| ZiShen HuoXue recipe is Master Liu Zhiming’s empirical prescription of treating CHD. Myocardial ischemia of coronary artery disease is in the range of chest discomfort of Chinese Medicine, Mr. Liu has proposed unique insight of chest discomfort pathogenesis, and condensed three treating methods "nourishing kidney, dredging yang, and eliminating pathogen". With many years of treatment based on syndrome differentiation, effect is as great as drumming. The recipe is composed by a mulberry compatibility, Trichosanthes, Radix, scallion white, mistletoe, Panax and other drugs together, and is applied for national patent (patent NO.201110268875.6) Pre-clinical studies demonstrate the safety and effectiveness, pharmacological studies show that the prescript of protective effect of myocardial ischemia may be associated with anti-inflammation and plaque stabilization.Coronary heart disease makes great threat to human life and health. China has more than 70 million people die of coronary heart disease each year, the morbidity and mortality continues to rise, which brings a tremendous burden on people’s lives and national finances. Western medicine treatment is mainly to relieve myocardial ischemia and hypoxia, and improve the prognosis of patients with coronary heart disease. In severe cases, subjects have to take revascularization surgery, percutaneous coronary intervention (PCI) and/or coronary artery bypass grafting (CABG), et al. But long-term treatment sometimes always cause side effects., and after surgery, it still took a long time of anti-platelet aggregation drugs, there again tubes may be narrow, there are several contraindications and possible complications. Chinese medicine is suitable for long-term treatment. Chinese medicine treatment of coronary heart disease starts from the present theory of governance, the overall treatment, with fewer side effects, treating the symptoms, low cost and other advantages, so the search for effective prevention and treatment of coronary heart disease in Chinese medicine safety and reduce the recurrence is an important topic of current research, especially in mining of old and famous TCM doctors’treatment of coronary heart disease experience has important clinical significance and value. This study is done on the platform of Beijing Natural Science Foundation (Project Name:proteomics and molecular mechanism study of old and famous TCM doctor’s ZiShen HuoXue recipe on rat ischemic myocardium, NO:7122153), detailed research are as following.Experimental study:Objective:From rat electrocardiogram, myocardial pathological changes and vasomotor factors, regulatory factors point of view channel information to verify the efficacy of ZiShen HuoXue recipe on kidney and blood stasis of coronary heart disease, explore ZiShen HuoXue recipe treat coronary heart disease by screening for protein expression difference molecular mechanisms of ischemia. State Medical Masters experience for the treatment of CHD, and provide new ideas for the scientific basis for the prevention and treatment of coronary heart disease.Methods:Using coronary artery disease with kidney and blood stasis rats model as research subject which produced by coronary artery ligation combined TCM preparation of syndrome preparation, using two-dimensional gel electrophoresis and tandem quadrupole time of flight mass spectrometry to screen the disease syndrome and normal group the difference between protein expression, and observing ZiShen HuoXue recipe on rats with myocardial difference protein expression; while the use of Western blotting is applied for detection of apoptosis associated factor of heat shock protein 27, mitogen-activated protein kinase p38 expression, enzyme-linked immunosorbent assay is applied for plasma plaque formation rats factor nitric oxide, endothelin and inflammatory cytokine IL-18 levels.Result1 Combining disease and syndrome model preparationComparing with normal groups and kidney deficiency and blood stasis group and combination of disease and syndrome group before modeling, rat temperature drops and breathing rate increases in kidney deficiency and blood stasis group, combination of disease and syndrome group (P<0.05). Comparing with normal group, rat weight decreases or grows slowly, and whole blood viscosity and casson viscosity increases (P<0.05, P<0.01). There is statistical significance in whole blood viscosity and electrocardiogram of the two groups (P<0.05).2 ECG and pathological comparisonZiShen HuoXue recipe can prevent the Q wave from ischemia in rats with kidney deficiency and blood stasis model. Under the microscope, the model rats’ hearts shows that:myocardial fibers swell and deform, the distance between muscle fibers broaden, tissues are loose and have edema syndrome, blood vessels in myocardial interstitial are dilated, more inflammatory cells are infiltrated. Myocardial cells markedly swell or degenerate or have necrosis phenomenon, nuclear dissolved, and part of the nuclear disappeared, much collagen fibrous tissue became hyperplasia. Endocardial blood vessels swell and degenerate, also a few inflammatory cells infiltrate. Epicardial tissue’s structures are destroyed and get hyperplasia, vessels swell or degenerate or have necrosis, many inflammatory cells infiltrate and collagen deposite. ZiShen HuoXue recipe can reduce