To Explore The Molecular Mechanism Of Qiliqiangxin Capsule In The Treatment Of Diabetic Cardiomyopathy Based On Network Pharmacology

Objective:As the number of patients with diabetes grows year by year,the number of people with diabetic cardiomyopathy(DCM)also increases.But it has no obvious symptoms for a long time,and its molecular mechanism has not been elucidated.Therefore,this thesis used H9c2 cardiomyocytes to establish a vitro model of DCM,combined with transcriptome sequencing(RNA-seq)technology and bioinformatics analysis,in order to reveal the pathogenesis of DCM.In addition,the potential mechanism of traditional Chinese medicine on DCM has less been investigated,and the previous studies are mainly about compound Chinese medicine prescriptions,but not active ingredients of Chinese medicine.Therefore,we used network pharmacology and literature research to screen the compounds of Qili Qiangxin capsules and obtain their targets,and further integrated the analysis with RNA-seq results to find compounds with targets that were consistent with disease targets.The efficacy of the drug was verified through biological experiments.This project explored the pathogenesis of diabetic cardiomyopathy,the effect of the effective components of Qili Qiangxin capsule on diabetic cardiomyopathy and its mechanism.It provided theoretical and data support for Qili Qiangxin capsule in the treatment of diabetic cardiomyopathy,and promoted the process of Chinese herbal medicine for the treatment of diabetic cardiomyopathy.Materials and methods:(1)H9c2 cardiomyocytes were cultured with high glucose(HG,DMEM medium containing 33 mmol·L-1 glucose)to establish an in vitro cardiomyocyte injury model,and RNA-seq technology was used to obtain differentially expressed genes GO-rich and KEGG were performed to predict the function of differentially expressed genes and the key pathways for diabetic cardiomyopathy.(2)By searching the TCMSP database and consulting the literature,the main compounds of Qili Qiangxin capsules were screened,and the corresponding targets were integrated.The DAVID database was used for KEGG enrichment analysis of targets,and was further integrated with RNA-seq results to obtain the main functional ingredient in Qili Qiangxin Capsule.(3)H9c2 cardiomyocytes were cultured with high glucose(HG,DMEM medium containing 33 mmol L-1 glucose)to establish an in vitro DCM model,and the effects of HG and Tan IIA on cell viability were determined by the CCK-8 method.The change of H9c2 myocardial cell cycle after HG and drug intervention was tested.Furthermore,the expression of cell cycle-related mRNA was detected by real-time fluorescent quantitative PCR(qRT-PCR).Result:RNA-seq analysis obtained a total of 1416 genes with different expression level in the high glucose group and the control group,of which 546 genes were up-regulated and 870 genes were down-regulated.KEGG pathway enrichment analysis showed that differentially expressed genes were enriched in cell growth and death,transport and catabolism.The GO enrichment analysis results showed that these genes may play a role by participating in biological processes such as the cellular process.These results indicated that cell growth and death processes were altered in diabetic cardiomyopathy.Network pharmacological analysis screened 230 Qili Qiangxin capsule compounds,and obtained a total of 56 main constituents through literature research.These two methods were integrated to obtain 5 active drugs(hypaconitine,quercetin,tanshinone IIA,cryptotanshinone,formononetin),of which quercetin corresponded to 154 targets,Tan IIA corresponded to 41 targets,cryptotanshinone corresponded to 30 targets,and formononetin corresponded to 23 targets.By comparing and analyzing the target components of each compound and the RNA-seq results,Tan IIA was the main effective component in Qili Qiangxin capsule that could affect the cell cycle process.The protective effects of Tan IIA on rat H9c2 cardiomyocyte injury induced by high glucose were verified by biological experiments.The results suggested that Tan IIA had a significant protective effect on H9c2 cardiomyocyte injury induced by high glucose by regulating the cell cycle.Conclusion:This study has provided transcriptome data for an in vitro model of diabetic cardiomyopathy.Functional analysis of differentially expressed genes has suggested that high glucose can affect cell growth and death,then influence cell cycle.Network pharmacological analysis has indicated that the Tan ?A,a functional component in Qili Qiangxin capsule,may alleviate high glucose-induced cell damage by regulating cell cycle-related mRNA.Biological experiments have shown that Tan IIA can improve cell cycle arrest caused by high glucose by regulating cell cycle related gene expression.