The Effect Of Spleen-Strengthening And Lung-Supplementing Liquid On Immune Function And Airway Inflammation Of Chronic Obstructive Pulmonary Disease With Lung-Spleen Qi Deficiency Syndrome In Rats

[Objective]This study aims to explore the effect of Spleen-Strengthening and Lung-Supplementing Liquids on immune function and airway inflammation in rats during stationary COPD phage, and to demonstrate whether through the NF-κB signaling transduction pathway Spleen-Strengthening and Lung-Supplementing Liquid regulates immune function and inhibit airway inflammation in rats with COPD, and to provide the experimental basis for the clinical treatment of Spleen-Strengthening and Lung-Supplementing Liquids to patients with COPD. Further more, this paper is to demonstrate the feasibility and significance of the treating method of Strengthening Spleen and Supplementing Lung as the treatment of stable COPD phage, and to have a further understanding of the relationship between TCM syndrome and objective indicators (immune dysfunction, and airway inflammation indicators, etc).In addition, the study is aimed to deepen the understanding of lung-spleen deficiency syndrome as the nature of the stable COPD phage, and to make TCM syndromes objective. All above is going to have great significance to basic research of guiding Chinese medical therapy for stable COPD phage.[Methods]1. Theoretic exploring aspect:TCM pathogenesis of chronic obstructive pulmonary disease was in-depth explored. In addition, the basis of spleen-strengthening and lung-supplementing treatment was deeply analyzed. And furthermore, the description explanation of Spleen-Strengthening and Lung-Supplementing Liquids was discussed. What's more, the feasibility of Spleen-Strengthening and Lung-Supplementing Liquids on regulating immune function and inhibiting airway inflammation in patients with chronic obstructive pulmonary disease.2. experimental research aspect:65SD adult male rats were randomly divided into six groups(10of the blank control group,11of every other groups), i.e. the model group, Spleen-Strengthening and Lung-Supplementing Liquids group respectively in large dose, middle dose and small dose, and positive drug (Dexamethasone) control group. Rats was treated with cigarette smoke and LPS complex factors to build the model of COPD in syndrome pattern of lung-qi deficiency. And then the model of spleen-qi deficiency was derived from gavaging rats with rhubarb liquid. Model-making lasted for28days. Rats were administrated in accordance with the experimental design for14days.28days later, the indicators were determined. Afterwards, the whole situation of the rats in each group was observd, and the basic pathological changes were observed after HE staining. And then after MASSON staining, collagen content and smooth muscle thickness in each group was observed. The bronchial smooth muscle thickness, internal diameter of the trachea was determined by application image analysis system, while the index of bronchial smooth muscle and area of collagen fibers were calculated. With arterial blood treated of anticoagulation, PaCO2, PaO2, PH and SaO2and other arterial blood gas indicator in rats were measured by arterial blood gas analysis. After the thymus, spleen of rat peeled, and thymus index, spleen index was calculated according to body mass. Next, blood was taken after anesthesia, then treated with anticoagulation, and expression rate of CD3+, CD4+, CD8+in the blood were determined by flow cytometry. Meanwhile CD4+/CD8+ratio was calculated. In addition, partial blood dealt with centrifugeserum, keeping serum, IgG, IgA, and IgM levels and IL-8and TNF-alpha activity were measured by ELISA. After blood taken, mRNA was extracted by trizol grinding after rat lung tissue frozen in liquid nitrogen. After reverse transcription, designed primers were dealt with real-time PCR amplification. With the internal reference for the gene expression of control, IL-8, TNF-alpha mRNA in the lung tissue were observed. tissue proteins of lung tissue were extracted by the kit, and with Western blot (Western blot) method, expression of NF-κB and I-κB in the lung tissue were determined.[Results]1.Though overall observation of animal models and pulmonary histopathologic findings, we confirmed the replication of animal models of patients with COPD of lung and spleen Qi deficiency syndrome was successful, and thus provided a basis for the smooth progress of the experiment.2.Spleen-Strengthening and Lung-Supplementing Liquids was able to significantly improve the overall situation of experimental rats with COPD, such as weight, cough, mental status.3.Spleen-Strengthening and Lung-Supplementing Liquids apparently inhibited the airway inflammation in rats, reduced collagen content, collagen fiber area index of bronchial smooth muscle, improve airway remodeling, with the efficacy similar to or superior to Dexamethasone, especially the efficacy in Large and middle dosage groups was better.4.Spleen-Strengthening and Lung-Supplementing Liquids was able to improve the hypoxic state of the rats, thus improving the survival rate of rats as well as the life quality. However, there was no significant improvement in PaCO2, PH.5.Spleen-Strengthening and Lung-Supplementing Liquids can significantly improve cellular and humoral immune function of the rats with COPD, and reduce the number of acute COPD attack.6.Spleen-Strengthening and Lung-Supplementing Liquids was able to inhibit the concentration of inflammatory markers such as IL-8and TNF-alpha and the expression of TNF-amRNA,IL-8and TNF-alphain in the lung tissue, with the efficacy in Large and middle dosage groups better.7. Spleen-Strengthening and Lung-Supplementing Liquids increased body weight of the COPD rats, thymus index, spleen index,and enhanced immunity, and disease prevention and disease resistance.8. Spleen-Strengthening and Lung-Supplementing Liquids reduced expression of NF-κB in lung tissue, but increased I-κB expression. NF-κB signaling transduction pathway plays an important role in airway inflammation reaction and immune regulation during COPD, and Spleen-Strengthening and Lung-Supplementing Liquids improved immune function, inhibited airway inflammation, at the same time downregulated NF-κB in lung tissue expression, upregulated expression of I-κB, from which we would conclude that Spleen-Strengthening and Lung-Supplementing Liquids might adjust immune function and inhibit airway inflammation through the NF-κB signaling pathway in COPD.[Conclusions]1.Lung and spleen Qi deficiency is directly related to the occurrence and development of COPD, which is the critical pathogenesis of chronic obstructive pulmonary disease stable period. Starting from the pathogenesis of Lung and spleen Qi deficiency syndrome, we established the strengthening spleen and supplementing lung treatment, so as to achieve the purpose of supplementing lung Qi.2.Spleen-Strengthening and Lung-Supplementing Liquids, with its description available, selected pathogesis and significant efficacy, enhanced the immune function in rats with COPD, inhibited airway inflammation, and improved the airway remodeling, especially the efficacy in Large and middle dosage groups was better. And some indicators of efficacy were dose-dependent. The mechanism is that Spleen-Strengthening and Lung-Supplementing Liquids may via NF-κB signaling transduction pathway inhibit airway inflammation in rats with COPD, regulate the immune function; and be possible to regulate immune function in rats with by down-regulating inflammatory factors.3.It is Lung and spleen Qi deficiency syndrome combined with animal models of chronic obstructive pulmonary disease to make this study more targeted, and what's more, it builds a bridge between chinese basic study and clinic study, and promotes study of Chinese medical efficacy mechanism. In addition, this study plays an important role in exploring the deep inner link between medical theory and Chinese medicine theory.4. This study deepens the understanding of lung and spleen Qi deficiency syndrome in COPD stable period, and plays an important role in promoting clarification of Chinese medical efficacy mechanism and research and development of new Chinese medicine. In addition, this study provides basis for exploring the super link and target of Chinese medicine treatment of chronic obstructive pulmonary disease.