Effects Of Two Kinds Of Equisetum Extract On Smooth Muscle Cells In Early Atherosclerotic Rats

Objective: Early experiments have verified that traditional chinese medicine Equisetum can regulate the metabolic disorder of blood-lipid, protect aorta endothelial cell and inhibit the development of atherosclerosis. The present Experiment is to extract the effective part of Equisetum and study the effects of the two kinds of extract on the blood-fat, proliferation, apoptosis, related genes expression, morphological changes of smooth muscle cells in early Atherosclerotic rats and its possible mechanisms.Methods1 Extract of the effective part of Equisetum The Equisetum was cut into pieces and put into water. Heat it to the boil for three hours. Extract the liquid with n-butano. The n-butano extract and the rest of extract were heated and concentrated to 1.25g/ml separately. The concentrate which was extracted by n-butano named extractⅠ, the rest named extractⅡ. Both the concentrates were stored in a freezer.2 Animal experimentFifty male mature Sprague-Dawley rats were divided randomly into five groups: normal group, model group, positive control group, extractⅠgroup and extractⅡgroup. The normal group were fed with standard chow diet and four other groups with a high fat diet. In these high fat diet groups, positive control group were fed with Gemfibrozil of 0.16 g /kg·d, extractⅠgroup were fed with n-butano extract of 12.5g/kg·d, extractⅡgroup with the rest of n-butano extract of 12.5g/kg·d.3 Detection of indicesThe plasma levels of triglyceride(TG), total cholesterol(TC), low density lipoprotein(LDL), very low density lipoprotein (VLDL) and high density lipoprotein (HDL) were measured after nine weeks. The rate of apoptosis, proliferation cycle, proliferation index and the expression of Fas, mutant p53, Bcl-2 and Caspase-3 were detected by the way of flow cytometry. Light microscopy and transmission electron microscopy were adopted to assess the degree of morphological changes of smooth muscle cells.Results1 TC, LDL in model group were higher than normal group(P<0.01). ExtractⅠgroup and extractⅡgroup could decrease the levels of TG, VLDL(P<0.01). ExtractⅡgroup could increase the level of HDL(P<0.01).2 The proliferation index in model group was higher than normal group(P<0.01). Proliferation index (PI) in extractⅠgroup and extractⅡgroup were lower (P<0.01) than model group (P< 0.01).3 The expression of mutant p53 and Caspase-3 in model group were higher than normal group(P<0.01), Bcl-2 was lower (P<0.01). Fas (P<0.01), Caspase-3(P<0.01) in extractⅠgroup and extractⅡgroup was higher than model group. Mutant p53 and Bcl-2 were lower than model group(P<0.01). The expression of mutant p53 in extractⅠgroup was lower than in extractⅡgroup (P<0.01). The apoptosis rate of smooth muscle cells in model group were higher than normal group(P<0.01). The rate of apoptosis in extractⅠgroup and extractⅡgroup was higher than model group (P<0.01).4 Morphological changes in model group were obvious. The quantity of smooth muscle cells increased and their array were disorder. Smooth muscle cells were paramorphia: nuclei, organelle and the body of cells were bigger, cell matrix was fat. Lipid was easily found in cytoplasma .These changes were less in both extractⅠgroup and extractⅡgroup.Conclusion: The two kinds of extract of Equisetum can decrease the level of TG, VLDL to regulate the metabolic disorder of blood-lipid. They can both regulate cell cycle to prohibit the proliferation of smooth muscle cells and regulate the expression of genes related to apoptosis of smooth muscle cells to promote the apoptosis of proliferating smooth muscle cells. As a result, two kinds of extract of Equisetum both have the effects of inhibiting development of experimental atherosclerosis. The effects in extractⅠgroup are more obvious than extractⅡgroup .