Gollasch M, Haase H, Ried C, Lindschau C, Morano I, Luft F C, Haller H
Franz-Volhard Clinic at the Max-Delbrück Center for Molecular Medicine, Humboldt University of Berlin, Germany.
FASEB J. 1998 May;12(7):593-601. doi: 10.1096/fasebj.12.7.593.
Despite intensive interest in understanding the differentiation of vascular smooth muscle cells (VSMC), no information is available about differential regulation of ion channels in these cells. Since expression of the L-type Ca2+ channel can be influenced by differentiation in other cell types, we tested the hypothesis that the L-type (C class) channel is a specific differentiation marker of VSMC and that expression of these channels depends on the state of cell differentiation. We used rat aortic (A7r5) VSMC, which express functional L-type Ca2+ channels, and induced dedifferentiation by cell culture in different media. Treatment with retinoic acid was used to redifferentiate the VSMC. We characterized the differentiated state of the cells by using immunohistochemistry and Western blot analysis for smooth muscle (SM) alpha-actin and SM-myosin heavy chain (MHC). The number of functional Ca2+ channels was significantly decreased in dedifferentiated VSMC and increased upon differentiation with retinoic acid. Ca2+ channel function was assessed by whole-cell voltage clamp techniques. Using Western blot and dihydropyridine binding analysis, we found that the expression of the Ca2+ channel alpha1 subunit, and to a lesser extent the beta2 subunit, was directly correlated with the expression of SM alpha-actin and SM-MHC. We conclude that expression of L-type Ca2+ channel alpha1 subunits, and thus a functional Ca2+ channel, is highly coordinated with expression of the SM-specific proteins required for specialized smooth muscle cell functions. Furthermore, our results demonstrate that the L-type Ca2+ channel is a novel marker for differentiation of VSMC. The data suggest that regulation of ion channel expression during differentiation may have physiological importance for normal smooth muscle function and may influence VSMC behavior under pathophysiological conditions.
尽管人们对了解血管平滑肌细胞（VSMC）的分化有着浓厚兴趣，但关于这些细胞中离子通道的差异调节尚无相关信息。由于L型Ca2+通道的表达在其他细胞类型中可受分化影响，我们检验了以下假设：L型（C类）通道是VSMC的一种特异性分化标志物，且这些通道的表达取决于细胞分化状态。我们使用表达功能性L型Ca2+通道的大鼠主动脉（A7r5）VSMC，并通过在不同培养基中进行细胞培养诱导其去分化。用视黄酸处理使VSMC重新分化。我们通过免疫组织化学和蛋白质印迹分析平滑肌（SM）α-肌动蛋白和SM-肌球蛋白重链（MHC）来表征细胞的分化状态。去分化的VSMC中功能性Ca2+通道的数量显著减少，而用视黄酸分化后则增加。通过全细胞电压钳技术评估Ca2+通道功能。使用蛋白质印迹和二氢吡啶结合分析，我们发现Ca2+通道α1亚基的表达，以及程度较轻的β2亚基的表达，与SMα-肌动蛋白和SM-MHC的表达直接相关。我们得出结论，L型Ca2+通道α1亚基的表达，以及由此产生的功能性Ca2+通道，与特殊平滑肌细胞功能所需的SM特异性蛋白的表达高度协调。此外，我们的结果表明L型Ca2+通道是VSMC分化的一种新型标志物。数据表明，分化过程中离子通道表达的调节可能对正常平滑肌功能具有生理重要性，并可能在病理生理条件下影响VSMC的行为。
