Study On The Mechanism Of Tongqiaomingmu ? On Inhibiting IL-6/HIF-1? Signaling Pathway

Objective:The data mining method was used to analyze the medication of Professor Sun he in the treatment of primary glaucoma.According to the results,the pharmacodynamic material basis of TQMM IV was analyzed based on network pharmacology.One of the possible effective targets of TQMM IV in the treatment of primary glaucoma was predicted to be IL-6/HIF-1?signal pathway.The intervention effect of TQMM IV in the treatment of primary glaucoma was observed,which might be the effect of TQMM IV in the treatment of primary glaucoma It is one of the mechanisms of optic nerve protection.Method:The first part:collect the medical records of Professor Sun he in the treatment of primary glaucoma from January 2010 to September 2020,select270 cases of them as the research object,establish a database,use frequency statistics,association rules and other methods to conduct data mining on the general situation,TCM syndrome types,pathogenesis,prescription frequency,etc.involved in the medical records This paper studies and analyzes the mining results,and summarizes Professor Sun he's clinical experience and academic thought in the treatment of this disease.The second part:select Professor Sun he's basic prescription TQMM IV for network pharmacology analysis,use tcmsp platform to search the active ingredients and core regulatory targets in TQMM IV,select disease-related targets through genecards,OMIM and other disease databases,and construct"drug disease"target network map;To predict the biological mechanism of TQMM IV in the treatment of primary glaucoma,go function analysis and KEGG enrichment analysis of"drug disease"target were carried out,and molecular docking was used to verify.The third part:the apoptosis model of mouse retinal ganglion cells was established by H₂O₂induction.The mice TQMM IV medicated serum was used to intervene,and the cell morphology was observed.The apoptosis rate of retinal ganglion cells was detected by flow cytometry and TUNEL,the level of ROS was detected by flow cytometry and immunofluorescence,and IL-6 mRNA and HIF-1?were detected by RT-PCR The expression of IL-6/HIF-1?pathway related proteins was detected by Western blotting to explore the protective mechanism of TQMM IV on retinal ganglion cells.Result:Part one57 drugs with frequency more than 10 times were 57,among which,the five drugs with the highest frequency were Chaihu(240 times),Shichanggpu(161 times),Gegen(144 times),Danggui(137 times),Chuanxiong(118times);the common drug combination was Chaihu+shichangpu(146 times),Chaihu+Gegen(130 times),Chaihu+Danggui(123 times),shichangpu+Gegen(122 times),Chaihu+Shichangpu+Gegen(113 times),The core combination is:Chaihu,Gegen,Shichangpu,Danggui,Lulutong and Fuling;core combination2:Chaihu,Shichangpu,Chenpi,Huangqin,Banxia and Fuling;core combination 3:Chaihu,Shanyao,Shichangpu,Gegen,Juhua and Danggui.Part twoThe main active components of TQMM IV are quercetin,chanol,isorhamnetin,?-glutasterol and naringenin.103 target points were obtained.The pre protein of degree was IL6,VEGFA,HIF1A,CASP3 and EGFR.The results of KEGG enrichment analysis of"drug disease"mainly play a role in the treatment of primary glaucoma by regulating HIF-1,T cell receptor,FOXO and other signal pathways.Part three1.morphological observation:the cells in model 1 and model 2 showed obvious morphological abnormalities at three time points,including the shrinkage of the cell,the irregular arrangement of the cell body and the inconspicuous changes of the protrusion.The cell morphology of each intervention group was improved and was closer to the blank group than that in the model group.2.the apoptosis rate of retinal ganglion cells showed that the apoptosis rate of each drug intervention group was lower than that of model 1 and model2,P<0.05;TUNEL method showed that the apoptosis rate of each drug intervention group was significantly reduced,P<0.05.3.The expression of ROS positive cells in each group showed that the expression of ROS positive cells in each drug intervention group was lower than that in model 1 group and model 2 group,P<0.05;The expression of ROS positive cells detected by immunofluorescence method showed that the expression of ROS positive cells in each drug intervention group was lower than that in the model group,P<0.05.4.The results of RT-PCR showed that:compared with the blank group,IL-6 mRNA and HIF-1 mRNA in model 1 group and model 2 group were significantly increased?mRNA The expression was significantly increased(P<0.05);Compared with the model group,IL-6 mRNA and HIF-1 mRNA in each drug intervention group were significantly higher?mRNA The expression was decreased(P<0.05).5.The results of Western blot showed that compared with the blank group,IL-6 and HIF-1 in model 1 group and model 2 group were significantly higher,The expression was increased(P<0.05);Compared with the model group,the levels of IL-6 and HIF-1 in each drug intervention group were significantly higher,The expression was significantly decreased(P<0.05).Conclusion1.Professor Sun he's core drugs for glaucoma mainly include Chaihu,Shichangpu,Gegen,Danggui,Yujin,Danshen,etc.2.The main active components of TQMM IV have anti-inflammatory,anti-oxidation,and improve the survival rate of retinal ganglion cells.The regulation of inflammatory related pathways,such as HIF-1,T cell receptor,FOXO and so on,may be the mechanism of TQMM IV in the treatment of primary glaucoma.3.TQMM IV can inhibit the apoptosis of retinal ganglion cells induced by purified H₂O₂in vitro and enhance the cell activity.Down regulating the expression of IL-6 mRNA and HIF-1?mRNA and inhibiting the activation of IL-6/HIF-1?pathway may be one of the mechanisms of its prevention and treatment of primary glaucoma.