基于不同钙释放机理的花萼海绵诱癌素A与去乙酰毛花苷的正性肌力作用研究
Comparative Studies of Calyculin A and Deslanoside on Rat Isolated Heart Inotropy and Their Underlying Mechanism
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摘要: 目的 与洋地黄类正性肌力代表药去乙酰毛花苷比较,研究蛋白磷酸酶(PP)抑制剂花萼海绵诱癌素A(Calyculin A)对心脏收缩力作用特点,分析Calyculin A正性肌力作用的钙释放机理。探讨能否以PP为靶点,开发正性肌力药物治疗心力衰竭。方法 分别用Calyculin A(1,4,10 nmol/L)和去乙酰毛花苷(0.1,1,10 μmol/L)对大鼠离体心脏灌流,分析大鼠心率(HR)、左心室舒张压(LVDP)和最大上升速率(+dp/dtmax)的变化。采用Calyculin A(100 nmol/L)和去乙酰毛花苷(10 μmol/L)对大鼠左心室心肌细胞进行灌流,测定钙释放幅值、舒张期胞浆钙浓度(Ibaseline)及胞浆钙浓度恢复到静息状态值50%水平的时间(CTD50);分析Calyculin A(100 nmol/L)和去乙酰毛花苷(10 μmol/L)对咖啡因(20 mmol/L)诱导的大鼠心肌细胞钙释放的影响,分析肌浆网钙泵活性(SERCA2a)、钠-钙交换体(NCX)及肌膜钙泵(PMCA)综合活性的变化。结果 Calyculin A(1,4,10 nmol/L)与去乙酰毛花苷(0.1,1,10 μmol/L)均能显著增加离体大鼠LVDP和+dp/dtmax(P<0.05),且减慢HR(P<0.05),并呈现浓度依赖性。Calyculin A(100 nmol/L)与去乙酰毛花苷(10 μmol/L)均显著增加钙释放幅值(P<0.05);Calyculin A还能显著降低Ibaseline(P<0.05),显著缩短CTD50(P<0.05)。Calyculin A(100 nmol/L)显著增加肌浆网SERCA2a的速率常数(P<0.05);去乙酰毛花苷(10 μmol/L)增加(NCX+PMCA)的速率常数(P<0.05),导致肌浆网SERCA2a在胞浆回吸收中所发挥作用的百分比下降(P<0.05)。结论 Calyculin A与去乙酰毛花苷对大鼠离体心脏作用没有显著区别,但Calyculin A通过增强肌浆网SERCA2a的活性,增加钙释放幅值并降低Ibaseline实现其正性肌力作用。Calyculin A的作用靶点PP,可作为一个潜在靶点用于开发正性肌力药物。Abstract: OBJECTIVE To investigate the inotropic effect and its underlying mechanism of calyculin A, a protein phosphatase (PP) inhibitor, compared to deslanoside, a digitalis positive inotropic agent. To provide inputs whether PP can be served as a potential therapeutic target to develop pharmacological inotropic agents in treatment of heart failure. METHODS Ex vivo study was used to record the effects of calyculin A and deslanoside on rat isolated contractilities. They were perfused to follow in order: normal perfusion solution→calyculin A(1, 4, 10 nmol/L) or normal perfusion solution→deslanoside(0.1, 1, 10 μmol/L). Ca2+ transients triggered by field stimulation and by caffeine (20 mmol/L) were measured to analyze the Ca2+ handling effects of calyculin A (100 nmol/L) and deslanoside (10 μmol/L). RESULTS Both calyculin A (1, 4, 10 nmol/L) and deslanoside (0.1, 1, 10 μmol/L) significantly increased the left ventricular developed pressure and the peak rate of rise of left pressure (P<0.05) and decreased the heart rate (P<0.05). Calyculin A (100 nmol/L) and deslanoside (10 μmol/L) significantly increased the amplitude of Ca2+ transient and shortened the Ca2+ transient duration at 50% full recovery level (P<0.05);Calyculin A also lowered diastolic cytoplasm Ca2+ concentration (P<0.05). Calyculin A (100 nmol/L) significantly increased the SERCA2a activity and combinational activities of Na+-Ca2+ exchanger (NCX) and plasma membrane Ca2+-ATPase (PMCA) (P<0.05); Deslanoside (10 μmol/L) only increased combinational activities of NCX and PMCA (P<0.05),resulting in lowering percentage of contribution of SERCA2a in Ca2+ reuptake phase of Ca2+ transient (P<0.05). CONCLUSION Calyculin A increases the amplitude of Ca2+ transient by enhancing activity of SERCA2a and combinational activities of NCX and PMCA, resulting in the positive inotropy and favorable relaxation. PP can be a potential therapeutic target to develop pharmacological inotropic agents in treatment of heart failure.
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Key words:
- calyculin A /
- deslanoside /
- isolated heart /
- Ca2+ transient
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