| Background:With the increase in elderly people and the improvement of people's living standard, morbidity and mortality rates of coronary heart disease rises rapidly and have become the major threat to China's public health, and thus has been called "the first killer of human." With the development of modern medical technology, diagnosis and treatment of coronary heart disease have made a significant progress. With the use of revascularization techniques, such as intravenous thrombolysis, percutaneous transluminal coronary angioplasty, the mortality caused by coronary heart disease decreased significantly, but the side effect such as myocardial ischemia-reperfusion-injury(MI/RI) became an important factor to threat patient's life. Thus, it is of great significance to study on pathological mechanism and effective drugs.MI/RI refers to the myocardial blood supply disruption in a short time to restore blood supply within a certain period of time, the original occurrence of ischemic myocardium to restore blood supply in much more serious damage. The main symptoms are a series of clinical symptoms of disease progression such as reperfusion arrhythmia,no-reflow phenomenon,myocardial stunning,microcirculation, and even sudden death.The pathological process is complex, involving multiple links. In the past, the basic and clinical research for the MI/RI were mainly focus on suppression of oxygen free radicals, antagonize intracellular calcium overload and so on, but recently some people believe that MI/RI is primarily a process of inflammatory response:the process was polymorphonuclear (PMN) cells as the main body, co-participation with platelets, vascular endothelial cells, a variety of biologically active substances such as cytokine and complement, mediated by the PMN adhesion molecule and vascular endothelial cell adhesion to infiltrate myocardium and release large amounts of oxygen free radicals and enzymes, cause various forms of-damage in the level of cell,sub-cellular and genetic. Research shows that adhesion, permeability, chemotaxis, activation and migration through endothelium of PMN when MI/RI depends on a variety of adhesion molecules and cytokines, the more influential ones are tumor necrosis factorα(TNF-α), interleukin factor-1 (Interleakin-1,IL-1), interleukin-6 (Interleakin-6,IL-6),etc. These cytokines can directly inhibit myocardial contractile function, resulting in decreasing the cardiac output, precipitating cardiomyocyte apoptosis and directly and indirectly increase the expression of PMN adhesion molecules CDllb and the surface of endothelial cells and myocardial intercellular adhesion molecule-1 (ICAM-1).In recent years, Chinese medicine in prevention and treatment of cardiovascular disease has made a considerable progress.There was no concept of ischemia-reperfusion injury in ancient TCM literatures.The disease were described with many terms such as "thoracic obstruction", "heart-throb", "precordial pain with cold limbs" "prostration syndrome" according to the etiology, clinical manifestation and prognosis.The total pathogenesis can be summarized as deficiency in the origin and excess in the superficiality.Deficiency in the base includes deficiency of Qi and blood, Yin and Yang that caused by weak constitution, deficiency of kidney or blood asthenia, especially deficiency of heart Qi and Yin rooted in spleen and kidney. While excessiveness in the superficies includes Qi stagnation, blood stasis, turbid phlegm, cold coagulation, heat accumulation that caused by improper diet,emotional stress, exhaustion, especially the complicated with turbid phlegm and blood stasis to repress thoracic Yang, occlude heart collaterals, so that extend chest pain.Clinical symptoms are asthenia complicated with sthenia,reciprocal translocation.Principles of treatment are replenishing Qi and blood, regulating Yin and Yang, promoting blood circulation and resolving phlegm.Objective:Based on early study, we used rats with ischemia-reperfusion injury as animal models, from both the sides of Arrhythmia scores and myocardial pathology to observe the protective effects of DingXin Recipe and tanshinoneⅡA on MI/RI; Research the expression of inflammatory factor IL-6 in rat serum and IL-6mRNA myocardial tissue changes and the intervention of DingXin Recipe and tanshinoneⅡA.At the same time use proteomics technology to research the Prohibitin expression after MI/RI and interventional roles of DingXin Recipe and tanshinoneⅡA on the expression of Prohibitin.The aim of this study is to confirm feasibility that Chinese herbal compound DingXin Recipe and tanshinoneⅡA can treatment ischemic heart disease. To clarify the pathogenesis of MI/RI will provide some experimental basis for developping new, safety traditional Chinese medicines to prevention and treatment of ischemic heart disease.Methods:32 adult male Wistar rats were randomized into 4 equal groups: Sham operated group (SH),myocardia ischemia-reperfusion group (MI/RI), DingXin Recipe group (DXR) and TanshinoneⅡA group (Tan).All the rats were fed for 7 days for accommodation. Rats of SH, MI/RI and Tan groups were drenched with normal sodium, and rats in DXR group were drenched with DXR physic liquor. After successive intragastric administration for 7 days, the rats of MI/RI, DXR and Tan groups accepted operation, and the rats of SH group also accepted the operation except no ligation and reperfusion.The rats of Tan group were administed of Tan by tail vein injection.We made models of MI/RI according to Pharmacological Empirical Methodology. Rats were Combined with electrocardiograph and electrocardioscope to observe the occurrence of arrhythmia and made a note of arrhythmia that happened during reperfusion stage and gave a mark of ventricular arrhythmia. After the above-mentioned operation, took blood from abdominal aorta and then prepared serum. Eventually, took out of the heart, washed it in cool normal sodium and separated the left ventricle, then stored it in refrigerator with the temperature of-80℃.Arrhythmia induced by myocardial ischemia-reperfusion injury mainly was ventricular