Adenovirus-mediated transfer of the smooth muscle cell calponin gene inhibits proliferation of smooth muscle cells and fibroblasts
Adenovirus-mediated transfer of the smooth muscle cell calponin gene inhibits proliferation of smooth muscle cells and fibroblasts
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DOI：
10.1016/s0014-5793(97)00944-7
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发表时间：
1997-08-25
期刊：
影响因子：
3.5
通讯作者：
Kamm, KE
中科院分区：
文献类型：
--
作者：
Jiang, Z;Grange, RW;Kamm, KE
平滑肌细胞钙调蛋白（h1或碱性同种型）是一种肌动蛋白结合蛋白，其抑制肌动球蛋白MgATP酶活性，并在分化的平滑肌中大量表达，Western印迹搁置的牛气管（BT）平滑肌细胞在培养物中仅表达组织中钙调蛋白量的2+/-1%（n=8），而NIH-3 T3成纤维细胞不表达，我们测试了将钙调蛋白引入培养的BT和3 T3细胞会抑制与细胞增殖相关的细胞骨架活性的假设。为了实现高效表达，通过在293细胞中同源重组产生编码CMV-钙调蛋白构建体（Adv-CaP）的腺病毒，在用Adv-CaP感染的BT和3 T3细胞中，超过90%的钙调蛋白表达（分别为32和11 μ g/mg总蛋白）与平滑肌组织中的钙调蛋白表达（51 μ g/mg）相似。用Adv-CaP感染细胞48 h，以低密度重新铺板，通过细胞密度和[H-3]胸苷掺入来评估增殖速率。细胞生长和DNA合成的Adv-CaP感染的细胞被抑制到三分之一的控制值为BT和3 T3细胞，表达的calponin主要定位于应力纤维在两种细胞类型。钙调蛋白可以在细胞骨架水平起作用，以阻碍通常导致细胞形状和DNA合成之间紧密耦合的信号通路。(C)1997年欧洲生物化学学会联合会。
Smooth muscle cell calponin (h1 or basic isoform) is an actin-binding protein that inhibits actomyosin MgATPase activity and is abundantly expressed in differentiated smooth muscle, Western blots shelved bovine tracheal (BT) smooth muscle cells in culture expressed only 2+/-1% (n=8) of the amount of calponin in tissues, while NIH-3T3 fibroblasts expressed none, We tested the hypothesis that introduction of calponin to cultured BT and 3T3 cells would inhibit cytoskeletal activities associated,vith cell proliferation, To achieve high-efficiency expression, an adenovirus encoding the CMV-calponin construct (Adv-CaP) was generated by homologous recombination in 293 cells, With greater than 90% of BT and 3T3 cells infected with Adv-CaP, calponin expression (32 and 11 mu g/mg total protein, respectively) was similar to that in smooth muscle tissues (51 mu g/mg). Cells were infected with Adv-CaP for 48 h, replated at low density and proliferation rates were assessed by cell density and [H-3]thymidine incorporation. Cell growth and DNA synthesis by Adv-CaP-infected cells were inhibited to one-third control values for both BT and 3T3 cells, Expressed calponin was localized primarily on stress fibers in both cell types. Calponin may act at the cytoskeletal level to retard signaling pathways that normally lead to tight coupling between cell shape and DNA synthesis. (C) 1997 Federation of European Biochemical Societies.