A Study On The Effects Of Melatonin And Yupingfeng Powder On Immune System In Mice Based On "The Correspondence Between Nature And Human" Theory And Network Pharmacology

Object:Based on the theory of "correspondence between nature and human" in traditional Chinese medicine,the artificial simulation system is used to study and observe the effects of Yupingfeng Power and melatonin on the immunity of mice under sudden changes in temperature.Screening potential targets of Yupingfeng Powder and melatonin using pharmacology network of traditional Chinese medicine,predicting possible mechanisms and key signal pathways,analyzing and exploring the Molecular biology mechanism of effects of sudden temperature change intervention and Yupingfeng Powder combined with melatonin intervention on the immune system.Methods:1.Firstly,the potential targets of Yupingfeng Powder and Melatonin were predicted and screened by using the traditional Chinese medicine network pharmacology.Use the SymMap database to query the potential targets of Huangqi,Baizhu and Fangfeng in Yupingfeng Powder,and overlap the target of Yupingfeng Powder with the targets of melatonin in the CTD disease database to obtain Yupingfeng Powder-melatonin's common targets.The potential drug targets were used to constructed a protein interaction network(PPI network)by using the STRING protein interaction online analysis tool.CluGO in Cytoscape software was used to perform GO function analysis and KEGG signal pathway enrichment analysis on the constructed PPI network.The difference is a statistically significant result.2.Animal experiments were used to further verify the effects of Yupingfeng Powder and Melatonin on the immune system of mice under the intervention of sudden temperature changes.D-galactose was used to make the mouse model of aging-immunity inferiority.The mice were randomly grouped and divided into two drug intervention factors according to the two factor 4x3 factorial design:Yupingfeng powder group,melatonin group,Yupingfeng&Melatonin group,physiological saline group;intervention factors of sudden temperature change:sudden temperature drop group,sudden temperature rise group,normal temperature control group,a total of 12 groups,6 in each group.Mice were administrated to with Yupingfeng powder and melatonin for 6 weeks.After gavage,the mice were placed in artificial temperature simulation system for intervention.The temperature dropped sharply from 30 degrees Celsius to 8 degrees Celsius within 30 minutes,and the temperature rose sharply from 8 degrees Celsius to 30 degrees Celsius within 30 minutes.The relative humidity keeps the humidity in natural conditions,and the lighting time refers to natural light for 10 hours.After the experiment,ELISA was used to detect the expression of IL-1?,IL-6,TNF-? in mouse serum and NFkB,NLRP3,SIRT1,and SIRT2 proteins in mouse lung tissues.Qrt-PCR was used to detect mouse lungs NRP2 gene expression level in tissues.Results:1.According to the pharmacological analysis of traditional Chinese medicine network,there are 85 joint targets of Yupingfeng Powder and melatonin.Using STRING to build a protein interaction network to obtain 20 key nodes with the highest degree of node,TP53,AKT1,ALB,JUN,IL6,CASP3,VEGFA,MAPK3,MAPK1,TNF,ESR1,PTGS2,FOS,CAT,HMOX1,SIRT1,CCND1,RELA,IL1B,BCL2L1.Using CluGO to perform GO function analysis and KEGG enrichment analysis on 20 key targets and 85 common targets respectively,we learned that Yupingfeng Powder and melatonin are related to multiple immunological processes,including thymus development,giant Phage differentiation,regulation of immune response cytokines,T cell homeostasis,negative regulation of apoptosis induced by oxidative stress,regulation of acute inflammatory response,regulation of chemokines,etc.,also involve the NF-kB signaling pathway,mitochondria Autophagy,longevity regulatory pathway,T cell receptor signaling pathway,TNF signaling pathway,IL-17 signaling pathway and other signaling pathways closely related to immune,aging and oxidative stress processes.2.Yupingfeng Powder can increase serum IL-1?(P<0.05),melatonin can reduce IL-1?(P<0.05)and TNF-?(P<0.05),the combination of Yupingfeng Powder and melatonin can increase IL-1?(P<0.05)and IL-6;Yupingfeng Powder and melatonin can increase the expression of NF-kB,SIRT1,SIRT2,NLRP3 protein in the lungs of mice;in the case of combination of Yupingfeng powder and melatonin,the expression of NRF2 gene was significantly up-regulated(P<0.05).Sudden changes in temperature have no significant effect on serum IL-1?,IL-6,TNF-a and other indicators.Sudden decrease of temperature significantly reduced the expression of NFkB,NLRP3,SIRT1,and SIRT2 proteins in mouse lungs(P<0.05).The sudden increase in temperature reduced the expression of SIRT1 relative to the control group(P<0.05).Both the sudden drop in temperature and the rapid rise in temperature significantly reduced the expression level of NRF2 gene in the lung tissue of mice(P<0.05).Conclusion:The study found that Yupingfeng Powder and Melatonin shared some of the same targets and signal pathway mechanisms for regulating the immune system.Yupingfeng Powder and Melatonin have dual regulatory effects that can promote inflammation as well as resist inflammation.The combination of Yupingfeng Powder and Melatonin can produce synergistic effects and have obvious anti-inflammatory and antioxidant effects,which is consistent with the inference that Yupingfeng Powder and melatonin have the same targets and mechanisms as predicted by the network pharmacology mentioned above.The drastic changes in temperature have interfered with the normal functions of the body's immune system's anti-inflammatory and anti-oxidation to a certain extent,and the cold factors with the sudden drop in temperature have a more significant impact on the immune system.This study provides the basis for further research on using TCM theory to prevent immune system dysfunction caused by climate change and abnormal biological rhythms.