| Objective:Based on the efficacy study,the protective effects of Dalbergia odorifera and its substitute on myocardial ischemia in rats with Qi stagnation and blood stasis syndrome and its possible mechanism were studied.To evaluate the pharmacological equivalence of Dalbergia odorifera and Dalbergia cochinensis in myocardial protection to provide new experimental data and research basis for the replacement application of Dalbergia odorifera and Dalbergia cochinensis.PartⅠDynamic observation of TCM symptoms and biological indexes in rats with Qi stagnation and blood stasis syndrome prepared by adrenaline combined with water-ice bathMethods:Forty male SD rats were randomly divided into 5 groups,namely the normal group and the observation group of Qi stagnation and blood stasis syndrome at different time points（12 h,24 h,48 h,96 h）.In the experiment,a rat model of Qi stagnation and blood stasis syndrome was built via subcutaneous injection of epinephrine hydrochloride（0.45g/kg/time,2 times/d）combined with ice water bath（10min/time/d）.The changes of TCM symptoms,hemodynamics,hemorheology,blood lipids and other biological indexes of rats were dynamically observed 12,24,48,and 96 h after modeling.The damage of blood vessels and myocardium in rats 24 h after modeling was further detected.Results:Compared with the normal group,the rats at 12 h,24 h,48 h,and 96 h after modeling all showed obvious TCM symptoms,the syndrome score was significantly increased,and the R value of tongue,foot,and tail Significant decline.At the same time,the rats’hemodynamics were impaired,and the ECG parameters were obviously abnormal.Moreover,the blood rheology became worse.The blood rheology indexes of the 12 h group and the 24 h group were significantly increased,and most of the blood rheology indexes were significantly increased at 48 h and 96 h.Blood lipids were abnormally increased,of which LDL-C level in the 12 h group and 24 h group were remarkably improvement,and TC,HDL-C and LDL-C levels in the 96 h group enhanced markedly.Comparison of the vascular endothelial injury,myocardial injury and pathological changes in the serum of the normal group and the 24 h of Qi stagnation and blood stasis syndrome group showed that the serum NO content of the Qi stagnation and blood stasis syndrome rats was significantly reduced,and ET-1content increased significantly,serum enzymes AST,LDH,CK-MB levels were abnormally increased.HE staining results of the abdominal aorta and myocardium of the rats in the 24h group also showed that the blood vessels and myocardium were obviously damaged.PartⅡStudy on the Protective Effect of Dalbergia Odorifera and Its Alternative Medicines on Myocardial Ischemia Rats with Qi Stagnation and Blood StasisMethods:Healthy male SD rats with normal ECG were got into 11 groups at random,which were the normal group（n=8）,the sham operation group（n=8）,the ligation group（n=10）,and the blood stasis group（n=8）.The compound model group（n=10）,the Danshen group（0.36g/kg,n=10）,the propranolol group（0.024 g/kg,n=10）,low-dose odoriferous group（1.5 g/kg,n=10）,Dalbergia high-dose group（3 g/kg,n=10）,Dalbergia cochinensis low-dose group（1.5 g/kg,n=10）,Dalbergia cochinensis high-dose group（3 g/kg,n=10）.Each group was given corresponding drugs or 0.5%CMC-Na solvent orally for 11 consecutive days.Except for no treatment in the normal group,the blood stasis group established a Qi stagnation and blood stasis syndrome model,the sham operation group only threaded the coronary artery without ligation,while the ligation group established a myocardial ischemia model,and the model of Qi stagnation and blood stasis syndrome together with myocardial ischemia was prepared in the compound model group and each administration group.Half an hour after 11d administration,the color changes of the tongue,foot and tail of rats were observed as evaluation indexes of TCM symptoms.The hemodynamic parameters were measured with Power Lab electrophysiological recorder.Hemorheological indicators were detected by hemorheological apparatus and corresponding kit.Serum lipid index and myocardial enzyme index were tested by biochemical analysis instrument.Vascular endothelial function was detected by ELISA.TTC staining and HE staining were used to evaluate myocardial infarction area and histopathological changes in rats.The energy metabolism kit