Study On The CircRNA Clinical Relevant Mechanism Of Xuefu Zhuyu Capsule Intervention In Patients With Stable Angina Pectoris With Qi Stagnation And Blood Stasis Syndrome

Coronary heart disease has become a serious clinical and public health problem because of its high morbidity and high mortality.Stable angina pectoris is one of its important classifications.Chinese medicine refers to coronary heart disease as "chest palsy" and "heartache." It considers that Qi block and blood vessel occlusion are important pathogenesis.Qi stagnation and blood stasis syndrome is one of the common TCM syndromes.Representative prescription for stasis syndrome.Xuefu Zhuyu Capsule is the representative prescription.Although clinical treatment of coronary heart disease currently has standardized traditional Chinese and western medicine treatment and secondary prevention,there are still problems such as resistance to aspirin and clopidogrel,anxiety and fear after intervention,coronary restenosis,etc.There are long history and definite curative effect of traditional Chinese medicine treatment of coronary heart disease.The simple use of Western medicine indicators or scales cannot fully reflect the curative effect of traditional Chinese medicine.PRO scale is one of the methods of curative effect evaluation,which can be used to evaluate the curative effect of traditional Chinese medicine from the subjective feelings of patients before and after treatment.As a research hot area in recent years,circRNA not only participates in the pathogenesis of coronary heart disease,but also plays an important role in the diagnosis,treatment and prognosis of coronary heart disease.Our team also has some practical explorations in the field of circRNA.Therefore,we relied on the research project on the evaluation of the efficacy of new Chinese medicines with syndromes based on the research project of the Chinese medicine industry,and evaluated the effectiveness of Xuefu Zhuyu Capsule in intervening stable angina pectoris with qi stagnation and blood stasis syndrome based on the Chinese medicine PRO scale for qi stagnation and blood stasis syndrome.Combined with the team's previous research in the field of circRNA,the circRNA-related mechanism of Xuefu Zhuyu Capsule intervening in stable angina pectoris and qi stagnation and blood stasis syndrome was discussed.Objective1.Evaluate the effectiveness of Xuefu Zhuyu Capsule in the intervention of stable angina pectoris with qi stagnation blood stasis syndrome based on the Chinese medicine PRO scale of Qi stagnation blood stasis syndrome2.To study the mechanism of circRNA related to Xuefu Zhuyu Capsules in the intervention of stable angina pectoris with qi stagnation and blood stasis syndrome.MethodPart 1 A nested case-control study of Xue's Zhuyu Capsule based on the Chinese medicine PRO scale of Qi stagnation and blood stasis syndrome in the intervention of stable angina pectorisA retrospective nested case-control study design was adopted,and 24 patients with stable angina pectoris with blood stasis syndrome from three hospitals in the Beijing area were included according to the standard,including 12 cases in the experimental group and the control group.After collecting general information,the online randomized system of the Chinese Academy of Chinese Medical Sciences was used to randomly generate a test group and a control group at a 1:1 ratio.The experimental group was treated with Xuefu Zhuyu Capsule,and the control group was treated with Xuefu Zhuyu Capsule Simulator,all of which were 6 capsules/time,2 times/day.They were taken with warm water,the introduction period was 1 week,and the course of treatment was 7 weeks.On the day of enrollment,the Chinese medicine PRO scale?main outcome index?of the subject's qi stagnation and blood stasis syndrome was recorded on the 2,4,6,and 7 weeks of the enrollment,and on the day of enrollment,the patient's TCM symptoms and signs scale?secondary?Outcome indicators),the number and duration of angina attacks?therapeutic indicators?,blood routine,urine routine,stool routine,liver and kidney function,electrocardiogram and other indicators?safety index?,and record adverse events throughout.Epidata3.0 was used for data entry,and SPSS25.0 was used for data analysis.The normality test was performed on the measurement data.The measurement data that conformed to the normal distribution were expressed as mean±standard deviation?Mean±SD?.The paired t test was used for comparison.The parameters of the test group and the control group were compared using the independent sample t test.The count data was expressed in terms of number of cases and percentage.The Fisher's exact test?T<1 or n<40?was used.The statistical tests are both two-sided tests,the confidence interval is 95%confidence level,*P<0.05,**P<0.01,***P<0.005,****P<0.0001 to determine the difference is statistically significant,Graph Pad Prism6.0 is used for drawing.Part 2 Study on CircRNA Related Mechanism of Xuefu Zhuyu Capsule Intervention in Stable Angina Pectoris with Qi Stagnation