乳化异氟醚对缺氧复氧乳鼠心肌细胞的保护作用及bcl-2、bax表达的影响

Effect of emulsified isoflurane on myocardial apoptosis and expressions of bcl-2 and bax induced by hypoxia-reoxygenation

目的观察乳化异氟醚(8%体积分数)对培养乳鼠心肌细胞模拟缺血再灌注损伤后心肌细胞凋亡及凋亡相关基因bcl-2、bax的mRNA与蛋白水平表达的影响。方法利用原代培养的Wistar乳鼠心肌细胞建立模拟心肌缺血再灌注模型,分正常培养组、模型组、脂肪乳组、乳化异氟醚(终浓度0.28mmol/L)。各组心肌细胞做相应分组处理后,利用倒置显微镜观察心肌细胞的生长状态和搏动频率,并用透射电镜观察细胞超微结构改变;流式细胞仪检测心肌细胞凋亡率;RT-PCR检测心肌细胞凋亡基因bcl-2 mRNA、bax mRNA的表达。Western blot定量检测Bcl-2、Bax蛋白表达。结果乳化异氟醚组和脂肪乳组均可显著降低细胞凋亡率(P<0.05),但乳化异氟醚组与脂肪乳组比较降低更明显(P<0.05)。乳化异氟醚可显著上调Bcl-2表达(P<0.05)、下调Bax表达(P<0.05);脂肪乳对Bcl-2和Bax表达无影响(P>0.05)。结论乳化异氟醚及其其中的成分之一脂肪乳可抑制模拟缺血再灌注损伤心肌细胞的凋亡;乳化异氟醚对缺氧复氧损伤的抗凋亡作用与其心肌细胞Bcl-2表达上调和Bax表达下调有关。

Objective To investigate the effects of emulsified isoflurane on myocardial apoptosis, and the expressions of Bcl-2 and Bax after hypoxia/reoxygenation in cultured myocardial cells of neonatal rats. Methods Myocardial hypoxia/reoxygenation model was established on cultured primary myocardial cells of neonatal rat. The myocardial cells were divided into four groups： the normal group, model group, fat milk group, and emulsified isoflurane group. The cellular morphologic changes and pulsatile frequency of each group were observed under inverted microscope and the uhrastructure of myocardial cells was observed under transmission electron microscope. The apoptotic rate of myocardial cells was determined by flow cytometry, the expression of bcl-2 mRNA and bax mRNA were detected by RT-PCR and the protein expressions of Bcl-2/Bax were observed by Western blot. Results The apoptotic rate were significantly decreased in both emulsified isoflurane and fat milk groups （ P 〈 0. 05 ） , especially in emulsified isoflurane group. Bcl-2 expressions was significantly up-regulated and Bax expressions was down-regulated in emulsified isoflurane group （P 〈 0. 05 ） , while those showed no significant changes in fat milk group （P 〉0.05）. Conclusion Emulsified isoflurane and one of its compo- nents, fat milk, can inhibit myocardial apoptosis induced by hypoxia/reoxygenation in cultured myocardial cells. Emulsified isoflurane exerts its anti-apoptosis effect maybe through up-regulating Bcl-2 expressions and downregulating Bax expressions.