Therapeutic Effect Of Jilizhiyangfang On Psoriasis Vulgaris In Mice And Based On Network Pharmacological Analysis And Experimental Verification

Objective:This study was to explore the therapeutic effect of jilizhiyangfang(JLZYF)on Imiquimod(IMQ)induced psoriasis model mice and the effect on skin barrier function;and based on network pharmacological analysis and experimental verification of the potential molecular mechanism of JLZYF in the treatment of psoriasis,so as,to lay a foundation for the study of Chinese herbal compound with definite therapeutic effect on psoriasis.Methodology:1.At first,six BALB/c mice were divided into blank control group and model control group,and the models were treated according to the relevant literatures.After the models were established,the skin tissues of the IMQ-induced psoriasis model mice were observed pathologically,the formation of Munro microabscess,the characteristic pathological manifestation of psoriasis,was found under the light microscope.2.Sixty BALB/C mice were randomly divided into Blank control group(Blank),Model control group(Model),Positive control group(Positive),JLZYF low concentration group(J-L),JLZYF middle concentration group(J-M)and JLZYF high concentration group(J-H),the mice were given orally for 7 days,during which the diet and activity of the mice were recorded,the body weight of the mice was taken and the trend of body weight change was recorded,the skin lesions on the back of the mice were observed,and the data of the Organ Index were obtained,the area and severity index(PASI)of psoriatic lesions were recorded and the skin histopathology of the mice were observed,the expression levels of IL-17 A and TNF-α were detected by enzyme-linked Elisa assay(ELISA).The surface water content of mice skin was measured and recorded.The relative expression levels of FLG were detected by Western Blot(WB),to investigate the therapeutic effect of JLZYF on IMQ-induced psoriasis in mice and its effect on skin barrier function.3.Through the research methods of network pharmacology,TCMSP database was used to search the effective components of JLZYF and collect the corresponding action targets;Using Gene Cards,OMIM,Drug Bank,Pharm Gkb,TTD databases to screen targets gene related to psoriasis,and using the online VENNY platform to obtain intersect the targets gene of psoriasis,and to screen the predictive targets genes of JLZYF in the treatment of psoriasis;Using STRING database to build PPI network;Cytoscape3.8.0software was used to build “JLZYF-active ingredients-target genes-psoriasis” interaction network and screen out core action target genes.Then perform KEGG metabolic enrichment analysis and enrichment analysis of GO on the selected target genes;The core action target and active ingredients were selected as receptor proteins and ligand small molecules.moreover,using Auto Dock Tools-1.5.6 to carry out molecular docking of ligand and receptor,and the molecular docking results was visualized by Pymol software,to analyze its potential mechanism of action on psoriasis from molecular level.4.Experimental verification,thirty BALB/C mice were randomly divided into Blank control group(Blank),Model control group(Model)and JL Group(JL),oral administration for 7 days.At the end of the experiment,skin tissue was taken from the back of the mice.Real-time fluorescent quantitative PCR was used to detect the expression of HSP90AA1 mRNA,in order to supplement the experimental data of potential mechanism of JLZYF in treating psoriasis.Results:1.In the model experiment,compared with the blank control group,IMQ was used to daub the skin on the back of mice for 7 days continuously to make the model,The typical psoriatic changes such as erythema,scales and thickened skin lesions were observed.The results of HE staining showed that the keratinocytes were thickened,inflammatory cells infiltrated,and the formation of Munro microabscess,It is suggested that the model is successful.2.In the formal experiment,(1)the general condition of mice in each group was better;except Blank group,the weight of mice in each group was decreased;(2)Compared with the model control group,the skin lesion of d8 mice was improved and the PASI score was decreased,especially in the JLZYF low concentration group;(3)Compared with Blank group,Thymus index was decreased and spleen index was increased in all groups,and IMQ stimulation could cause thymus atrophy and spleen enlargement.The spleen indexes of Positive,J-L,J-M and J-H groups were lower than those of Model group,indicating that JLZYF can regulate the immune response of psoriasis;(4)HE staining of the skin tissue showed that the skin histopathology of the mice in each group were changed compared with those in the Blank group,the epidermis of Positive,J-L,J-M and J-H groups became thinner;(5)ELISA showed that the expression of IL-17 A and TNF-α in the serum of Positive,J-L,J-M and J-H groups were decreased compared with that of Model group,and the decrease of IL-17 A and TNF-α in J-L group was the most significant,it is suggested that JLZYF can alleviate the inflammatory reaction of psoriasis;(6)Compared with the Model group,the water content in the epidermis of Positive,J-L,J-M and J-H groups was increased,which suggested that the JLZYF could improve the dry and desquamation symptoms of psoriasis;(7)The expression of FLG protein in the skin of Positive and J-L groups was higher than that of the Model group,and the expression of FLG protein in J-L group was higher than that of the Model group,it is suggested that the JLZYF can repair the damage of skin barrier function of psoriasis by regulating the expression of FLG protein.3.Through the research methods of network pharmacology,such as isorhamnetin,luteolin,quercetin,kaempferol,β-Carotene,β-sitosterol effective components of JLZYF.Through CCND1,MYC,TP53,CTNNB1,TNF,HSP90AA1,CDK4,CCL2,NFKBIA,RELA and other key targets;And PI3K-Akt signaling pathway,apoptosis,Toll like receptor signaling pathway,apoptosis,Th17 cell differentiation,JAK-STAT signaling pathway and other major pathways,Psoriasis was treated by anti inflammation,regulating immunity,regulating cell proliferation and apoptosis.Molecular docking verified that luteolin,quercetin,kaempferol,isorhamnetin,β-Sitosterol,β-Carotene combines stably with core targets CCND1,MYC,TP53,TNF,HSP90AA1,NFKBIA and RELA.This study has a certain reference significance for clarifying the molecule mechanism of JLZYF in the remedy of psoriasis.This study provides a direction for future experiments to verify the therapeutic mechanism of JLZYF on psoriasis.4.Experimental verification,Compared with Blank,the expression of HSP90AA1 mRNA was significantly decreased in the skin of Model and JL mice(p<0.01);and the expression of HSP90AA1 mRNA was increased in the skin of JL mice compared with Model,the difference was significant(p<0.01),suggesting that JLZYF can regulate cell proliferation and apoptosis,participate in the body’s immune regulation and inflammatory response by regulating the expression of HSP90AA1 mRNA,to improve the scaling,pruritus and other symptoms of psoriasis.Conclusions:1.The JLZYF can effectively relieve the clinical manifestations of erythema,scale and epidermis thickening in the lesions of psoriasis.JLZYF can down-regulate the expression of IL-17 A and TNF-α in a dose-dependent manner,and affect the organ index of of immune organs thymus and spleen,it is suggested that JLZYF can alleviate the clinical symptoms of psoriasis by participating in inflammatory reaction and immune regulation.2.JLZYF can effectively improve the water content of skin epidermis and significantly increase the expression of FLG protein in order to repair the skin barrier function of psoriatic lesions.3.the potential mechanism of JLZYF is anti-inflammatory,anti-pruritic,repairing skin barrier,regulating apoptosis and proliferation of cells and regulating immune response in vivo;Furthermore,multi-component,multi-target and multi-pathway play a synergistic role in the treatment of psoriasis.4.the experiment proved that the JLZYF can promote the expression of HSP90AA1,so as to inhibit the apoptosis of cells,participate in the immune regulation and inflammatory reaction of the body and slow down the clinical manifestation of psoriasis.