| Background and ObjectiveCoronary heart disease (CHD) is a kind of global disease. The incidence of coronary heart disease is increasing with economic development, improvement of living standards, changes of dietary and the pace of life and reduction of the amount of exercise. Framingham study from the United States found that the occurrence of overwhelming majority cardiovascular events is caused by coronary heart disease in the amount of the elderly aged over65years. According to the World Health Organization (WHO), myocardial infarction will become the first reason of human death in the next two decades.Currently, diabetes mellitus (DM) is the main cause of the increase of coronary heart disease morbidity and mortality. In regard to CHD patients with DM, the prognosis of myocardial infarction and revascularization is worse than CHD patients not associated with DM, and the degree of progress in their coronary arteries is also faster than the CHD patients not associated with DM. Thus, diabetes mellitus is taking a more and more important role in the generation and development of coronary heart disease. In2001the U.S. National Cholesterol Education Program (NCEP) Adult Treatment third report (ATPⅢ), diabetes has been increased to the same high degree of risk to coronary heart disease.Until now, the damage mechanisms of diabetes to coronary heart disease are still unclear. More and more evidences suggest that a close connection exist between hyperglycemia and endothelial function and morphologic abnormalities. The high periphery glucose quantity has been discovered that it can intervene cell cycle in cultured endothelial cells, increase DNA damage, postpone the repair of endothelium and result in excessive cell death.Sirturin protein family is NAD+-dependent enzyme. Sirturin family of mammals plays a role in the fight against stress and a variety of metabolic pathways, including adipose production, gluconeogenesis, and insulin and glucose biosynthesis. Sirturin family has been considered as a potential target in the treatment of metabolic and age-related diseases, due to its functions of regulating various physiological processes. Although the exact role of Sirturin family to mammalian aging need further elucidated, more and more studies have revealed that mitochondrial protein SIRT3can adjust calorie restriction, and plays an important role in surveillance of aging process. The present studies show that high expression of SIRT3is the unique protein related to human longevity.PPAR-a (peroxisome proliferator-activated receptor) belongs to type II nuclear receptor superfamily and it is ligand-activated transcription factor, and plays a key role in lipid and glucose metabolism. PPAR-a is the main regulator molecular of mitochondrial biogenesis, which can regulate the expression of a variety of energy metabolism-related genes, and it has attracted more and more attentions in cardiovascular field.Therefore, this study is divided into two parts:1. Clinical research aimed at observing the influence of blood glucose control to the progression of coronary artery disease in CHD patients with diabetes, and having a deeply knowledge of the impact of diabetes on coronary atherosclerosis progress, and deepening our understanding of the importance of blood glucose control in CHD patients with DM.2. Experimental section is intended to explore the changes in the expression level and the roles in arterial endothelial injury of cytoplasmic full SIRT3and PPAR-α, in the basis of high glucose conditions, and observe whether the SIRT3and PPAR-α have synergism, in order to provide new ideas and research bases for revealing the mechanism of endothelial injury of arterial atherosclerosis in the pathophysiologic process in molecular level.Methods1. Clinical Research:First,60patients of the inclusion criteria will be divided into two groups, one group is30CHD patients with DM in good control of blood glucose, which is named as control group; the other one is30CHD patients with DM in bad control of blood glucose, which is named as exposed group. This two groups have good balance in age、sex、the average of course of CHD and DM、history of hypertension、clinical classification of CHD result of coronary angiography、HbA1c、LDL-C、systolic blood pressure(SBP)、diastolic blood pressure(DBP) and so on.Then, observe the glucose and lipid control situation, the frequency of angina pectoris, the readmission rates and the result of another coronary angiography three years later between the two groups.2. Experimental Research:(1) The HAECs, as a kindly gift of professor Zhang ming-xiang, were cultured carefully before divided randomly into the following groups:normal control group,11mmol/L glucose group,16.5mmol/L glucose group,22mmol/L glucose group,27.5mmol/L glucose group,33mmol/L glucose group, and38.5mmol/L glucose group;(2) Those cells were harvested after being treated for24hours;(3) The expression of full-length SIRT3, PPAR-α, iNOS, eNOS at protein level was examined by means of Western-blot. Results1. Clinical Research:(1) Through t text andχ2text, the two groups have good balance in age、sex、the average of course of CHD and DM、history of hypertension、clinical classification of CHD、result of coronary angiography、HbA1c、LDL-C、systolic blood pressure(SBP)、 diastolic blood pressure(DBP) and so on(P>0.05);(2) In three years, the blood pressure has been controlled relatively good between the two groups. So we can exclude the effect of high blood pressure on the experimental results(P>0.05);(3) The level of BMI、HbA1c、LDL-C has increased obviously in the exposed group, which compares with the control group three years later. And compares with three years before(P<0.05), the level of BMI、 HbA1c、LDL-C also increases obviously in the exposed group(P<0.05);(4) The frequency of angina pectoris has no evidently differences between two groups(P>0.05), but the readmission rates and the result of another coronary angiography have apparently differences between two groups(P<0.05).2. Experimental Research:(1) After being dealt with gradient concentration glucose for24hours, the endothelial cells grew slower as the concentration of glucose getting higher, evidenced by smaller cell density and more dead cells. Time limited, we did not meter these changes with precise methods;(2) Full-length SIRT3, PPAR-α, iNOS, eNOS protein was found in the cytoplasm of normal control group;(3) Full-length SIRT3, PPAR-α, iNOS, eNOS protein was found in the cytoplasm of each and every high-glucose groups;(4) There were no significant differences among β-actin expression levels of all the groups;(5) Compared with the normal control group, each and every group that was dealt with high glucose got a different protein expression level of full-length SIRT3, PPAR-α, iNOS, eNOS in cytoplasm, and the differences were obvious; and the difference among different glucose concentration groups are obvious; and as the glucose concentration getting higher, the expression of cytoplasm full-length SIRT3, PPAR-α, eNOS protein has gradually increased and then decreased, and expressed the largest amount in27.5mmol/L, and the expression of iNOS protein has earlier increased, and expressed the largest amount in11mmol/L, and then gradually decreased, and the expressed level is low in27.5mmol/L.Conclusion1. Clinical Research:Diabetes is one high risk factor for coronary heart disease. Nowadays diabetes has been considered as the same high degree of risk to coronary heart disease in the guideline. So the control of blood glucose regularly is crucial important in CHD patients with DM.2. Experimental Research:(1) Full-length SIRT3and PPAR-a protein presents in the cytoplasm of normal HAECs;(2) The protein expression level of full-length SIRT3and PPAR-a in cytoplasm is up-regulated under transient high glucose stimulation, and these expression levels were obviously increased and then decreased as the concentration of glucose getting higher and expressed the largest amount in27.5mmol/L, suggesting that SIRT3and PPAR-a protein is likely to take part in the mechanism against the damage caused by high concentration glucose in endothelial cells;(3) After stimulated by different concentrations of glucose, SIRT3and PPAR-a expresses a positive linear correlation with eNOS, and expresses a negative linear correlation with iNOS, and synergy exists between SIRT3and PPAR-a protein in the process to resist the stress injury, and PPAR-a may play an important role in SIRT3anti-stress injury mechanisms;(4) We build models of high glucose transient stimulation on endothelial cells that are similar with in vivo, which could be helpful for further studies about full-length SIRT3and PPAR-a, and endothelial damage caused by glucose, and the relationship between full-length SIRT3and PPAR-a and Oxidative Stress. |
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