Automated analysis of contractile force and Ca2+ transients in engineered heart tissue
Automated analysis of contractile force and Ca2+ transients in engineered heart tissue
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DOI：
10.1152/ajpheart.00705.2013
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发表时间：
2014-05-01
影响因子：
4.8
通讯作者：
Hansen, Arne
中科院分区：
文献类型：
--
作者：
Stoehr, Andrea;Neuber, Christiane;Hansen, Arne
收缩和舒张是心肌细胞功能生物学的基本方面。它们反映了收缩机制对细胞质Ca2+浓度的收缩期升高和舒张期降低的反应。因此，分析收缩功能和Ca2+瞬态对于区分肌丝反应性和Ca2+稳态变化是重要的。本文描述了一种自动化技术，用于在灌注和电刺激下对多达11条条带格式、基于纤维蛋白的大鼠、小鼠和人类fura-2负载的工程心脏组织（EHTs）进行收缩力和Ca2+瞬态的序列分析。在增加细胞外Ca2+浓度和对异丙肾上腺素和碳乙醇的反应下，对eht的测量表明，eht概括了心脏组织功能生物学的基本原理。大鼠、小鼠和人eht的Ca2+浓度-反应曲线显示，三组大鼠、小鼠和人的最大抽搐力（大鼠、小鼠和人分别为0.22、0.05和0.08 mN, P < 0.001）和对外部Ca2+的敏感性（大鼠、小鼠和人的EC50分别为0.15、0.39和1.05 mM Ca2+, P < 0.001）不同。相比之下，在皮肤大鼠和人eht之间没有检测到肌丝Ca2+敏感性的差异，这表明对外部Ca2+浓度敏感性的差异是由于Ca2+处理蛋白的变化。最后，本研究证实fura-2具有Ca2+缓冲作用，从而改变对细胞外Ca2+的力响应。
Contraction and relaxation are fundamental aspects of cardiomyocyte functional biology. They reflect the response of the contractile machinery to the systolic increase and diastolic decrease of the cytoplasmic Ca2+ concentration. The analysis of contractile function and Ca2+ transients is therefore important to discriminate between myofilament responsiveness and changes in Ca2+ homeostasis. This article describes an automated technology to perform sequential analysis of contractile force and Ca2+ transients in up to 11 strip-format, fibrin-based rat, mouse, and human fura-2-loaded engineered heart tissues (EHTs) under perfusion and electrical stimulation. Measurements in EHTs under increasing concentrations of extracellular Ca2+ and responses to isoprenaline and carbachol demonstrate that EHTs recapitulate basic principles of heart tissue functional biology. Ca2+ concentration-response curves in rat, mouse, and human EHTs indicated different maximal twitch forces (0.22, 0.05, and 0.08 mN in rat, mouse, and human, respectively; P < 0.001) and different sensitivity to external Ca2+ (EC50: 0.15, 0.39, and 1.05 mM Ca2+ in rat, mouse, and human, respectively; P < 0.001) in the three groups. In contrast, no difference in myofilament Ca2+ sensitivity was detected between skinned rat and human EHTs, suggesting that the difference in sensitivity to external Ca2+ concentration is due to changes in Ca2+ handling proteins. Finally, this study confirms that fura-2 has Ca2+ buffering effects and is thereby changing the force response to extracellular Ca2+.