Calcium-sensing receptors induce apoptosis in cultured neonatal rat ventricular cardiomyocytes during simulated ischemia/reperfusion
Calcium-sensing receptors induce apoptosis in cultured neonatal rat ventricular cardiomyocytes during simulated ischemia/reperfusion
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DOI：
10.1016/j.cellbi.2008.03.009
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发表时间：
2008-07-01
影响因子：
3.9
通讯作者：
Li, Wei-ming
中科院分区：
文献类型：
--
作者：
Jiang, Chun-ming;Han, Li-Ping;Li, Wei-ming
钙敏感受体（calcium -sensing receptor, CaSRs）是调节全身钙稳态的g蛋白偶联受体，参与细胞增殖、分化和凋亡。我们之前已经证明，CaSR可以诱导离体大鼠成年心脏和正常大鼠新生心肌细胞凋亡。然而，CaSR在新生儿心肌细胞缺血再灌注诱导的细胞凋亡中的作用尚不清楚。因此，在本研究中，我们将新生大鼠心室心肌细胞在模拟缺血溶液中孵育2 h，然后在正常培养基中再孵育24 h，建立模拟缺血/再灌注（I/R）模型。流式细胞术检测心肌细胞凋亡率；透射电镜观察形态学改变；Western blotting分析caspase-3、Bcl-2、CaSR、细胞外信号调节蛋白激酶（ERK）、Fas/Fas配体（FasL）的表达；用激光共聚焦扫描显微镜测定细胞内钙浓度。结果表明，模拟I/R增加了CaSR的表达和心肌细胞凋亡。GdCl3是一种特异性的CaSR激活剂，在I/R期间，随着心肌细胞内钙和细胞凋亡的增加，进一步增强了CaSR的表达。激活CaSR可下调Bcl-2表达，上调caspase-3和Fas/FasL表达，刺激ERK1/2磷酸化。综上所述，CaSR通过抑制Bcl-2、诱导钙超载、激活Fas/FasL死亡受体通路参与新生大鼠心室心肌细胞I/R损伤和凋亡。(C) 2008国际细胞生物学联合会。Elsevier Ltd.出版。版权所有。
Calcium-sensing receptors (CaSRs) are G-protein coupled receptors which regulate systemic calcium homeostasis and also participate in cell proliferation, differentiation and apoptosis. We have previously shown that CaSR can induce apoptosis in isolated rat adult hearts and in normal rat neonatal cardiomyocytes. However, no knowledge exists concerning the role of CaSR in apoptosis induced by ischemia and reperfusion in neonatal cardiac myocytes. Therefore, in the present study, we incubated primary neonatal rat ventricular cardiomyocytes in ischemia-mimetic solution for 2 h, then re-incubated them in a normal culture medium for 24 h to establish a model of simulated ischemia/reperfusion (I/R). We assayed the apoptotic ratio of the cardiomyocytes by flow cytometry; observed morphological alterations by transmission electron microscope; analyzed the expression of caspase-3, Bcl-2, CaSR, extracellular signal-regulated protein kinase (ERK), and Fas/Fas ligand (FasL) by Western blotting; and measured the concentration of intracellular calcium by Laser Confocal Scanning Microscopy. The results showed that simulated I/R increased the expression of CaSR and cardiomyocyte apoptosis. GdCl3, a specific activator of CaSR, further enhanced CaSR expression, along with increases in intracellular calcium and apoptosis in cardiomyocytes during I/R. Activation of CaSR down-regulated Bcl-2 expression, upregulated caspase-3 and Fas/FasL expression and stimulated ERK1/2 phosphorylation. In summary, CaSR is involved in I/R injury and apoptosis of neonatal rat ventricular cardiomyocytes by inhibiting Bcl-2, inducing calcium overload and activating the Fas/FasL death receptor pathway. (C) 2008 International Federation for Cell Biology. Published by Elsevier Ltd. All rights reserved.