北京大学生命科学学院王世强教授领导的实验室与北京大学第三医院张幼怡研究员领导的实验室通力合作，利用心衰动物模型开展了心衰早期发病的分子病理机制研究，业已取得重要科研成果。1月9日，有较高学术影响的国际期刊《PloSBiology》发表了他们的研究论文。
据介绍，心衰是威胁人类生命的重大疾病之一，患者最终因心肌收缩能力不足而死亡。心衰往往由心肌肥厚演变而来，因此阐明心肌肥厚到心衰的演变规律对防治心衰有重要意义。
王世强教授和张幼怡研究员的协作组为了探索这一演变的分子机制，对控制心肌兴奋收缩耦联的钙信号转导过程进行了细胞和亚细胞水平的功能鉴定，并从分子间相互作用的角度系统研究了细胞膜钙离子通道与肌质网Ryanodine受体相互耦联的分子动力学过程，首次发现两分子的耦联效率在心肌肥厚发生早期、细胞收缩功能尚无变化的时候就已经发生了衰退。衰退的原因是锚定两分子所在膜结构的蛋白分子Junctophilin的表达量下降。Junctophilin的减少使细胞膜钙离子通道与肌质网Ryanodine受体间耦联效率进行性地衰退，最终导致细胞整体钙信号下降和心肌收缩能力的降低。这一研究为心衰病理发生提供了重要分子机制。
根据上述发现，他们还提出了生理功能“稳定余量”的概念，并证明稳定余量的存在是细胞钙信号等生理系统保持功能稳态的前提；分子耦联效率的降低在一定范围内只是在消耗稳定余量，不会引起细胞整体功能的变化；当稳定余量最终被耗竭后，分子耦联效率的进行性下降将表现为细胞钙信号和心脏收缩功能的不断恶化。这一论述阐释了心衰病理发生的演变规律，并为心衰的早期诊断、防治的必要性和可行性提供了理论依据。
部分英文原文：
Intermolecular Failure of L-type Ca2+ Channel and Ryanodine Receptor Signaling in Hypertrophy
Ming Xu, Peng Zhou, Shi-Ming Xu, Yin Liu, Xinheng Feng, Shu-Hua Bai, Yan Bai, Xue-Mei Hao, Qide Han, Youyi Zhang*, Shi-Qiang Wang*
1 State Key Lab of Biomembrane and Membrane Biotechnology, Ministry of Education Key Lab of Molecular Cardiovascular Sciences and Institute of Vascular Medicine, Third Hospital, College of Life Sciences, Peking University, Beijing, China
Pressure overload–induced hypertrophy is a key step leading to heart failure. The Ca2+-induced Ca2+ release (CICR) process that governs cardiac contractility is defective in hypertrophy/heart failure, but the molecular mechanisms remain elusive. To examine the intermolecular aspects of CICR during hypertrophy, we utilized loose-patch confocal imaging to visualize the signaling between a single L-type Ca2+ channel (LCC) and ryanodine receptors (RyRs) in aortic stenosis rat models of compensated (CHT) and decompensated (DHT) hypertrophy. We found that the LCC-RyR intermolecular coupling showed a 49% prolongation in coupling latency, a 47% decrease in chance of hit, and a 72% increase in chance of miss in DHT, demonstrating a state of “intermolecular failure.” Unexpectedly, these modifications also occurred robustly in CHT due at least partially to decreased expression of junctophilin, indicating that intermolecular failure occurs prior to cellular manifestations. As a result, cell-wide Ca2+ release, visualized as “Ca2+ spikes,” became desynchronized, which contrasted sharply with unaltered spike integrals and whole-cell Ca2+ transients in CHT. These data suggested that, within a certain limit, termed the “stability margin,” mild intermolecular failure does not damage the cellular integrity of excitation-contraction coupling. Only when the modification steps beyond the stability margin does global failure occur. The discovery of “hidden” intermolecular failure in CHT has important clinical implications.