myocardial pathology damage in the model rats.3 Testing on plaque factors and inflammatory factorsZiShen HuoXue recipe could decrease plasma endothelin and interleukin-18 levels in blood circulation, and increase levels of nitric oxide. Compared with the blank control group, model group, plasma endothelin, interleukin-18 are significantly increased, while the concentration of nitric oxide concentration decreased significantly, there are significant differences (P<0.01). Comparing to the model group, the plasma endothelin concentration of treatment groups are reduced to varying degrees, in high dose and middle dose groups, they are also significantly different comparing to the model group (P<0.01). Comparing with the model group, plasma interleukin-18 concentrations in different dose are lower, the high dose had significant difference (P<0.01), the low-dose group are statistically significant (P<0.05). Comparing with model control group, different doses of oral ZiShen HuoXue recipe groups show different increases in plasma concentration of nitric oxide, high-dose is significant difference (P<0.01), low and middle doses are significantly different (P <0.05). Comparison of different dose groups, plasma endothelin concentrations decreased with increasing concentration of administration, there are significant differences (P<0.01) between the two groups; IL-18 are decreased while the dosage increased, middle and low dose groups have significant difference to high dose group (P<0.01); nitric oxide of high dose group have statistically significant (P<0.05) with the dose group.4 Detection of apoptosis-related factorsContrast to control group, the expression of heat shock 27 protein in model group was decreased(P<0.01); and mitogen-activated protein kinase p38 expression is increased (P<0.01). Comparing with the model group, different dose groups of heat shock protein 27 gray scale values have increased to some extent; treatment groups mitogen-activated protein kinase p38 are reduced (P<0.05). These show myocardial ischemia could make the expression of heat shock protein 27 in myocardial tissue decreased, mitogen-activated protein kinase p38 expression increased, while ZiShen HuoXue recipe could elevate heat shock protein 27 in myocardial tissue content, and reduce mitogen expression of p38-activated protein kinase.5 Screening of differentially expressed proteinsComparing with the control group, model group found 66 differentially expressed proteins; compared with the control group and the model group, each dose group of ZiShen HuoXue recipe have different target protein, there are lactate dehydrogenase B chain, carnitine palmitoyl transferase -2, isocitrate dehydrogenase, ATP synthase a subunit and myosin-6.Conclusion1 There is no obvious difference between the model of combining the myocardial ischemic coronary disease with the syndrome of kidney deficiency and blood stasis which is established by surgical method and non-disease of kidney deficiency and blood stasis syndrome model. They all meet the TCM clinical diagnosis criteria of kidney deficiency and blood stasis syndrome.2 The appropriate amount of ZiShen HuoXue recipe could improve the ECG of myocardial ischemia model rats. The model rats with kidney deficiency and blood stasis made by the combined method have various degrees of damage in its myocardial. It proves it’s a successful method in building the model, also there is a close relationship between kidney deficiency and blood stasis with myocardial. ZiShen HuoXue recipe can reduce myocardial damage in the model rats with ischemia, and regulate the function, thus improve the effect of this disease.3 ZiShen HuoXue recipe could reduce synthesis and increase release of endothelin synthesis, inhibition of the inflammatory response mediated by IL-18, promote the synthesis of nitric oxide, through ways of anti-inflammation, stabilize plaque, dilate blood vessels, reduce the degree of myocardial ischemia, so as to improve kidney Blood Syndrome of coronary heart disease treatment.4 ZiShen HuoXue recipe could promote the expressions of heat shock protein 27, suppress mitogen-activated protein kinase p38 expression in myocardial tissue, inhibition of cardiomyocyte apoptosis, reduce inflammation, regulate stress factors and cellular channel factor, thus play active role in the treatment of ischemic heart disease.5 The model rats proteome changed, and these proteomic spots may be the molecular basis for the development of myocardial ischemia in coronary heart disease; ZiShen HuoXue recipe regulated energy metabolism and protected the cytoskeleton in rats with kidney deficiency and blood stasis syndrome, which may be the mechanism for the treatment of coronary heart disease.In summary, the use of ZiShen HuoXue recipe on coronary artery disease and kidney and blood stasis rats, can effectively inhibit the inflammatory response, reducing myocardial apoptosis, stable plaque, improve myocardial energy metabolism, protect the cytoskeleton, significantly relieve myocardial ischemia. ZiShen HuoXue recipe is the treatment of kidney and blood stasis of coronary heart disease and effective way, the present study was to further explore the mechanism of action Pharmacological indicate the direction and laid a good foundation. |
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