| Objective: Based on the previous research of the research group,clinical research was carried out to explore the clinical efficacy and theoretical connotation of prescriptions,and the core of the research was to screen the core targets and enrich the main pathways and biological processes through network pharmacology to predict its mechanism of action,and to verify the specific regulatory effects and potential molecular mechanisms at the animal level,so as to broaden the TCM research ideas of ischemic heart disease and provide more references for the prevention and treatment of ischemic heart disease.Methods:Clinical study: From May 12,2023 to December 31,2023,72 patients with unstable angina pectoris who were treated in the outpatient and treatment area of the Department of Cardiology of the Affiliated Hospital of Changchun University of Chinese Medicine were collected,and were randomly divided into the control group(36 cases)and the experimental group(36 cases),of which the control group was given conventional treatment with Western medicine,and the experimental group was combined with Xuan Bi Tong Yu Formula,and the treatment cycle was 4 weeks.Seattle angina scale and other efficacy indicators,and the presence or absence of abnormalities in routine laboratory tests were used as safety indicators to study their treatment effects and safety,and the serological indicators VEGF and IL-6 were selected according to the previous research basis of the research group,and their changes before and after treatment in the two groups were observed,so as to preliminarily explore the direction of the effect of Xuan Bi Tong Yu Formula in the treatment of unstable angina.Network pharmacology: The active ingredients of Xuan Bi Tong Yu Formula were screened out through the TCMSP database,and Gene Cards,The disease targets were obtained from the OMIM and GEO databases,the intersection targets of Xuan Bi Tong Yu Formula and ischemic heart disease were screened by Venny2.1.0 software,the PPI network was constructed in the STRING database and visualized by Cytoscape software,the potential targets were uploaded to the DAD database for GO function and KEGG pathway enrichment analysis,and finally Auto Dock was used Vina(1.1.2)performed molecular docking of the active ingredient and key target to verify its interaction activity.Experimental study: Based on the results of the previous study of the research group and the network pharmacology results of this study,the protective mechanism of Xuan Bi Tong Yu Formula on myocardial ischemia model rats was further revealed.A myocardial ischemia model was constructed based on SPF SD rats,and the experimental animals were randomly divided into sham operation group,model group,high-dose Xuan Bi Tong Yu Formula high-dose group,low-dose group,Xuan Bi Tong Yu Formula high-dose + HIF-1αinhibitor group and positive control(Musk Baoxin Pill)group,a total of 6 groups,with 10 animals in each group.After successful modeling,each group was administered by gavage,and the relevant indexes were detected after 4 weeks:(1)automatic biochemical analyzer to detect serum CK-MB and LDH activities,(2)HE staining to observe the morphological changes of myocardial tissue,(3)immunohistochemistry to detect the expression of HIF-1α,CD34,VEGF and Ki-67 in myocardial tissue,(4)TUNEL method to observe myocardial cell apoptosis,(5)Western blot to detect HIF-1α in myocardial tissue,(6)q RT-PCR was used to detect the expression of IL-6,IL-1β,TNF-α,CASP3 and MMP9,the core indexes of network pharmacology in myocardial tissue.Results:Clinical Studies:(1)A total of 72 subjects were enrolled in this study,3 cases were dropped out in the control group and 4 cases were dropped out in the experimental group during the study,and a total of 65 valid research cases were finally included,33 cases in the control group and 32 cases in the experimental group.(2)There were no significant differences between the two groups in terms of gender,age,course of disease,body temperature,respiration,blood pressure,heart rate,smoking history,alcohol history,angina grade,predisposing factors,family history and other observation indicators before treatment(P>0.05).(3)The total TCM pattern score,individual score of each symptom,angina pectoris,and Seattle angina scale were significantly improved after treatment(P<0.05),and the post-treatment score of the experimental group was significantly better than that of the control group(P<0.05).(4)The efficacy of TCM syndrome and nitroglycerin reduction rate in the experimental group were significantly better than those in the control group,and the results were statistically significant(P<0.05),but although the experimental group was better than the control group in terms of angina pectoris efficacy and electrocardiogram efficacy,statistical analysis showed that there was no significant difference between the groups(P>0.05).(5)There were significant differences between the two groups after treatment with VEGF and IL-6(P<0.05),and the experimental group was significantly better than the control group after treatment,and the results were statistically significant(P<0.05).