Introduction
In response to pressure overload, the heart produces an adaptive response in the form of cardiac hypertrophy to maintain adequate cardiac output and tissue perfusion [1–3]. In the early stage of hypertrophy, cardiac contractile dysfunction is not present, and the ventricle is hemodynamically compensated. When the pressure stimuli are persistent, the heart usually undergoes functional deterioration, eventually leading to heart failure [3,4]. In the failure stage, the heart becomes incapable of generating sufficient pumping power. To prevent the pathogenesis of heart failure, one strategy has been to stop or postpone the transition of hypertrophy from the compensated stage toward the decompensated stage [4]. Therefore, understanding the cellular and molecular mechanisms involved in cardiac hypertrophy is important for developing clinical therapies against heart failure.
At the cellular level, the contractile power during excitation-contraction coupling (E-C coupling) is governed by a mechanism known as Ca2+-induced Ca2+ release (CICR) [5,6]. In this process, Ca2+ influx through L-type Ca2+ channels (LCCs) on the cell surface membrane (including T-tubules) activates ryanodine receptor (RyR) Ca2+ release from the sarcoplasmic reticulum (SR) to generate cell-wide Ca2+ transients [7–9]. Besides LCCs and RyRs, Ca2+ cycling proteins, e.g., SR Ca2+ pumps (SERCA), Na+-Ca2+ exchangers, and their regulatory mechanisms, are also important in determining the amplitude and kinetics of Ca2+ transients [8]. All these mechanisms have been studied in a wide variety of hypertrophy and heart failure models [8,10–14]. Most studies support the idea that the LCC activity does not change much during hypertrophy and heart failure [11]. However, the Ca2+ transients triggered by comparable LCC currents are decreased in amplitude and/or slowed in kinetics in most models of decompensated hypertrophy (DHT) and heart failure [11,13]. These studies lead to the notion that the Ca2+ influx through LCCs becomes less effective in triggering RyR Ca2+ release [13]. Yet the molecular details underlying defective E-C coupling remain unknown. On the other hand, studies on compensated hypertrophy (CHT), a stage prior to DHT, show that the cellular aspects of E-C coupling still appear to be normal or even slightly enhanced [15]. It is thus intriguing to know whether and when the intermolecular process of CICR is modified and how the modification eventually leads to cellular failure in E-C coupling.
During past years, we have developed a local Ca2+ imaging protocol in conjunction with a loose-seal patch clamp technique to investigate LCC-RyR intermolecular coupling [9,16]. In the present study, we utilized this technique and an aortic stenosis model to test the hypothesis that the intermolecular coupling between an LCC and RyRs undergoes a progressive modification during the development of hypertrophy. Our results showed that hypertrophy resulted in an increase in LCC-RyR coupling latency and a decrease in intermolecular signaling efficiency, which started at the early, compensated stage when cellular E-C coupling appeared normal. Our findings provided intermolecular insights into the remodeling of Ca2+ signaling during the pathogenesis leading to heart failure.
Results
To elucidate the microscopic modification of E-C coupling during hypertrophy, we created pressure-overload hypertrophy models induced by aortic stenosis [17]. About 7–11 wk after aorta banding, hemodynamic and echocardiographic measurements identified the status of CHT by increased left ventricle (LV) wall thickness and normal contractile indices, and the status of DHT by the onset of mild depression of contractile indices in addition to thickened LV walls (Figure 1A, 1B, and 1C; Table S1). To characterize the cellular aspects of E-C coupling, we combined a whole-cell patch clamp technique and confocal line-scan imaging to record simultaneously LCC Ca2+ current (ICa) and intracellular Ca2+ transients when the cell membrane was depolarized to 0 mV (Figure 1D). Cell capacitance (Figure S1A) and contraction (Figure S1B) were also measured. In DHT, despite the unchanged ICa density and kinetics (Figure 1E and Figure S1C and S1D), both the amplitude of Ca2+ transients and cell contraction decreased significantly (Figure 1F and Figure S1B). As a result, the gain of E-C coupling was significantly lower than that of the control (Figure 1G). By contrast, neither the amplitude of Ca2+ transients nor the gain of E-C coupling was altered in CHT, indicating that the hypertrophy-associated E-C coupling deficiency occurs only in the late, decompensated stage, but not in the early, compensated stage.
更多原文链接：
http://biology.plosjournals.org/perlserv/?request=get-document&doi=10.1371/journal.pbio.0050021
