Effects of mutant and antisense RNA of phospholamban on SR Ca2+-ATPase activity and cardiac myocyte contractility
Effects of mutant and antisense RNA of phospholamban on SR Ca2+-ATPase activity and cardiac myocyte contractility
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DOI：
10.1161/01.cir.100.9.974
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发表时间：
1999-08-31
期刊：
影响因子：
37.8
通讯作者：
Dillmann, WH
中科院分区：
文献类型：
--
作者：
He, HP;Meyer, M;Dillmann, WH
研究背景-心力衰竭时心脏舒张延迟是由于肌浆网Ca 2 +-ATP酶（SERCA 2）活性降低所致，受磷蛋白（PLB）抑制SERCA 2活性，因此是改善心力衰竭时心脏功能的潜在靶点。用重组腺病毒在心肌细胞中表达PLB（Adv/mPLB）ol反义RNA（Adv/asPLB），观察其对心肌细胞SERCA 2活性和收缩力的影响。一个mPLB，K3 E/R14 E，在新生儿心肌细胞中与内源性PLB五聚化，并导致SERCA 2对Ca 2+的亲和力增加45%，舒张期Ca 2+下降加快27%，分别通过SR Ca-45摄取测定和indo I促进的Ca 2+瞬时测量测定。成人心肌细胞收缩力的边缘检测分析显示，缩短分数增加74%，伴随着115%的速度增加的再延长和25%的时间减少到一半的最大再延长。同时，Adv/asPLB感染新生心肌细胞可使内源性PLB水平降低54%，这与SERCA 2的Ca 2+亲和力增加35%和舒张期Ca 2+下降加快21%相关。然而，在成人心肌细胞，腺病毒/asPLB未能显着改变内源性PLB水平，SERCA 2活性，或大多数的contractile parameters. Conclusions-K3 E/R14 E是一个显性负突变体的PLB，破坏了结构的完整性和功能的内源性PLB，从而增强SERCA 2活性和心肌细胞收缩性。在新生心肌细胞中，asPLB导致的PLB稳态丰度的降低也导致SERCA 2活性的增加。
Background-The delayed cardiac relaxation in failing hearts has been attributed to a reduced activity of sarcoplasmic reticulum Ca2+-ATPase (SERCA2), Phospholamban (PLB) inhibits SERCA2 activity and is therefore a potential target to improve the cardiac performance in heart failure.Methods and Results-Mutants of PLB (Adv/mPLB) ol antisense RNA of PLB (Adv/asPLB) was expressed in cardiac myocytes by recombinant adenovirus, and their effects on SERCA2 activity and myocyte contractility were studied. One mPLB, K3E/R14E, pentamerized with endogenous PLB in neonatal myocytes and resulted in a 45% increase in the affinity of SERCA2 for Ca2+ and 27% faster diastolic Ca2+ decline as determined by SR Ca-45 uptake assays and by indo I-facilitated Ca2+ transient measurement, respectively. Edge-detection analysis of adult myocyte contractility showed a 74% increase in fractional shortening, accompanied by 115% increase in velocity of relengthening and 25% decrease in time to half-maximal relengthening. In parallel, infection of neonatal cardiac myocytes by Adv/asPLB decreased the endogenous PLB level by 54%, which was associated with a 35% increase in Ca2+ affinity of SERCA2 and 21% faster diastolic Ca2+ decline. However, in adult cardiac myocytes, Adv/asPLB failed to significantly alter the endogenous PLB level, the SERCA2 activity, or most of the contractile parameters.Conclusions-K3E/R14E is a dominant negative mutant of PLB that disrupts the structural integrity and function of the endogenous PLB and consequently enhances SERCA2 activity and myocyte contractility. In neonatal myocytes, the decrease in steady-state abundance of PLB by asPLB also leads to increased SERCA2 activity.