurrence of fibrosis.2.Ginkgo biloba extract is less effective against fibrosis than pirfenidone,which is approved in the guidelines for the treatment of IPF.3.Ginkgo biloba extract can affect the process of EMT by increasing the expression of e-cadherin,a marker of epithelial cells,and inhibiting the expression of α-SMA,a marker of mesenchymal cells.4.Ginkgo biloba extract can reduce the expression level of P38 MAPK protein and inhibit P38 MAPK.Regulation of P38 MAPK signaling pathway may be one of the anti-pulmonary fibrosis mechanisms. |
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