Shneyvays V, Nawrath H, Jacobson K A, Shainberg A
Department of Life Sciences, Bar-Ilan University, Ramat-Gan, Israel.
Exp Cell Res. 1998 Sep 15;243(2):383-97. doi: 10.1006/excr.1998.4134.
The effects of the selective adenosine (ADO) A3 receptor agonist IB-MECA (N6-(3-iodobenzyl)adenosine-5'-N-methylcarboxamide) on cultured newborn rat cardiomyocytes were examined in comparison with ADO, the ADO A1 receptor-selective agonist R-PIA (N6-R-phenylisopropyladenosine), or the ADO A3 selective antagonist MRS 1191 (3-ethyl-5-benzyl-2-methyl-6-phenyl-4-phenylethynyl-1, 4-(+/-)-dihydropyridine-3,5 dicarboxylate), using digital image analysis of Feulgen-stained nuclei. At high concentration, IB-MECA (>/=10 microM ) and ADO (200 microM) induced apoptosis; however, R-PIA or MRS 1191 did not have any detectable effects on cardiac cells. In addition, DNA breaks in cardiomyocytes undergoing apoptosis following treatment by IB-MECA were identified in situ using the nick end labeling of DNA ("TUNEL"-like) assay. In the presence of >/=10 microM IB-MECA, disorder in the contraction waves appeared, and a decrease in the frequency of beats was observed. Analysis with light microscopy revealed that the number of contracting cells decreased in a concentration-dependent manner. The A3 receptor agonist IB-MECA caused an increase in intracellular free calcium concentration ([Ca2+]i). The drug produced a rapid rise followed by a sustained increase in [Ca2+]i, which lasted for 40-60 s. Finally, cessation of beating and Ca2+ transients were observed. Full recovery of contractile activity and rhythmical Ca2+ transients were observed 15-20 min after IB-MECA treatment. The induction of apoptosis in the cardiocytes by IB-MECA led to the appearance of features of apoptotic nuclei: the onset of condensation, compacting, and margination of nuclear chromatin. These effects were accompanied by the disintegration of the structural framework of the nucleus and nuclear breakdown. The results suggest that activation of the A3 adenosine receptor may participate in the process of apoptosis in cardiomyocytes.
使用Feulgen染色细胞核的数字图像分析技术，将选择性腺苷（ADO）A3受体激动剂IB-MECA（N6-(3-碘苄基)腺苷-5'-N-甲基甲酰胺）与ADO、ADO A1受体选择性激动剂R-PIA（N6-R-苯基异丙基腺苷）或ADO A3选择性拮抗剂MRS 1191（3-乙基-5-苄基-2-甲基-6-苯基-4-苯乙炔基-1,4-(+/-)-二氢吡啶-3,5-二羧酸酯）相比较，研究其对培养的新生大鼠心肌细胞的影响。高浓度时，IB-MECA（≥10 μM）和ADO（200 μM）可诱导细胞凋亡；然而，R-PIA或MRS 1191对心肌细胞无任何可检测到的影响。此外，使用DNA缺口末端标记（“TUNEL”样）分析法原位鉴定了经IB-MECA处理后发生凋亡的心肌细胞中的DNA断裂。在存在≥10 μM IB-MECA的情况下，收缩波出现紊乱，且观察到搏动频率降低。光学显微镜分析显示，收缩细胞数量呈浓度依赖性减少。A3受体激动剂IB-MECA导致细胞内游离钙浓度（[Ca2+]i）升高。该药物使[Ca2+]i迅速升高，随后持续增加，持续40 - 60秒。最后，观察到搏动停止和Ca2+瞬变。在IB-MECA处理后15 - 20分钟观察到收缩活性和节律性Ca2+瞬变完全恢复。IB-MECA诱导心肌细胞凋亡导致凋亡细胞核特征出现：核染色质开始凝聚、致密化和边缘化。这些效应伴随着细胞核结构框架的解体和核破裂。结果表明，A3腺苷受体的激活可能参与心肌细胞凋亡过程。
