| Objective: As scaffolding structures of cardiomyocytes, the cytoskeleton plays vital role in normal myocardium structure and function. Investigation shows that the cytoskeletal alteration is dominant in the pathogenesis of cardiac hypertrophy and congestive heart failure. However, little is known about the relationship between cytoskeleton and some vital neuroendocrine factors such as Angiotensin Ⅱ. This study is aimed to investigate the alterations of microtubules in hypertrophied cardiomyocytes induced by Angiotensin Ⅱ as well as the relationship between calcineurin pathway and microtubules. Methods: 1. The isolated neonatal Wistar rat ventricular cardiomyocytes were cultured and purified with the method of differential attachment. 2. The cultured cells were identified by morphology, spontaneous contraction and specific immunocytochemical stain. 3. Hypertrophied cardiomyocyte model induced by Angiotensin Ⅱwas set up. 4. By [~3H]-Leu incorporation, model of hypertrophied cardiomyocytes was assessed. 5. The ultrastructural characteristics in hypertrophied cardiomyocytes were investigated with the method of transmission electron microscopy as well as structural reorganization of microtubules with immumofluorescence. 6. Content of β-tubulin in hypertrophied cardiomyocytes and other groups was measured by western blotting, and specific activity of calcineurin was detected with the method of chromatometry. Results: 1. The shapes of primary cultured cardiomyocytes are spindle ,triangle and irregular. Among total cultured cells, the spindle cells are majority of primary cultured cardiomyocytes. Spontaneous contraction of the spindle cell is apparent. There have abundant brownish red filaments in cytoplasm of primary cultured cardiomyocytes with the method of specific immunocytochemical stain. 2. Ultrastructural changes consist of enlargement and varying shape of nuclei, enlarged mitochondria, abundant heterochromatin and accumulation of lipid droplets and glycogen. The ultrastructural alteration is unspecified performance of compensatory hypertrophy. 3. [~3H]-Leu incorporation in cardiomyocyte of AngiotensinⅡgroup is significantly higher than that of the control group which indicats synthesis of protein increased induced by AngiotensinⅡ. Whereas [~H]-Leuincorporation in cardiomyocytes increase or decrease treated respectively with taxol or colchicine. 4. With the method of immumofluorescence and western blotting, the quantity of microtubule and β-tubulin increases in AngiotensinⅡgroup. On the basis of AngiotensinⅡ, the quantity of microtubules increased or decreased treated respectively with taxol and colchicine.5. Specific activity of calcineurin in AngiotensinⅡgroup is much higher than that in control group. Whereas specific activity of calcineurin increased or decreased treated respectively with taxol or colchicine.Conclusions: 1.With the method of collagenase digestion, it is convenient to obtain primary cultured neonatal Wistar rat ventricular cardiomyocytes. This method is one of vital research tools in the fields of cardiac hypertrophy, heart failure, hypertension as well as coronary heart disease. 2. By 10~-7mol/L concentration, AngiotensinⅡcould accelerate cellular proliferation,structural protein biosynthesis and cardiac hypertrophy. 3. Density and stability of microtubule as well as β-tubulin biosynthesis are increased in hypertrophy cardiomyocyte induced by AngiotensinⅡ. 4. Spacial microtubule conformation was rearranged in hypertrophy cardiomyocyte induced by AngiotensinⅡ. With this structural remodeling, microtubule has very important function in the pathogenesis of cardiac hypertrophy. 5. Colchicine and taxol could bind with microtubule and change spacial conformation of microtubule. In this way, colchicine and taxol can adjust signal transduction in the process of cardiac hypertrophy. 6. With the method of structural remodeling, microtubule has effect on specific activity of calcineurin, and has very important function in the calcium metabolism and pathogenesis of cardi...
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