was used to detect glucose,pyruvate,lactic acid and ATPase.Result:Observing the appearance and symptoms of the rats,it was found that the weight of the compound model group was significantly reduced,and the tongue,feet,and tail appeared dark and purple.Compared with the compound model group,Dalbergia odorifera high dose significantly relieved weight loss（P<0.05,P<0.01）.The R value of the tongue surface of Dalbergia odorifera high and low dose groups was visibly rise（P<0.05,P<0.01）.The R value and G value of the sole of the Dalbergia odorifera high-dose group and the Dalbergia cochinensis high-dose group were significantly increased（P<0.05,P<0.01）,and the R value of the Dalbergia cochinensis low-dose group was significantly increased（P<0.05）.The G value and B value of the tail of the Dalbergia cochinensis high-dose group were significantly increased（P<0.01）.The determined results of hemodynamic measurement are expressed that compared with the sham operation group,SBP,DBP,MBP,MAP,LVSP,±dp/dt max,and mean blood flow of the composite model group were significantly reduced（P<0.05）,LVEDP increased but not significantly（P>0.05）.Dalbergia high-dose group can significantly increase arterial systolic blood pressure,diastolic blood pressure,mean blood pressure,left ventricular systolic blood pressure,+dp/dtmax,mean blood flow（P<0.05,P<0.01）.Low dose of Dalbergia odorifera+dp/dtmax,mean blood flow increased significantly（P<0.05）,and low dose of Dalbergia cochinchinensis left ventricular systolic blood pressure and mean blood flow increased significantly（P<0.05）.The results of electrocardiogram parameters revealed that compared with the sham operation group,the QRS duration and QT interval of the compound model group were distinctly prolonged,the R wave amplitude was visibly decreased,the ST value was clearly increased,and the heart rate was evidently slowed down（P<0.05,P<0.01）.The Q-T interval of the high dose of Dalbergia odorifera in the administration group was noticeably shortened（P<0.05）.The results of hemorheology showed that,compared with the normal group,the high-cut,mid-cut,and low-cut whole blood viscosity,HCT,erythrocyte electrophoresis time,RBC,erythrocyte aggregation index,erythrocyte aggregation coefficient,and hemoglobin concentration,FIB,flow resistance and Casson viscosity,and Casson stress of the compound model group were all evidently increased（P<0.05,P<0.01）.Compared with the compound model group,the low dose of Dalbergia odorifera in the administration group markedly reduced high-cut,mid-cut,and low-cut whole blood viscosity,HCT,red blood cell electrophoresis time,RBC,and hemoglobin concentration,and reduced high-cut,mid-cut,and low-cut flow resistance,Casson viscosity and Casson stress（P<0.01,P<0.05）;high-dose odorifera group significantly reduced high-cut whole blood viscosity and Casson stress（P<0.05）;Dalbergia cochinensis low-dose group can markedly reduce mid-cut whole blood viscosity,shorten red blood cell electrophoresis time,reduce high-cut,mid-cut flow resistance and Carson stress（P<0.05）;Dalbergia cochinensis high-dose group significantly reduced high-cut,mid-cut whole blood viscosity,and lower mid-cut Flow resistance and Carson stress（P<0.01,P<0.05）.After detecting the platelet aggregation index,it was found that compared with the normal group,the TXB2 content of the composite model group was obviously rise（P<0.01）.The TXB2 content of the high and low doses of Dalbergia odorifera in the administration group was obviously reduced（P<0.01,P<0.05）.The results of coagulation and fibrinolysis system index showed that the PAI-1 level of the compound model group was clearly lower than that of the normal group,and the t-PA level was evidently higher in the Dalbergia odorifera administration group and the high dose of Dalbergia cochinensis（P<0.01,P<0.05）.The detection results of vascular endothelial injury indicators showed that compared with the normal group and the sham operation group,the NO level of the compound model group was distinctly lower.The NO levels in the high-dose Dalbergia odorifera high-dose group and Dalbergia cochinchinensis high-dose group increased markedly（P<0.01,P<0.05）.Detection of myocardial enzyme indicators found that compared with the normal group and the sham operation group,the levels of AST,LDH,and CK-MB in the composite model group were