Experimental Study Of Mechanism Of Diabetic Cardiomyopathy And Cardiac Protective Effect From Energy Metabolism Intervention

PartⅠModel construction and effect of PPARα-FFA signal pathway in STZ–induced diabetic cardiomyopathy ratsObjective To construct ideal diabetic cardiomyopathy model in streptozotocin (STZ) induced type 2 diabetic rats,and investigate the effect of peroxisome proliferator -activated receptor alpha(PPARα)and free fatty acids (FFA) signals transduction pathway in the development of diabetic cardiomyopathy through detecting cardiac expression changes of PPARαand metabolic alteration of FFA.Methods 65 female Wistar rats were randomized into control group (CON, n=8), feeding with general diet all the time, and experimental group ( n=57), which were fed with high glucose and hyperlipid diet for 4 weeks, to induce insulin resistance. Then diabetes mellitus was induced by a single injection of STZ (35 mg/kg; i.p.), Hyperglycemia (≥16.7mmol/L) was confirmed 48h later.The diabetic rats were fed with high glucose and hyperlipid diet for 16 weeks continually and 8 of them were randomized into diabetic cardiomyopathy group(DCM), Hemodynamic characterization was determined by echocardiography,the myocardial pathologic and ultrastructure changes was observed by microscope and electron microscope. The collagen volume fraction(CVF, VG staining)and expression of types I, III collagen and PPARαwere quantified by digital image analysis. Levels of blood glucose, insulin, total cholesterol, triacylglycerol and FFA were determined using biochemical or RIA methods.Results (1) Hyperglycemia was displayed in 40 STZ-induced rats followed with hyperinsulinemia, 2 rats of DCM group were died at the end of experiment. Diastolic function deteriorated in DCM-group(n=6) exhibited low E wave to A wave ratio (E/A) (DCM, 0.83±0.21 versus CON, 1.28±0.18, P<0.01)and shortened isovolumic relaxation time (IVRT)(DCM, 29.13±7.10 versus CON, 45.06±5.03, P<0.01). Histopathology revealed ventricular remodeling in STZ–DCM heart. The HW/BW ratio, ColⅠ/ ColⅢratio in DCM group (4.42±0.46, 15.49±0.87, respectively) is higher than that in CON group(3.54±0.34, 6.51±0.96, P<0.01), the CVF of LV was significantly higher in DCM (2.21±0.56) than CON (1.61±0.36, P<0.01) rats. The ultrastructure of DCM heart presented focal degeneration and disarranged order of myofilament, increased interstitial collagen and glycogen granule deposite, etc.(2) Systemic and cardiac metabolism changes: Compared with CON group, there were elevated plasma concentrations of glucose, insulin, Chol, Tg and FFA in DCM group. This was paralleled by abundant PPARαexpression in DCM-group (65.34±3.67) versus CON-group (48.69±4.88, P<0.01).Conclusion STZ-induced insulin-resistant type 2 diabetic and diabetic cardiomyopathy rats model were established successfully. The expression alterations of PPARαand metabolism changes of FFA involve in the abnormality of myocardial metabolism and myocardial remodeling, and possibly play an important role in the development of diabetic cardiomyopathy.PartⅡPPARαagonist fenofibrate inhibits myocardial inflammation and remodeling in STZ-induced diabetic cardiomyopathy ratsObjective To construct diabetic cardiomyopathy(DCM)model in streptozotocin (STZ) induced type 2 diabetic rats,and investigate the effect of peroxisome proliferator-activated receptor alpha(PPARα)agonist fenofibrate on myocardial inflammatory factors and myocardial remodeling.Methods Control rats were fed with common diet. Type 2 diabetes rats was induced by high energy diet, then treated with lower dose of STZ and continuous high glucose and hyperlipid diet. The model rats were randomized into three groups : diabetic cardiomyopathy (DCM) group, intervention group treated with low dose fenofibrate(LFIB, 4 mg·kg-1·day-1) and intervention group treated with high dose fenofibrate(HFIB, 40 mg·kg-1·day-1)for 16 weeks, respectively, with 8 rats in each group. Hemodynamic characterization was determined by echocardiography, the myocardial ultrastructure changes was observed by electron microscope, and the plasma biochemical indicators,heart weight index,typeⅠandⅢcollagens, collagen volume fraction, myocardial cytokine gene and protein expression (TNF-α, IL-10, PPARα) were detected respectively by biochemical methods, semi-quantity RT-PCR and immunohistochemistry at the end of experiment.Results (1) The decreased left ventricular systolic and diastolic function, myocardial remodeling were