Thiel William H, Chen Biyi, Hund Thomas J, Koval Olha M, Purohit Anil, Song Long-Sheng, Mohler Peter J, Anderson Mark E
Vanderbilt University, Nashville, TN, USA.
Circulation. 2008 Nov 25;118(22):2225-34. doi: 10.1161/CIRCULATIONAHA.108.788067. Epub 2008 Nov 10.
Timothy syndrome (TS) is a disease of excessive cellular Ca(2+) entry and life-threatening arrhythmias caused by a mutation in the primary cardiac L-type Ca(2+) channel (Ca(V)1.2). The TS mutation causes loss of normal voltage-dependent inactivation of Ca(V)1.2 current (I(Ca)). During cellular Ca(2+) overload, the calmodulin-dependent protein kinase II (CaMKII) causes arrhythmias. We hypothesized that CaMKII is a part of the proarrhythmic mechanism in TS.
We developed an adult rat ventricular myocyte model of TS (G406R) by lentivirus-mediated transfer of wild-type and TS Ca(V)1.2. The exogenous Ca(V)1.2 contained a mutation (T1066Y) conferring dihydropyridine resistance, so we could silence endogenous Ca(V)1.2 with nifedipine and maintain peak I(Ca) at control levels in infected cells. TS Ca(V)1.2-infected ventricular myocytes exhibited the signature voltage-dependent inactivation loss under Ca(2+) buffering conditions, not permissive for CaMKII activation. In physiological Ca(2+) solutions, TS Ca(V)1.2-expressing ventricular myocytes exhibited increased CaMKII activity and a proarrhythmic phenotype that included action potential prolongation, increased I(Ca) facilitation, and afterdepolarizations. Intracellular dialysis of a CaMKII inhibitory peptide, but not a control peptide, reversed increases in I(Ca) facilitation, normalized the action potential, and prevented afterdepolarizations. We developed a revised mathematical model that accounts for CaMKII-dependent and CaMKII-independent effects of the TS mutation.
In TS, the loss of voltage-dependent inactivation is an upstream initiating event for arrhythmia phenotypes that are ultimately dependent on CaMKII activation.
Timothy 综合征（TS）是一种因心脏主要的 L 型钙通道（Ca(V)1.2）突变导致细胞钙（Ca(2+)）过度内流和危及生命的心律失常的疾病。TS 突变导致 Ca(V)1.2 电流（I(Ca)）正常的电压依赖性失活丧失。在细胞钙超载期间，钙调蛋白依赖性蛋白激酶 II（CaMKII）会引发心律失常。我们推测 CaMKII 是 TS 致心律失常机制的一部分。
我们通过慢病毒介导的野生型和 TS Ca(V)1.2 转移，建立了成年大鼠心室肌细胞的 TS（G406R）模型。外源性 Ca(V)1.2 含有赋予二氢吡啶抗性的突变（T1066Y），因此我们可以用硝苯地平使内源性 Ca(V)1.2 沉默，并在感染细胞中将 I(Ca)峰值维持在对照水平。在不允许 CaMKII 激活的钙缓冲条件下，感染 TS Ca(V)1.2 的心室肌细胞表现出标志性的电压依赖性失活丧失。在生理钙溶液中，表达 TS Ca(V)1.2 的心室肌细胞表现出 CaMKII 活性增加和致心律失常表型，包括动作电位延长、I(Ca)易化增加和后去极化。对 CaMKII 抑制肽而非对照肽进行细胞内透析，可逆转 I(Ca)易化的增加，使动作电位正常化，并防止后去极化。我们开发了一个修订的数学模型，该模型考虑了 TS 突变的 CaMKII 依赖性和 CaMKII 非依赖性效应。
在 TS 中，电压依赖性失活的丧失是心律失常表型的上游起始事件，这些表型最终依赖于 CaMKII 的激活。
