TNF-α-mediated caspase-8 activation induces ROS production and TRPM2 activation in adult ventricular myocytes
TNF-α-mediated caspase-8 activation induces ROS production and TRPM2 activation in adult ventricular myocytes
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DOI：
10.1093/cvr/cvu112
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发表时间：
2014-07-01
影响因子：
10.8
通讯作者：
Fauconnier, Jeremy
中科院分区：
文献类型：
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作者：
Roberge, Stephanie;Roussel, Julien;Fauconnier, Jeremy
TRPM 2是瞬时受体电位（TRP）超家族的Ca 2+渗透性阳离子通道，其与凋亡信号传导相关。其在心脏病理生理学中的作用尚不清楚。本研究的目的是确定促凋亡细胞因子肿瘤坏死因子-α（TNF-α）是否在小鼠心室心肌细胞中诱导TRPM 2样电流。Western印迹显示TRPM 2表达，其在TNF-α孵育后没有改变。使用膜片钳在全细胞配置，非特异性阳离子电流被记录后，暴露于TNF-α（I-TNF），达到最大稳态振幅后3小时孵育。半胱天冬酶-8抑制剂z-IETD-favorite、抗氧化剂N-乙酰半胱氨酸和TRPM 2抑制剂克霉唑、N-（P-戊基肉桂酰基）邻氨基苯甲酸和氟芬那酸（FFA）可抑制I-TNF。TRPM 2先前已被证明是由ADP-核糖激活，这是由聚（ADP-核糖）聚合酶1（PARP-1）产生。TNF-α暴露导致蛋白质的聚ADP核糖基化增加，PARP-1抑制剂3-氨基苯甲酰胺抑制I-TNF。TNF-α暴露增加了活性氧（ROS;用荧光指示剂MitoSOX Red测量）的线粒体产生，并且这种增加被caspase-8抑制剂z-IETD-fetamine阻断。克霉唑和TRPM 2抑制性抗体降低TNF-α诱导的心肌细胞死亡。这些结果表明，TNF-α诱导成年心室心肌细胞中的TRPM 2电流。TNF-α诱导半胱天冬酶-8活化，导致ROS产生、PARP-1活化和ADP-核糖产生。TNF诱导的TRPM 2活化可能导致心肌细胞死亡。
TRPM2 is a Ca2+-permeable cationic channel of the transient receptor potential (TRP) superfamily that is linked to apoptotic signalling. Its involvement in cardiac pathophysiology is unknown. The aim of this study was to determine whether the pro-apoptotic cytokine tumour necrosis factor-alpha (TNF-alpha) induces a TRPM2-like current in murine ventricular cardiomyocytes.Adult isolated cardiomyocytes from C57BL/6 mice were exposed to TNF-alpha (10 ng/mL). Western blotting showed TRPM2 expression, which was not changed after TNF-alpha incubation. Using patch clamp in whole-cell configuration, a non-specific cation current was recorded after exposure to TNF-alpha (I-TNF), which reached maximal steady-state amplitude after 3 h incubation. I-TNF was inhibited by the caspase-8 inhibitor z-IETD-fmk, the antioxidant N-acetylcysteine, and the TRPM2 inhibitors clotrimazole, N-(P-amylcinnamoyl) anthranilic acid and flufenamic acid (FFA). TRPM2 has previously been shown to be activated by ADP-ribose, which is produced by poly(ADP-ribose) polymerase 1 (PARP-1). TNF-alpha exposure resulted in increased poly-ADP-ribosylation of proteins and the PARP-1 inhibitor 3-aminobenzamide inhibited I-TNF. TNF-alpha exposure increased the mitochondrial production of reactive oxygen species (ROS; measured with the fluorescent indicator MitoSOX Red), and this increase was blocked by the caspase-8 inhibitor z-IETD-fmk. Clotrimazole and TRPM2 inhibitory antibody decreased TNF-alpha-induced cardiomyocyte death.These results demonstrate that TNF-alpha induces a TRPM2 current in adult ventricular cardiomyocytes. TNF-alpha induces caspase-8 activation leading to ROS production, PARP-1 activation, and ADP-ribose production. TNF-induced TRPM2 activation may contribute to cardiomyocyte cell death.