Study On Mechanism Of The Decoction Of Gubiaozhiti In Regulating The Balance Of Th17/Treg Cells And Mediating Anti-allergic Rhinitis

Objective:This study aims to verify the therapeutic effect of the Decoction of Gubiaozhiti,an empirical prescription of a prestigious TCM doctor,on allergic rhinitis,and reveal the mechanism of the Decoction of Gubiaozhiti in regulating the balance of Th17/Treg cells and mediating anti-allergic rhinitis,so as to provide scientific basis for clinical application of the Decoction of Gubiaozhiti in the treatment of allergic rhinitis,and inherit and innovate academic thoughts of the prestigious TCM doctor.Methods:Allergic rhinitis models were established by nasal drops of ovalbuminn（OVA）and AL（OH）₃ powder,and divided into six groups randomly,i.e.,normal control group,model group,positive group（loratadine group,0.5mg/kg）,and low,medium and high dose groups of the Decoction of Gubiaozhiti（respectively 7.5g/kg,15g/kg and 30g/kg）.Then,weight changes and nasal symptoms（such as rhinocnesmus,runny nose and sneezing）of the mice were observed.In addition,H&E staining was applied to detect the pathological changes of mouse’s nasal tissue.Enzyme linked immunosorbent assay（ELISA）was adopted to detect TGF-βand IL-17 levels in mouse serum of each group.Flow cytometry（FCM）was employed to detect the expression levels of intracellular Th1 cells（CD4⁺IFN-g⁺T cells）,Th17 cells（CD4⁺IL-17⁺T cells）and Treg cells（CD4⁺Foxp3⁺regulatory T cells）in the spleen and lymph nodes.Moreover,the data of pharmaceutical ingredients and their targets,disease-related targets,gene names,etc.were collected and screened out from TCMSP,Uniprot,Gene Cards,Dis Ge NET and other databases.These data were also visually analyzed by Metascape,Cytoscape and other software to predict the potential targets of the Decoction of Gubiaozhiti in treating allergic rhinitis.Results:1.In terms of general symptoms,compared with the normal control group,the allergic rhinitis model group had a significant increase in the scores of mouse’s nasal symptoms,with a significant difference（P<0.01）.2.H&E staining results showed that a large number of inflammatory cells infiltrated into the nasal mucosa,there was obvious glandular hypertrophy in lamina propria,and the ciliated columnar epithelial cells were not arranged with integrity.Compared with the model group,the positive group and the low,medium and high dose groups of the Decoction of Gubiaozhiti had a significant decrease in symptom scores,with a significant difference（P<0.05）.Moreover,compared with the model group,the positive group and the medium and high dose groups of the Decoction of Gubiaozhiti had a significant decrease in the infiltration of inflammatory cells into the nasal mucosa,the glandular hypertrophy in lamina propria was improved,and the ciliated columnar epithelial cells were arranged well.The positive group and the medium dose group of the Decoction of Gubiaozhiti were of the best effect.3.It can be seen from ELISA detection results that compared with the normal control group,the model group had an increase in the level of IL-17 in mouse serum,but there were no significant changes in the level of TGF-β.Compared with the model group,the positive group and the low,medium and high dose groups of the Decoction of Gubiaozhiti had a lower level of IL-17,and there was a significant difference for the middle dose（P<0.05）.Additionally,compared with the model group,the high dose group of the Decoction of Gubiaozhiti had a higher level of TGF-β（a kind of serum cytokine）,with a significant difference（P<0.05）.4.As can be seen from FCM detection results,compared with the model group,the low,medium and high dose groups of the Decoction of Gubiaozhiti had a significant decrease in the proportion of Th17 cells（CD4⁺IL-17⁺T cells）in mouse spleen,with a significant difference（P<0.05）.However,there was no difference in the expression level of intracellular Th17 cells（CD4⁺IL-17⁺T cells）in lymph nodes between the two groups（P>0.05）.Besides,there was also no significant difference in the expression level of Th1 cells（CD4⁺IFN-γ⁺T cells）in both lymph nodes and the spleen between the two groups（P>0.05）.Moreover,the pharmacological intervention of the Decoction of Gubiaozhiti significantly increased the proportion of Treg cells（CD4⁺Foxp3⁺regulatory T cells）in the spleen and lymph nodes of the allergic rhinitis model mice,with a significant difference（P<0.05）.5.It was found in the network pharmacology research that there were a number of effective components that inhibited AR-related genes in the Decoction of Gubiaozhiti,and the corresponding targets mainly focused on the reaction of oxides and oxidative stress,the reaction of cells to chemical stress stimulation,and the substance metabolic reaction.Conclusion:1.The Decoction of Gubiaozhiti has a significant therapeutic effect on allergic rhinitis,and its mechanism is related to inhibiting the inflammation induced by Th17 cells（CD4⁺IL-17⁺T cells）,increasing the expression level of Treg cells（CD4⁺Foxp3⁺regulatory T cells）and regulating the balance of Th17/Treg cells.2.According to the network pharmacology research,it is speculated that the mechanism of the Decoction of Gubiaozhiti in treating allergic rhinitis is synergistic with multiple components,multiple targets and multiple paths.