and Blood Stasis SyndromeScreening of the first part of Xuefu Zhuyu Capsule for intervention in patients with stable angina pectoris and stagnation of blood stasis syndrome:5 patients in the experimental group and 5 patients in the control group;combining literature search and pre-scientific research results of the team,screening target circRNA;On the next day of enrollment and the next day of enrollment.4 ml of venous blood from these 10 patients in the morning fasting state was drawn to complete the separation of white blood cells,total RNA was extracted,and RNA quality inspection was completed using agarose gel electrophoresis and NanoDrop2000 spectrophotometer.Real-time quantitative polymerase chain reaction?qRT-PCR?was used to verify the changes of these 4 circRNAs before and after the first part of treatment.Gene GO function enrichment,gene KEGG signal pathway analysis.miRNA Predict,construct circRNA-miRNA-mRNA network,analyze and discuss PCR results.SPSS25.0 was used for statistical analysis of data.The comparison of various parameters before and after treatment in the group was performed by paired t test.The comparison of each parameter between the two groups was by independent sample t test.All statistical tests were performed by two-sided test,*P<0.05,**P<0.01,***<0.005,****P<0.0001 to determine the difference is statistically significant,Graph Pad Prism6.0 is used for drawing.ResultPart 1 A nested case-control study of Xue's Zhuyu Capsule based on the Chinese medicine PRO scale of Qi stagnation and blood stasis syndrome in the intervention of stable angina pectoris1.There is no significant difference in baseline comparison between the experimental group and the control group before treatment?P>0.05?,which is comparable;2.The main outcome indicators:PRO scale score reminder,after 2,4,6,7 weeks of treatment,the experimental group The total effective rate was higher than that of the control group.After 7 weeks of treatment,the improvement of the syndrome of qi stagnation and blood stasis was better than that of the control group?P=0.027?,the difference was statistically significant;P=0.023),palpitation?P=0.028?and Shantaixi?P=0.041?were significantly better than the control group,the difference was statistically significant;4.curative effect index:the number of angina attacks?P=0.006?and duration in the experimental group The time?P=0.001?improvement was significantly better than the control group,the difference was statistically significant;5.Safety indicators:basic vital signs such as body temperature,heart rate,breathing,blood pressure,urine routine There were no abnormalities in routine and electrocardiogram.There were no significant statistical differences in blood routine,liver and kidney function,and coagulation?P>0.05?.There were no adverse events in the subjects during treatment and follow-up.Part 2 Study on CircRNA Related Mechanism of Xuefu Zhuyu Capsule Intervention in Stable Angina Pectoris with Qi Stagnation and Blood Stasis Syndrome1.There is no significant difference between the experimental group and the control group before baseline treatment?P>0.05?,which is comparable;2.Screening hsa_circ₀029589,circ_Lrp6,circ_ARF3,hsa_circ₀036007 is the target circRNA of this experiment;3.Blood sample quality test results Display:RNA integrity,total quantity,purity and other quality all meet the experimental requirements,and the rating is A grade,which can be used for subsequent experiments;4.The melting curves of hsa_circ₀029589 and hsa_circ₀036007 are single peaks,and the specificity of PCR amplification is good;5.The relative expression levels of hsa_circ₀029589 and hsa_circ₀036007 were compared between the control group and the experimental group before treatment?P=0.2046,P=0.0873?,before and after the control group treatment?P=0.4517,P=0.7539?,the comparison between the control group and the experimental group after treatment?P=0.6476,P=0.1821?,there is no statistical difference;hsa_circ₀029589 before treatment in the experimental group and after treatment in the experimental group?P=0.0782?,no statistical difference;hsa_circ₀036007 test group before and after treatment group comparison?P=0.0006?,the expression of hsa_circ₀036007 in the test group after treatment was about 0.304 times before treatment,hsa_circ₀036007 was significantly down-regulated.Conclusion1.The PRO scale for Qi stagnation and blood stasis syndrome can be used as an evaluation index for the clinical efficacy of stable angina pectoris syndrome;Xuefu Zhuyu Capsule has good efficacy and safety in the intervention of stable angina pectoris with stagnation of blood stasis syndrome;2.hsa_circ₀036007 may be a potential biomarker for predicting the efficacy of Xuefu Zhuyu Capsule in intervening stable angina pectoris and qi stagnation and blood stasis syndrome,and may be combined with hsa-let-7d-5p,hsa-let-7a-5p,hsa-miR-93-5p or PI3K-Akt signaling pathway is related to play a role,the next step in the study can be expanded sample size or increased animal model cell lines related experiments to verify.