markedly increased（P<0.01,P<0.05）.The high dose of Dalbergia odorifera in the administration group and the high and low dose of Dalbergia cochinensis markedly reduced the levels of LDH and CK-MB（P<0.01,P<0.05）.The results of HE staining showed that the myocardial pathological damage of the high and low doses of Dalbergia odorifera and the high and low dose of Dalbergia cochinensis was visibly improved,and the massive accumulation of inflammatory cells was significantly reduced.Part Ⅲ Study on Mechanism of Dalbergia Odorifera on Myocardial Ischemia Rats with Qi Stagnation and Blood Stasis Syndrome Methods:SD rats were randomly divided into 5 groups:sham operation group,compound model group,Danshen tablet group,Dalbergia odorifera high-dose group,and Dalbergia odorifera low-dose group.The experimental method used subcutaneous injection of epinephrine hydrochloride plus ice bath combined with ligation of the left anterior descending coronary artery to prepare a myocardial ischemia model with Qi stagnation and blood stasis.Rats in each group were given 0.5%CMC-Na solvent or corresponding drugs by intragastric administration for 11 consecutive days.The changes of blood lipids were evaluated by detecting the levels of serum TC,TG,HDL-C and LDL-C.Observe changes in glucose metabolism by detecting Glu,PA,LA content and ATPase activity.Detect GULT4,PFK2,PDH,LDHA,HIF-1α,CD36,ACC,CPT1,AMPK,LKB1,SIRT1,PGC-1αm RNA expression by RT-PCR to analyze the influence of fatty acid metabolism,glucose metabolism,and key enzymes of the tricarboxylic acid cycle.Result:（1）Compared with the compound model group,the low-dose Dalbergia odorifera group can significantly reduce the levels of TG and LDL-C（P<0.05）,and the high-dose Dalbergia odorifera group can significantly reduce the levels of TG and LDL-C and increase the level of HDL-C（P<0.01,P<0.05）.（2）Compared with the sham operation group,the LA content of the compound model group was significantly increased（P<0.05）,and the high and low doses of Dalbergia odorifera could significantly reduce the Glu content and LA content.（3）There was no significant difference in Na+K+-ATPase activity,Ca2+Mg2+-ATPase activity,and Ca2+-ATPase activity in each group.（4）PCR results showed that the low-dose Dalbergia odorifera group could significantly increase the expression levels of PFK-2 and CPT-1 m RNA,and reduce the expression level of LKB1 m RNA;the high-dose Dalbergia odorifera group could significantly increase GULT-4,PFK-2,PDH,CD36,ACC,CPT-1,LKB1and PGC-1αm RNA（P<0.01）.Conclusion:On the basis of previous studies on Qi stagnation and blood stasis model,the rat model of Qi stagnation and blood stasis syndrome can be successfully reproduced by subcutaneous injection of 0.9 mg/kg/d epinephrine hydrochloride,twice a day,soaking in 0～2℃frozen water for 10 minutes,and continuous modeling for 10 days.The established model of blood stasis syndrome is an animal model that is more in line with the symptoms of traditional Chinese medicine and the pathophysiological characteristics of Qi stagnation and blood stasis syndrome.The duration of blood stasis syndrome is also longer than the commonly used acute blood stasis model,and the most obvious period of Qi stagnation and blood stasis syndrome is found within24 h after modeling through dynamic observation and comparison.On the basis of this model,combined with coronary artery ligation,a myocardial ischemia model of Qi stagnation and blood stasis syndrome was established to study the efficacy of Dalbergia odorifera and its alternative medicine.The results confirmed that Dalbergia odorifera and its alternative medicinal material can protect Qi stagnation and blood stasis by improving blood stasis symptoms,abnormal hemorheology,hemodynamic disorders,dyslipidemia,myocardial damage,endothelial damage,and energy metabolism disorders.Syndrome myocardial ischemia in rats,which provides a certain experimental basis and theoretical basis for the use of Dalbergia cochinensis instead of Dalbergia odorifera to treat chest pain and heartache.In addition,studies have found that Dalbergia odorifera can improve the energy metabolism disorder of myocardial ischemia rats with Qi stagnation and blood stasis syndrome by promoting fatty acid oxidation,glucose oxidation and tricarboxylic acid circulation. |
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