manifested in all diabetic rats, and these abnormalities were more significant in DCM group. Fenofibrate intervention groups with two different doses significantly improved the cardiac function compared with DCM group(P<0.01). But the two intervention groups had no significant difference(P>0.05). (2) Compared with CON group, there were significant elevated plasma concentrations of glucose, insulin, Chol, Tg and FFA in DCM group. Fenofiberate treatment had no effect on the level of blood glucose and insulin, but significantly decreased the concentration of blood Chol, Tg and FFA. The level of FFA in HFIB group was significantly lower than that in LFIB group(187±13.6 vs 168±17.3μmol/L,P<0.05). (3) The expression of TNF-α, IL-10 and PPARαwas increased significantly,the myocardial impairment and remodeling was displayed.in DCM . In contrast , The expression of TNF-α, IL-10 and PPARαwas decreased and the myocardial remodeling was attenuated by fenofiberate treatment, moreover, RT-PCR analysis further suggested that mRNA expression of TNF-αwas upregulated in DCM group(0.83±0.08), which was down regulated in LFIB and HFIB intervention groups. The values in these two groups were 0.58±0.07 and 0.55±0.07, respectively, both of which had significant difference compared with that of DCM group(P<0.01), while which still had significant difference compared with that of CON group. On the contrary, mRNA expression of IL-10 increased significantly in DCM group, while the expression was obviously upregulated in two intervention groups(P<0.01).Conclusion The alteration of myocardial energy metabolism and expression of inflammatory factors, myocardial remodeling are close related with the development of DCM. The activation of PPARαpathway could balance the expression of inflammatory factors and then improve myocardial remodeling and cardiac function.PartⅢInfluence of trimetazidine on myocardial remodeling and transforming growth factor-beta 1 expression in STZ-induced diabetic cardiomyopathy ratsObjective:To investigate the influence of trimetazidine on myocardial remodeling in diabetic cardiomyopathy in STZ-induced diabetic cardiomyopathy rats and the alteration of transforming growth factor-beta 1( TGF-β1).Method:Control rats were fed with common diet. Type 2 diabetes rats was established by high energy diet, then treated with lower dose of STZ and continuous high glucose and hyperlipid diet. The diabetic model animals were randomized into three groups: untreated DCM group, TMZ group and CAP group. A normal control group was set up in advance(n=8/group)After STZ injection and drug administration for sixteen weeks, LV function was evaluated by echocardiogram and heart weight index, collagen volume fraction (CVF )were calculated. myocardial cytokine gene and protein expression ( TNF-α, IL-10, PPARα) were detected. Myocardial TGF-β1 content and mRNA expression was quantified by immunohistochemistry and RT-PCR .Result:(1)DCM hearts demonstrated diabetic cardiomyopathy phenotype, including a significant reduction of LV diastolic function , reflected by lowered E/A ratio, shortened isovolumic relaxation time (IVRT) and decreased -dp/dt max .The CVF was increased to 0.38±0.09. The immunohistochemistry IDP and the level of mRNA were increased, which were (1.03±0.12)and (1.01±0.1), respectively. Compared with CON group, both groups had significant difference (P<0.01 in both groups). (2) In contrast, the ratio of heart weight/body weight was significantly decreased in TMZ and CAP rat groups compared with DCM rats(P<0.05); The diastolic function in TMZ and CAP groups was improved, the CVF was decreased(P<0.01)the values in both groups were (0.27±0.05) and(0.24±0.06), respectively. Compared with DCM group, both groups had significant difference (P<0.01 in both groups). IDP in both intervention groups were (0.75±0.08) and (0.74±0.08), respectively, which were significantly decreased compared with that in CON groups (P<0.01 in both groups). The mRNA expression of TGF-β1 was down regulated significantly, the values of TMZ and CAP groups were (0.71±0.09) and (0.73±0.22), respectively. Compared with CON group, both groups had significant difference (P<0.01 in both groups).Conclusion: Trimetazidine improved myocardial collagen deposition and myocardial remodeling in diabetic rats,associated with the decrease in expression of TGFβ1. Optimization of myocardial energy metabolism may be useful in prevention and treatment of diabetic myocardial disease.