Doxycycline inducible expression of SERCA2a improves calcium handling and reverts cardiac dysfunction in pressure overload-induced cardiac hypertrophy.
Doxycycline inducible expression of SERCA2a improves calcium handling and reverts cardiac dysfunction in pressure overload-induced cardiac hypertrophy.
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DOI：
10.1152/ajpheart.00428.2004
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发表时间：
2004-11
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通讯作者：
Jorge A. Suarez;B. Gloss;D. Belke;Ying Hu;Brian T. Scott;T. Dieterle;Yun-Kyung Kim;M. Valencik;J. McDonald;W. Dillmann
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作者：
Jorge A. Suarez;B. Gloss;D. Belke;Ying Hu;Brian T. Scott;T. Dieterle;Yun-Kyung Kim;M. Valencik;J. McDonald;W. Dillmann
衰竭心脏中心脏舒张延迟归因于肌（内）质网 Ca2+-ATPase 2a (SERCA2a) 活性和/或表达降低。尽管SERCA2a的组成型过度表达已被证明可有效预防心脏功能障碍，但尚不清楚在先前存在肥厚的心脏中增加SERCA2a表达是否具有治疗作用。为了检验这一假设，我们生成了一个二元转基因 (BTG) 系统，该系统允许四环素诱导的心脏特异性 SERCA2a 表达。在该系统（tet-on SERCA2a）中，FLAG标记的SERCA2a转基因在多西环素（Dox）存在的情况下表达，但在Dox不存在的情况下不表达（mRNA增加2.3倍，SERCA2a蛋白增加45%）。在来自非带状 Dox 处理的 BTG 小鼠的分离心肌细胞中测量的钙瞬变显示，钙下降加速，收缩期 Ca2+ 峰值增加。 BTG 小鼠的肌浆网 (SR) 钙负荷增加了 45%。在压力超负荷（主动脉束带）的情况下，超声心动图分析显示 SERCA2a-FLAG 的表达导致缩短分数改善。 SERCA2a-FLAG 表达减轻了由此产生的心脏功能障碍。这可以通过钙瞬变下降速率的增加来说明。 SERCA2a 过表达后，从带状 BTG 小鼠中分离的心肌细胞的细胞缩短和 SR 钙负荷也得到改善。总之，我们培育了一种有条件地过度表达 SERCA2a 的新型转基因小鼠。该模型适用于受控 SERCA2a 表达对心脏功能影响的长期和短期研究。此外，SERCA2a 的诱导性过度表达改善了患有收缩功能障碍的小鼠的心脏功能和钙处理。
Delayed cardiac relaxation in failing hearts has been attributed to reduced activity and/or expression of sarco(endo)plasmic reticulum Ca2+-ATPase 2a (SERCA2a). Although constitutive overexpression of SERCA2a has proven effective in preventing cardiac dysfunction, it is unclear whether increasing SERCA2a expression in hearts with preexisting hypertrophy will be therapeutic. To test this hypothesis, we generated a binary transgenic (BTG) system that allows tetracycline-inducible, cardiac-specific SERCA2a expression. In this system (tet-on SERCA2a), a FLAG-tagged SERCA2a transgene is expressed in the presence of doxycycline (Dox) but not in the absence of Dox (2.3-fold more mRNA, 45% more SERCA2a protein). Calcium transients measured in isolated cardiac myocytes from nonbanded Dox-treated BTG mice showed an accelerated calcium decline and an increased systolic Ca2+ peak. Sarcoplasmic reticulum (SR) calcium loading was increased by 45% in BTG mice. In the presence of pressure overload (aortic banding), echocardiographic analysis revealed that expression of SERCA2a-FLAG caused an improvement in fractional shortening. SERCA2a-FLAG expression alleviated the resultant cardiac dysfunction. This was illustrated by an increase in the rate of decline of the calcium transient. Cell shortening and SR calcium loading were also improved in cardiac myocytes isolated from banded BTG mice after SERCA2a overexpression. In conclusion, we generated a novel transgenic mouse that conditionally overexpresses SERCA2a. This model is suitable for both long- and short-term studies of the effects of controlled SERCA2a expression on cardiac function. In addition, inducible overexpression of SERCA2a improved cardiac function and calcium handling in mice with established contractile dysfunction.