Effects of thapsigargin and phenylephrine on calcineurin and protein kinase C signaling functions in cardiac myocytes.
Effects of thapsigargin and phenylephrine on calcineurin and protein kinase C signaling functions in cardiac myocytes.
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DOI：
10.1152/ajpcell.00594.2008
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发表时间：
2009-02
期刊：
影响因子：
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通讯作者：
A. Prasad;G. Inesi
中科院分区：
文献类型：
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作者：
A. Prasad;G. Inesi
将新生大鼠心肌细胞暴露于10 nM毒胡萝卜素（TG）或20 μ M苯肾上腺素（PE），以比较Ca（2+）稳态的变化。任何一种治疗均导致静息胞质[Ca（2+）]升高和电刺激后肌细胞中Ca（2+）信号的降低。事实上，ATP依赖性Ca（2+）转运减少是由于TG催化抑制肌浆网ATP酶（SERCA 2）或PE减少SERCA 2蛋白表达所致。TG或PE产生转染的荧光素酶cDNA的活化T细胞核因子（NFAT）依赖性表达的显著升高，这依赖于通过活化的钙调磷酸酶增加的NFAT去磷酸化和通过失活的糖原合成酶激酶3 β减少的磷酸化。暴露于TG后SERCA2（失活）蛋白的表达增加，而不产生肥大。相反，SERCA2表达减少，尽管高CN活性，蛋白激酶C（PKC）激活PE（或佛波醇12-肉豆蔻酸酯13-乙酸酯）的条件下产生肌细胞肥大。TG和PE的作用均依赖于CN对NFAT的去磷酸化作用，如环孢菌素（CsA）对CN的抑制作用所示。然而，由于NFAT和/或其他转录因子的竞争性募集，由PKC激活触发的肥大程序绕过SERCA2转录和表达。对CN活化的类似依赖性，但在PKC活化条件下相对减少，涉及Na（+）/Ca（2+）交换器-1的转录和表达。另一方面，PKC激活后瞬时受体电位通道蛋白显著上调。观察到的Ca（2+）稳态的改变可能有助于收缩衰竭的发展。
Neonatal rat cardiac myocytes were exposed to 10 nM thapsigargin (TG) or 20 muM phenylephrine (PE) to compare resulting alterations of Ca(2+) homeostasis. Either treatment results in resting cytosolic [Ca(2+)] rise and reduction of Ca(2+) signals in myocytes following electrical stimuli. In fact, ATP-dependent Ca(2+) transport is reduced due to catalytic inhibition of sarcoplasmic reticulum ATPase (SERCA2) by TG or reduction of SERCA2 protein expression by PE. A marked rise of nuclear factor of activated T cells (NFAT)-dependent expression of transfected luciferase cDNA is produced by TG or PE, which is dependent on increased NFAT dephosphorylation by activated calcineurin and reduced phosphorylation by inactivated glycogen synthase kinase 3beta. Expression of SERCA2 (inactivated) protein is increased following exposure to TG, whereas no hypertrophy is produced. On the contrary, SERCA2 expression is reduced, despite high CN activity, following protein kinase C (PKC) activation by PE (or phorbol 12-myristate 13-acetate) under conditions producing myocyte hypertrophy. Both effects of TG and PE are dependent on NFAT dephosphorylation by CN, as demonstrated by CN inhibition with cyclosporine (CsA). However, the hypertrophy program triggered by PKC activation bypasses SERCA2 transcription and expression due to competitive recruitment of NFAT and/or other transcriptional factors. A similar dependence on CN activation, but relative reduction under conditions of PKC activation, involves transcription and expression of the Na(+)/Ca(2+) exchanger-1. On the other hand, significant upregulation of transient receptor potential channel proteins is noted following PKC activation. The observed alterations of Ca(2+) homeostasis may contribute to development of contractile failure.