Clinical Observation And Experimental Study Of Bushen Huoxue Decoction In The Treatment Of CTD-ILD And On Intervention Of Pulmonary Fibrosis Via Regulating Of Cell Autophagy Pathway

Objective:Clinical observation to explore the clinical efficacy of Bushen Huoxue Decoction（BHD）in the treatment of connective tissue disease associated interstitial lung disease（CTD-ILD）of kidney deficiency and blood stasis and its effect on the changes of Pa O₂ and FVC%pred levels.Experimental study to to evaluate the anti-pulmonary fibrosis effect and mechanism of BHD in bleomycin（BLM）-induced pulmonary fibrosis in mice.Method:In the clinical study,60 patients with kidney deficiency and blood stasis were randomly divided into treatment group（Bushen Huoxue recipe+western medicine standard treatment）and control group（western medicine standard treatment）.30 cases in each group were treated continuously for8 weeks.The improvement of Pa O2,FVC%pred,clinical symptom and sign score and clinical effective rate were observed before and after treatment.2.Experimental studies:（1）Animal modeling and grouping:120 healthy C57BL/6 male mice were randomly divided into four groups:blank control group,sham operation group,BLM group and BHD group,with 30 mice in each group.Mice in the BLM group and the BHD group were modeled by tracheal instillation of BLM,and the sham operation group model was constructed by tracheal instillation of equal volume of normal saline.After 24 hours of modeling,the mice in the BHD group were given daily intragastric administration,and the other three groups were given equal volume normal saline intragastric administration.（2）Samples collection and testing:On the 7th,14th and 28th day after drug treatment.The lung tissues were collected for histopathological examination（H&E and Masson three-color staining）,and the expression levels of COL III,IL-17A,PI3K-p110αsubunit and MTOR protein in the lung tissues of mice in each group were detected by immunohistochemistry.The expression levels of Col3a1 m RNA,Map1Lc3b m RNA,Il17a m RNA,Pik3ca m RNA and Mtor m RNA in lung tissues of mice in each group were detected by q RT-PCR.The expression levels of LC3-I,LC3-II,p-GSK3β（ser9）and GSK3βin lung tissues of mice in each group were detected by Western blot.（3）Data analysis:The experimental results of each group were statistically analyzed to explore the specific mechanism of regulating autophagy by BHD in the intervention of pulmonary fibrosis.Results:1.Clinical observation:（1）There were no statistical differences in age distribution,gender composition,primary morbidity and course of disease between the two groups,which were comparable.（2）After treatment,Pa O2 level of the two groups was significantly higher than before treatment,and the improvement degree of the treatment group was significantly better than that of the control group,with statistical significance.（3）The level of FVC%pred in the two groups after treatment was significantly higher than that before treatment,and the improvement degree in the treatment group was significantly better than that in the control group,with statistical significance.（4）After treatment,the scores of symptoms and signs of the two groups were significantly lower than before treatment,and the improvement of symptoms and signs in the treatment group was significantly better than that in the control group,with statistical significance.（5）The clinical effective rate of the treatment group was significantly higher than that of the control group.2.Experimental study:BHD can significantly reduce the inflammatory response of mouse lung tissue,and reduce the excessive deposition of ECM represented by type III collagen accumulation;At the same time,BHD can significantly increase the expression levels of Map1Lc3b m RNA and LC3-II/LC3-I,and decrease the expressions of IL17A protein,Il17a m RNA and its downstream signaling molecules PI3K-p110αsubunit,Pik3ca m RNA and p-GSK3β（ser9）.However,there was no significant effect on the expression of MTOR and Mtor m RNA.Conclusions:（1）BHD can improve clinical symptoms and signs,improve lung function and improve the quality of life of patients.（2）BHD can reduce chronic inflammation,reduce the accumulation of type III collagen,interfere with the deposition of extracellular matrix,and delay the destruction of alveolar structure in mouse pulmonary fibrosis model.This process may be achieved by inducing autophagy through inhibition of IL-17A/PI3K/GSK3βsignaling pathway.