体外培养心肌细胞代谢和微管蛋白对拉伸应变的响应

The Response of Metabolism and Tubulin of Cardiac Myocytes in vitro to Mechanical Stretch

探讨不同参数的拉伸应变与体外培养心肌细胞代谢和微管蛋白含量的关系。建立体外细胞基底膜拉伸装置,分析膜上应变分布。分离乳鼠心肌细胞接种在膜上,施加不同大小和频率的周期性应变,观察细胞形态,测定培养液中葡萄糖和乳酸含量,Western Blot法测定细胞内聚合态微管蛋白-βtubulin含量。自行研制的基底膜拉伸装置性能稳定。所得心肌细胞成活率较高,α-横纹肌肌动蛋白和结蛋白免疫细胞化学染色阳性。拉伸后细胞由多角形趋向于杆状,且排列方向改变。固定频率1 Hz时,8%应变组的葡萄糖比消耗速率,乳酸比生成速率和-βtubulin含量均明显高于对照组（P〈0.05）。固定应变8%时,1 Hz的周期性拉伸对细胞代谢的促进作用最为明显（P〈0.01）,频率0.5~2 Hz的拉伸应变均可使-βtubulin含量升高。适当的拉伸应变可促进体外培养心肌细胞的代谢和微管聚合。

The aim of this study is to investigate the effects of stretch on growth and metabolism of cardiac myocytes in vitro. Cardiac myocytes from ventricles of neonatal rats were isolated and cultured on silicone membranes. The silicone membranes were stretched cyclically and the two-Dimensional finite element model was developed to analyze the strain distribution of the membrane. The cells morphology was observed by a phase contrast microscope after cultured under cyclic stretch with different frequency. The contents of glucose and lactic acid in the supernatant were assayed to determine the metabolism of cells. Intracellular β-tubulin expression was analyzed by western blot. Most of the cardiac myocytes grew well on the membranes experiencing cyclic stretch and express α-sacromeric actin and desmin. Morphology of the cells changed from polygon to rhabditiform, and the orientation of cells altered. When the strain frequency was fixed at 1 Hz, the specific glucose consumption rate, the specific lactate production rate and the polymeric tubulin content of 8 % strain group were obviously higher than those of static group （ P 〈 0.05）. When the strain magnitude was fixed at 8 %, stimuli of 1 Hz strain gave the most enhancement to metabolism of the cells （ P 〈 0.01）. The polymeric tubulin contents in all frequency levels were higher than that of static group when strain magnitude was 8%. Proper mechanical stretch could improve the metabolism and polymeric tubulin contents of cardiac myocytes in vitro.