arrhythmias, including ventri cular extrasystole, ventricular tachycardia and ventricular fibrillation, so we gave a mark of ventricular arrhythmia based on the method of Cutis.The muscular tissues that fixed by formaldehyde was paraffin imbedded and made into slices.Used HE dyeing to observe the morphous changes of myocardium in each group and the protective effects of DingXin Recipe and Tanshinone II A.We detected IL-6 in serum from abdominal aorta by ABC-ELISA, and observed the effect of DingXin Recipe and TanshinoneⅡA on expression of serum IL-6.Total RNA was extracted from partial myocardium and reversely transcribed into cDNA.The Real Time RT-PCR was used to examine the level of IL-6 mRNA. Then we observed the effect of DingXin Recipe and TanshinoneⅡA on the expression of IL-6 mRNA.Total protein was extracted from partial myocardium for Western blot. The expression of PHB was determined by immunohistochemical method and western blotting method, and observed the effect of DingXin Recipe and TanshinoneⅡA on expression of PHB in rats with MI/RI.Arrhythmia score were multiple sets of grading data, we used Kruskal-Wallis for analysis, experimental data indicated with average ranks.The remaining data were completely random design of the measurement data, we adopted One-Way ANOVA and multiple comparing for analysis, homogeneity of variance with Bonferroni method and missing variance with Dunnett's T3 method.The experimental data were expressed mean±standard deviation (x±s), and the correlation between two variables was analyzed by Pearson correlation analysis. P <0.05 indicated statistical significance. SPSS13.0 software was used for statistical analysis.Results:1. The effect of DingXin Recipe and TanshinoneⅡA on arrhythmia induced by MI/RI in rats. By statistics analysis in total, the score of arrhythmia had significant difference (x2=22.360, P=0.000).The score of arrhythmia in MI/RI group was the highest, which confirmed that models were credible,and DingXin Recipe and TanshinoneⅡA showed a great effect on the prevention against arrhythmia induced by myocardial ischemia-reperfusion injury according to the score of DXR and Tan groups.The incidence of premature ventricualr contraction in SH group was low and without other kinds of arrhythmia. In MI/RI group, arrhythmia occurred in 8 rats, and arrhythmia started to appear when coronary artery was perfused again for 1 minute, mainly including premature ventricualr contraction, ventricular tachycardia and ventricular fibrillation, with a few atrial ventricular blocks, and 1 rat in this group died of ventricular fibrillation. The score of ventricular arrhythmia in MI/RI group was significantly higher than that in SH group. In DXR group, arrhythmia also occurred in 8 rats, mainly premature ventricualr contraction and ventricular tachycardia, and the score of ventricular arrhythmia was lower compared with MI/RI group. In Tan group, arrhythmias were also mainly premature ventricualr contraction and ventricular tachycardia, and there was only premature ventricualr contraction in 1 rat, the score of ventricular arrhythmia was also lower compared with MI/RI group.2. The effect of DingXin Recipe and TanshinoneⅡA on the structure of myocardium in rats with MI/RI. The observation of HE dyeing from light well-arranged myocardial cells, well-distributed HE dyeing and integrated cell membrane. While in MI/RI group, the myocardial cells were chaotic, HE dyeing was uneven, cells in some parts of the heart were cloudy swelling, the cardiac muscle fibers was unclear or disappeared. In DXR and Tan group, some myocardial cells developed degeneration and necrosis, but it was not severe. HE dyeing was relatively even, cell hydropic degeneration was lightened and necrosis scope was small.3. The effect of DingXin Recipe and TanshinoneⅡA on the expression of IL-6 in serum. The content of IL-6 had significant difference by statistics (F=27.783, P= 0.000). we found that compared with SH group, the content of IL-6 in MI/RI group increased obviously, and difference had statistics significance (P=0.000); In DXR and tan group,the content of IL-6 was lower than MI/RI group (P=0.001,0.004) and higher than the SH group (P=0.001,0.000); but compared DXR with Tan, difference had no statistics significance (P=1.000)4. The effect of DingXin Recipe and TanshinoneⅡA on the expression of IL-6mRNA in myocardial tissue. The content of IL-6mRNA had significant difference by statistics (F= 22.499,P=0.000).We found that compared with SH group, the content of IL-6mRNA in MI/RI group increased obviously, and difference had statistics significance (P=0.000); In DXR and tan group,the content of IL-6 was lower than MI/RI group (P=0.002,0.010) and higher than the SH group (P= 0.002,0.000); but compared DXR with Tan, difference had no statistics significance. (P=1.000)5. The effect of DingXin Recipe and TanshinoneⅡA on the expression of PHB in myocardial tissue. The content of PHB had significant difference by statistics (F =96.215,207.061,P=0.000). We found that compared with SH group, the content of PHB in MI/RI group increased obviously, and difference had statistics significance (P=0.000); due to the role of DXR and Tan the content of PHB higher than MI/RI group obviously, and difference had statistics significance (P=0.000); but compared DXR with Tan, difference had no statistics significance (P=1.000,0.381)Conclusions:1. Arrhythmia can easily be induced by MI/RI in rats, and it can effectively be prevented by DingXin Recipe and TanshinoneⅡA.2. DingXin Recipe and TanshinoneⅡA showed great protection against myocardium histological structure destruction in rats with MI/RI.3. The contents of serum IL-6 was obviously increased after MI/RI, but degraded by DingXin Recipe and TanshinoneⅡA, and the damage induced by MI/RI were lessened.4. The expression of myocardium IL-6mRNA was obviously increased after MI/RI, but degraded by DingXin Recipe and TanshinoneⅡA.5. The expression of myocardium PHB was obviously increased after MI/RI, DingXin Recipe and TanshinoneⅡA show protection probably through increased the expression of PHB.
|
PDF Full Text Request