MOUSE PHOSPHOLAMBAN GENE-EXPRESSION DURING DEVELOPMENT INVIVO AND INVITRO
MOUSE PHOSPHOLAMBAN GENE-EXPRESSION DURING DEVELOPMENT INVIVO AND INVITRO
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DOI：
10.1161/01.res.71.5.1021
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发表时间：
1992-11-01
影响因子：
20.1
通讯作者：
KRANIAS, EG
中科院分区：
文献类型：
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作者：
GANIM, JR;LUO, WS;KRANIAS, EG
为了建立一种小鼠模型，可以通过选择性扰动受磷蛋白基因来定义受磷蛋白在心肌收缩性中的确切作用，我们开始了对受磷蛋白在小鼠心脏中的作用的研究。在不存在或存在异丙肾上腺素的情况下灌注完整的跳动的心脏，并获得心脏性能的定量测量。异丙肾上腺素刺激与肌浆网Ca 2+泵对Ca 2+的亲和力增加有关，这是由于受磷蛋白磷酸化。为了评估受磷蛋白基因表达在小鼠发育过程中的调节，使用了北方印迹和聚合酶链反应分析。受磷蛋白mRNA首次在发育第九天（心管开始收缩的时间）的小鼠胚胎中检测到。在小鼠胚状体中，已经显示出重演心脏发生的几个方面，在第七天（首次观察到自发收缩的时间）检测到受磷蛋白mRNA。只有那些表现出收缩的胚状体表达受磷蛋白转录本，这些都伴随着蛋白质的表达，如免疫荧光显微镜所示。对胚状体中受磷蛋白的cDNA进行序列分析，结果表明其与成人心脏中的受磷蛋白完全同源。推导的小鼠受磷蛋白的氨基酸序列与兔心脏受磷蛋白相同，但与狗心脏和人心脏受磷蛋白相差一个氨基酸。这些数据表明，受磷蛋白，在心脏肌浆网中的Ca 2 +-ATP酶的调节剂，是目前在子宫内和体外的小鼠心脏发生非常早，这可能是一个重要的决定因素，为适当的发展心肌收缩力。
To establish a murine model that may allow for definition of the precise role of phospholamban in myocardial contractility through selective perturbations in the phospholamban gene, we initiated studies on the role of phospholamban in the murine heart. Intact beating hearts were perfused in the absence or presence of isoproterenol, and quantitative measurements of cardiac performance were obtained. Isoproterenol stimulation was associated with increases in the affinity of the sarcoplasmic reticulum Ca2+ pump for Ca2+ that were due to phospholamban phosphorylation. To assess the regulation of phospholamban gene expression during murine development, Northern blot and polymerase chain reaction analyses were used. Phospholamban mRNA was first detected in murine embryos on the ninth day of development (the time when the cardiac tube begins to contract). In murine embryoid bodies, which have been shown to recapitulate several aspects of cardiogenesis, phospholamban mRNA was detected on the seventh day (the time when spontaneous contractions are first observed). Only those embryoid bodies that exhibited contractions expressed phospholamban transcripts, and these were accompanied by expression of the protein, as revealed by immunofluorescence microscopy. Sequence analysis of the cDNA encoding phospholamban in embryoid bodies indicated complete homology to that in adult hearts. The deduced amino acid sequence of murine phospholamban was identical to rabbit cardiac phospholamban but different from dog cardiac and human cardiac phospholamban by one amino acid. These data suggest that phospholamban, the regulator of the Ca2+-ATPase in cardiac sarcoplasmic reticulum, is present very early in murine cardiogenesis in utero and in vitro, and this may constitute an important determinant for proper development of myocardial contractility.