Cortex Dictamni Inhibits Formation Of Advanced Atherosclerotic Lesions In Mice And The Protective Effects On Cardiocytes

Ischemic heart disease, which is known as coronary artery disease, has become the highest mortality rate of human diseases, especially in the developing countries. Meanwhile, in recent years, the tendency of patients’ age has become younger and younger, and the incidence cases increased year by year. Therefore, much attention has been focused on the treatment of the coronary artery disease.Atherosclerosis is the initial factor of ischaemic heart disease. For the treatment of ischaemic heart disease, it is necessary to repair the damage location after myocardial ischemia, but the most important thing is to inhibit the formation of atherosclerotic lesions from the early stage. Most researchers focused on the Chinese medicine of promoting blood circulation to remove blood stasis in the field of prevention of atherosclerosis by natural medicine. But in recent years, damp and heat are also the pivotal factors of coronary artery disease on the base of the pathological study. Therefore, aimed to improve the shortcomings of previous works, systematic screenings were carried out for the related heat-clearing Chinese medicine, which could prevent the formation of atherosclerotic lesions. The results showed that Cortex Dictamni has an obvious inhibitory effect on the formation of atherosclerotic lesions and has intervention effect on the development of pathological process of atherosclerosis.PART I Cortex Dictamni inhibits formation of advanced atherosclerotic lesions in ApoE-deficient miceAim To determine whether Cortex Dictamni aqueous extract (CDAE) has any effect on the advanced lesion formation of aortic atherosclerotic lesions in ApoE-/-mice and to explore the possible mechanisms involved.Methods40female ApoE-/-mice were randomly divided into four groups (n=10): the control group, high dose CDAE group, medium dose CDAE group and low dose CDAE group. All animals were fed a high fat diet and the three CDAE groups were orally administered with CDAE at different doses (3.2mg/g,1.6mg/g,0.8mg/g) for6weeks from12to18weeks, while the control group got the same volume of distilled water. At the end of the experiment, the serum levels of lipids including total cholesterol (TC), triglyceride (TG), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C) were determined. The heart and proximal aorta were removed from the mice and then cut directly under and parallel to the aortic cusps and the upper portions were embedded in OCT compound. The samples were cut into a total of80sections (6μm) and the size of the atherosclerotic lesion in each aortic section was evaluated. In vivo experiment, MTT assay was used to view proliferation of VSMCs. Cell migration was determined by Transwell assay.Conclusions CDAE administration could obviously reduce the advanced lesion size in ApoE-/-mice. The possible mechanism of action maybe reduce the serum lipid levels, and inhibit the proliferation and migration of VSMCs.PART II Protective effects of Cortex Dictamni on hypoxia-reoxygenation injury in H9C2cardiocytesAim To investigate the protective effects of CDAE on myocardial ischemia and to study the mechanism of action further.Methods The hypoxia-reoxygenation injury model in vitro was established in H9C2cardiocytes. The protective effects of CDAE on H9C2cells were initially determined by three indexes, including cell activity by MTT method, the release amount of LDH and cTn-I. In order to analyze the relation between oxidative stress and the protective effect of CDAE, the superoxide dismutase (SOD) activity in the supernatant and quantity of malondialdehyde (MDA) of lipid metabolites were determined. To study the apoptosis of cardiocytes, the cells were detected by flow cytometry with the method of FITC V/PI. Besides, the mitochondrial membrane potential (MMP), activity of caspase-3, amount of cytochrome c and Bcl-2/Bax ratio were determined to study the signal transduction pathway for the protection of cardiocytes.Conclusions CDAE significantly protected the hypoxia-reoxygenation injured H9C2cardiocytes and the action mechanism was realized by antioxidant effect and anti-apoptotic effect. The antioxidant effect of CDAE was to increase the activities of antioxidant enzyme and decrease the production of lipid peroxidation. The anti-apoptotic effect of CDAE was mainly realized by mitochondria pathway.