To Investigate The Therapeutic Effect Of Scutellarin On Age-related Macular Degeneration Based On Network Pharmacology

ObjectiveTo explore the protective effect and mechanism of Scutellarin on agerelated macular degeneration by network pharmacology,and to provide a new therapeutic strategy for clinical prevention and treatment of AMD.MethodsIn this study,a total of 30 male SD rats of about 200 g were randomly selected and divided into 3 groups,10 rats in each group,which were divided into 3 groups as follows:(1)control group,(2)sodium iodate model group and(3)Scutellarin treatment group,sodium iodate model group and Scutellarin treatment group were given sodium iodate(50 mg/kg)via tail vein to induce age-related macular degeneration model,and the Scutellarin treatment group was given Scutellarin(50mg/kg.d)intraperitoneally every day for 4 weeks after successful modeling.The control group and sodium iodate model group were given the same dose of normal saline by intraperitoneal injection.Retinal thickness was measured by optical coherence tomography(OCT)at 1 day,14 days and 28 days after the model was successfully established.Rat RPE cells were selected and divided into three groups:(1)control group,(2)sodium iodate model group and(3)Scutellarin treatment group.CCK8 assay was used to detect the proliferation of RPE cells.Annexin V-FITC and flow cytometry were used to detect the apoptosis of RPE cells.The level of reactive oxygen species(ROS)was detected to evaluate the oxidative stress injury of RPE cells induced by Scutellarin.TCMSP Genecard and Swiss Targe t Prediction databases were used to query and screen the targets of Scutellarin.Genecard database was used to collect the pathogenic gene targets related to oxidative stress and age-related macular degeneration,and the online Venn diagram website was used to obtain the common targets of Scutellarin,oxidative stress and age-related macular degeneration.The protein-protein interaction network was established by STRING database and 10 core targets were obtained.Cytoscape software was used for image optimization,and the gene ontology(GO)function analysis and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway analysis were performed online using the wechat website.Molecular docking was used to verify the interaction between Scutellarin and its core targets,and quantitative polymerase chain reaction(q PCR)was used to verify the expression of key genes in cells.ResultsThe OCT results of rats showed that the structure of each layer in the control group was uniform and orderly.Compared with the sodium iodate model group,the high reflective spots of RPE layer were reduced,the disorder of photoreceptors and outer nuclear layer was alleviated,and the retinal thickness was increased significantly in the Scutellarin treatment group.Cell CCK8 results showed that the number and viability of cells in the Scutellarin treatment group increased compared with the sodium iodate model group.Annexin V-PI apoptosis assay showed that the survival rate of RPE cells was significantly decreased and the apoptosis rate was increased after the treatment of sodium iodate.Compared with the sodium iodate model group,the apoptosis of RPE cells was effectively reduced in the treatment group of Scutellarin.The results of ROS detection showed that the survival rate of RPE cells was significantly decreased after the induction of sodium iodate,and the level of ROS was significantly increased and the expression of ROS was enhanced.Compared with the sodium iodate model group,the Scutellarin treatment group effectively reduced the expression of ROS in RPE cells in vivo.The potential targets of Scutellarin were predicted by TCMSP,Genecard and Swiss Target Prediction databases.A total of 148 targets were screened,and 121 specific targets of Scutellarin were screened.A total of 10022 oxidative stress targets and 4123 age-related macular degeneration disease targets were searched in Genecard database.A total of 66 intersection targets of Scutellarin,oxidative stress and AMD were identified.Cytoscape screened 10 core genes,including CASP3,STAT3,MTOR,PTGS2,TNF,HRAS,EGFR,AKT1,BCL2L1 and HIF1 A.Molecular docking results showed that there was a good binding activity between SCU and the core target.The results of PCR showed that the expression levels of CASP3,MTOR,PTGS2,TNF,EGFR,BCL2L1 and HIF1 A in the sodium iodate model group were significantly higher than those in the control group after SCU treatment of rat RPE cells.SCU treatment could effectively regulate gene expression,among which the contents of MTOR,PTGS2,TNF,EGFR and BCL2L1 were decreased,and the contents of CASP3 and HIF1 A were increased,which was statistically significant compared with sodium iodate model group.The expression levels of STAT3,HRAS and AKT1 genes in the sodium iodate model group were decreased,and SCU treatment effectively increased gene expression,and the difference was statistically significant.ConclusionsScutellarin plays an important role in the treatment of age-related macular degeneration by inhibiting apoptosis and protecting retinal cells from apoptosis.The pathways involved mainly include VEGF signaling pathway,AGE-RAGE signaling pathway,HIF-1 signaling pathway,etc.