(6)After treatment,there were no obvious abnormalities in vital signs,routine laboratory examinations,etc.,and no serious adverse reactions were found in the two groups,indicating good drug safety.Network pharmacology:The database collected 106 active ingredients and 206 drug targets in Xuan Bi Tong Yu Formula,mainly including quercetin,β-sitosterol,stigmasterol,kaempferol,blue violine,etc.,and screened the top 10 core targets IL-6,AKT1,TP53,IL-1β,TNF,EGFR,JUN,MYC,CASP3,MMP9,etc.,and Xuan Bi Tong Yu Formula mediated core interaction targets,regulated HIF-signaling pathway,PI3K-Akt signaling pathway,NOD-like receptor signaling pathway,calcium signaling pathway and other pathways are involved in biological processes such as apoptosis,cell proliferation,and inflammatory response,and then play a role in protecting cardiac function.Experimental Studies:(1)The automatic biochemical analyzer showed that the amount of CK-MB and LDH released into the model group was significantly higher than that in the sham group(P<0.01).There was no significant difference in serum CK-MB and LDH levels in the high-dose and positive control drug Musk Baoxin Pill group(P>0.05),and the CK-MB and LDH contents in the high-dose Xuanbi Tongyu Fang group were significantly lower than those in the model group(P<0.01),but the myocardial injury markers in the Xuanbi Tongyu Fang+HIF-1α inhibitor group were significantly higher than those in the Xuanbi Tongyu Fang group(P<0.05).(2)HE staining showed that compared with the sham operation group,the myocardial cells in the model group were hypertrophied and deformed,irregular arrangement,increased intercellular space,increased interstitial and extravascular matrix,and showed significant improvement after treatment with Xuan Bi Tong Yu Formula and the positive control drug Musk Bao Xin Pill,and the myocardial cell space of the rats in the Xuan Bi Tong Yu Formula + HIF-1α inhibitor group became larger again,and the intercellular and perivascular extracellular matrix increased significantly.(3)IHC and Western blot showed that the expression levels of HIF-1α and VEGF in the model group were significantly increased compared with the sham operation group(P<0.01),Compared with the model group,the expression of the two groups was significantly increased in the high-dose group(P<0.01),and compared with the high-dose group of Xuan Bi Tong Yu Formula,the expression of HIF-1α and VEGF was significantly inhibited by Xuan Bi Tong Yu Formula + HIF-1α inhibitor(P<0.01).(4)IHC showed that the expression of CD34 and Ki-67 in the model group was significantly reduced compared with the sham operation group(P<0.01).Compared with the model group,the expression of Xuan Bi Tong Yu Formula was significantly increased in the Xuan Bi Tong Yu Formula group(P<0.05),and compared with the high-dose group of Xuan Bi Tong Yu Formula,the expression of Xuan Bi Tong Yu Formula + HIF-1α inhibitor could be significantly inhibited(P<0.01),while the expression of CD34 and Ki-67 in the positive control group was not significantly changed(P>0.05).(5)TUNEL assay showed that the apoptosis index of cardiomyocytes in the mo del group was significantly higher than that of the sham operation group(P<0.01),Compared with the model group,the apoptosis index of cardiomyocytes in the low-d ose and high-dose groups of Xuan Bi Tong Yu Formula was significantly reduced(P<0.01),and compared with the high-dose group of Xuan Bi Tong Yu Formula,Xua n Bi Tong Yu Formula + HIF-1α inhibitor significantly promoted cardiomyocyte apo ptosis(P<0.01),while the cardiomyocyte apoptosis index of the positive control gro up did not change significantly(P>0.05).(6)The results of qRT-PCR showed that the expressions of CASP3,MMP9,IL-6,IL-1β and TNF-α in the model group were significantly increased compared with the sham operation group(P<0.01),Compared with the model group,the m RNA co ntent of the above factors was significantly increased in the Xuan Bi Tong Yu Form ula group(P<0.01),and compared with the high-dose group of Xuan Bi Tong Yu Formula,the m RNA content of the above factors was significantly increased in the Xuan Bi Tong Yu Formula + HIF-1α inhibitor(P<0.01),while the CASP3,There was no significant change in the expression of TNF-α(P>0.05),the expression of I L-6 was significantly increased(P<0.05),the expression of IL-1β was significantly i ncreased(P<0.01),and the expression of MMP9 was significantly decreased(P<0.01).Conclusion:Xuan Bi Tong Yu Formula can improve the efficacy of TCM syndrome in UA patients,improve angina symptoms,improve the quality of life of patients,increase the rate of nitroglycerin depletion,increase serum VEGF content,reduce IL-6 content,and have good drug safety.In myocardial ischemia,Xuanbi Tongyu Formula may act on core targets such as IL-6,IL-1β,TNF,CASP3,MMP9 through effective components such as quercetin,β-sitosterol,stigmasterol,kaempferol,and blue viol,so as to positively regulate the expression of HIF-1α,mediate the HIF-1α signaling cascade,promote tissue angiogenesis,reduce inflammatory damage,and inhibit apoptosis,thereby protecting ischemic myocardium. |
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