| Objection:Shexiang Baoxin pill(SBP)is a traditional formulation of Chinese patent medicine,used to treat myocardial infarction of coronary atherosclerotic heart disease in the clinic in China.However,the mechanism of its therapeutic effects has not been clearly elucidated yet.The aim of this study was to investigate the potential cardioprotective mechanism of SBP in the treatment of myocardial infarction(MI)model rats by applying proteomic approach.Materials and methods:1)The rat model of MI was generated by ligating the left anterior descending coronary artery.Eighteen rats were randomly divided into three groups(n = 6 each):the MI group,MI group treated with SBP(SBP),and sham-operated group(SOG).A label-free quantitative proteomic approach was utilized to investigate the whole proteomes of heart tissues from the groups above on the day of the operation(Day 0)and 15 days later(Day 15).2)Developed a model of MI in rats and treated them with SBP(MI+SBP: SBP),the effective component compounds in SBP(ECC)(MI+ECC),benazepril hydrochloride(MI+BH),or vehicle(MI + Vehicle)as a control.A sham operated group with vehicle treatment(SOG + Vehicle: SOG)was used as an additional control group.Tandem Mass Tag(TMT)-based quantitative proteomics methods was utilized to investigate that the effect and therapeutic mechanism of SBP on cardiovascular disease is achieved by regulating multiple pathways.In these two studies,Creatine kinase(CK)and lactate dehydrogenase(LDH)were measured to confirm successful establishment of the MI model and The advantage of SBP’s clinical therapeutic effect was established by measuring left ventricular fractional shortening(LVFS)and left ventricular ejection fraction(LVEF)after treatment for 15 days.The differentially expressed proteins were subsequently analyzed with bioinformatic methods after the quantitative proteomic analysis.Additionally,the expression levels of the promising proteins were validated by Western blotting.Results:1)In the label-free proteomic analysis,the echocardiography analyses showed that SBP treatment significantly preserved the cardiac function of MI rats.Additionally,quantitative proteomics identified 129 differentially expressed proteins,and 15 proteins were considered as logical candidates for explaining the cardioprotective effect of SBP.Bioinformatic analysis of these differentially expressed proteins revealed that the proteins involved in cellular mitochondrial energy metabolism processes,such as fatty acid beta-oxidation and aerobic respiration,were significantly regulated under SBP treatment,of which fatty acid-binding protein 3(FABP3,involved in the peroxisome proliferator-activated receptor(PPAR)signaling pathway and energy metabolism.)and myoglobin(MB,involved in the redox pathway,which is vital in antioxidant defense post-MI.)were significantly downregulated in the MI model group compared with the SOG group and returned to the basal level with SBP treatment,confirmed by Western blotting.2)In the TMT quantitative proteomics analysis,there were no significant difference in echocardiography and whole proteome analysis for ECC group and benazepril hydrochloride group(BH),compared with MI group.However,in the TMT-based proteomic study,the echocardiography analyses showed that SBP treatment significantly preserved the cardiac function of MI rats.A total of 210 significantly changed proteins were utilized to make a Kyoto Encyclopedia of Genes and Genomes(KEGG)analysis,which showed that 15 signaling pathways related to CVD were regulated by SBP.Among these pathways,33 differentially expressed proteins were found to be closely related to CVD.A thorough direct analysis of pathway enrichment and GO-annotation to these significantly changed proteins showed that down-regulation of Fibulin-1(FBLN1)may be related to regulation of transforming growth factor-beta(TGF-β)secretion,in addition to FABP3 and MB in SBP group.Furthermore,the expression levels of FBLN1 were confirmed by Western blot.Conclusions:The results of label-free quantitative proteomic study suggest that the cardioprotective effects of SBP are achieved through the preservation of energy metabolism in the heart tissue of MI rats.Compared with ECC group and BH group,SBP had better and definite efficacy.And that the TMT quantitative proteomics analysis indicated the proteins could be effective targets of SBP,and SBP could exert cardioprotective function at multiple targets through regulating multiple pathways.The results of two independent studies suggested that SBP may play a role in cardiac protection by regulating multiple targets in a variety of pathways,including PPAR signaling,energy metabolism,the redox pathway and TGF-β signaling pathway.The results also reveal the multi-effective advantages of SBP and its superiority to other treatments via mechanisms of action such as heart regeneration,angiogenesis processes,cardiac repair and remodeling,heart development and enhanced healing following myocardial infarction.The proteins found in different research strategies may be potential targets for SBP.SBP regulates the expression of multiple target proteins,and shows the characteristics of multi-component,multi-target and multi-link whole effect of traditional Chinese medicine prescription.These results suggested a theoretical basis for the future clinical research of the Chinese medicine formula in the prevention and treatment of cardiovascular diseases. |
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