Johnson Carolyn M, Kanter Evelyn M, Green Karen G, Laing James G, Betsuyaku Tetsuo, Beyer Eric C, Steinberg Thomas H, Saffitz Jeffrey E, Yamada Kathryn A
Department of Medicine, Washington University School of Medicine, St. Louis, MO 63110, USA.
Cardiovasc Res. 2002 Mar;53(4):921-35. doi: 10.1016/s0008-6363(01)00522-3.
Adult ventricular myocytes express two gap junction channel proteins: connexin43 (Cx43) and connexin45 (Cx45). Cx43-deficient mice exhibit slow ventricular epicardial conduction, suggesting that Cx43 plays an important role in intercellular coupling in the ventricle. Cx45 is much less abundant than Cx43 in working ventricular myocytes. Its role in ventricular conduction has not been defined, nor is it known whether expression or distribution of Cx45 is altered in Cx43-deficient mice. The present study was undertaken to determine (1) whether expression of Cx45 is upregulated and (2) whether gap junction structure and distribution are altered in Cx43-deficient mice.
Ventricular tissue from neonatal Cx43(+/+), Cx43(+/-) and Cx43(-/-) and adult Cx43(+/+) and Cx43(+/-) mice was analyzed by immunoblotting and confocal immunofluorescence microscopy.
Total Cx45 protein abundance measured by immunoblotting was not different in Cx43-deficient or null hearts compared to wild-type control hearts. However, the amount and distribution of Cx45 immunoreactive signal measured by quantitative confocal analysis were markedly reduced in both Cx43(+/-) and Cx43(-/-) hearts.
Although the total content of Cx45 is not upregulated in Cx43-deficient hearts, the localization of Cx45 to cardiac gap junctions depends on the expression level of Cx43 and is dramatically altered in mice that express no Cx43.
成年心室肌细胞表达两种缝隙连接通道蛋白：连接蛋白43（Cx43）和连接蛋白45（Cx45）。Cx43基因缺失的小鼠表现出心室心外膜传导缓慢，提示Cx43在心室细胞间偶联中起重要作用。在工作心室肌细胞中，Cx45的含量远低于Cx43。其在心室传导中的作用尚未明确，Cx43基因缺失的小鼠中Cx45的表达或分布是否改变也不清楚。本研究旨在确定：（1）Cx43基因缺失小鼠中Cx45的表达是否上调；（2）Cx43基因缺失小鼠中缝隙连接的结构和分布是否改变。
采用免疫印迹法和共聚焦免疫荧光显微镜技术，分析新生Cx43(+/+)、Cx43(+/-)和Cx43(-/-)小鼠以及成年Cx43(+/+)和Cx43(+/-)小鼠的心室组织。
免疫印迹法检测显示，与野生型对照心脏相比，Cx43基因缺失或无效的心脏中Cx45总蛋白丰度无差异。然而，定量共聚焦分析测得的Cx45免疫反应信号的数量和分布在Cx43(+/-)和Cx43(-/-)心脏中均显著降低。
虽然Cx43基因缺失的心脏中Cx45的总含量未上调，但Cx45在心脏缝隙连接中的定位取决于Cx43的表达水平，在不表达Cx43的小鼠中显著改变。
