Laminin responsiveness is associated with changes in fibroblast morphology, motility, and anchorage-independent growth: cell system for examining the interaction between laminin and EGF signaling pathways.
Laminin responsiveness is associated with changes in fibroblast morphology, motility, and anchorage-independent growth: cell system for examining the interaction between laminin and EGF signaling pathways.
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层粘连蛋白反应性与成纤维细胞形态、运动性和锚定无关生长的变化相关：用于检查层粘连蛋白和 EGF 信号通路之间相互作用的细胞系统。
DOI：
10.1002/jcp.1041640318
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发表时间：
1995
期刊：
影响因子：
--
通讯作者：
Bertics,PJ
中科院分区：
文献类型：
--
作者：
Lin,ML;Bertics,PJ
层粘连蛋白可以影响包括上皮细胞和神经细胞在内的几种细胞类型的粘附、分化和运动性。此外，含有表皮生长因子（EGF）样基序的层粘连蛋白可以刺激含有EGF受体的成纤维细胞的DNA合成，但层粘连蛋白并不竞争EGF的结合。为了进一步研究层粘连蛋白在成纤维细胞中的作用，以及层粘连蛋白与EGF受体功能之间的关系，我们开发了一种系统，其中从不能粘附层粘连蛋白的小鼠成纤维细胞系（B82L‐wt）中克隆出具有层粘连蛋白结合活性的细胞，但转染了野生型人EGF受体。虽然只有分离的克隆可以有效地附着在层粘连蛋白覆盖的板上，但所有的细胞都可以附着在塑料、纤维连接蛋白和胶原I上，并且都表现出相当水平的细胞表面相关的层粘连蛋白。配体结合实验表明，具有层粘连蛋白附着活性的细胞具有高亲和力的EGF结合（Kd ~ 0.4 nM），并且都表达相似水平的人EGF受体。然而，与B82L - wt细胞相比，具有层粘连蛋白结合活性的细胞表现出形态改变、不依赖于锚定的生长和运动性。具体来说，具有层粘连蛋白结合活性的成纤维细胞的形态看起来更细长，它们在塑料板上扩散得更广泛。对它们在软琼脂中生长的分析显示，与B82L‐wt细胞相比，克隆的集落形成增加了2‐5倍。具有层粘连蛋白附着能力的细胞也表现出层粘连蛋白诱导的运动，这种运动是定向的（趋化性）而不是随机的（趋化作用），表明层粘连蛋白受体和信号通路的功能。为了研究参与这些作用的特异性层粘连蛋白受体，我们评估了抗整合素亚单位抗体对细胞粘附和迁移的影响。这些研究表明，抗α6整合素抗体可以完全抑制克隆细胞对层粘连蛋白的附着和迁移，并且抗α6免疫印迹显示只有克隆表达可测量的α6水平。这些数据表明，含有α6的整合素有助于这些克隆的层状介导的附着和运动，并且该系统也可能影响这些成纤维细胞的形态和不依赖于锚定的生长。此外，这些细胞为检测EGF和层粘连蛋白受体之间的相互作用提供了一个独特的系统。©1995 Wiley‐Liss, Inc。
Laminin can influence the adhesion, differentiation, and motility of motility of several cell types, including epithelial and neural cells. In addition, laminin, which contains an epidermal growth factor (EGF)‐like motif, can stimulate DNA synthesis in fibroblasts possessing the EGF receptor, but laminin does not compete for EGF binding. To further investigate laminin action in fibroblasts, and the relationship between laminin and EGF receptor function, we have developed a system wherein cells containing laminin‐binding activity were cloned from a mouse fibroblast cell line (B82L‐wt) that cannot adhere to laminin but that have been transfected with the wild‐type human EGF receptor. Although only the isolated clones can efficiently attach to laminin‐coated plates, all the cells can adhere to plastic, fibronectin, and collagen I, and all exhibit comparable levels of cell surface‐associated laminin. Ligand‐binding assays showed that the cells with laminin attachement activity possess high‐affinity EGF binding (Kd ∼ 0.4 nM), and all express a similar level of the human EGF receptor. However, when compared to the B82L‐wt cells, the cells with laminin‐binding activity exhibit altered morphology, anchorage‐independent growth, and motility. Specifically, the morphology of the fibroblasts possessing laminin binding activity appears more elongated and they spread more‐extensively on plastic plates. Analysis of their growth in soft agar revealed that the clones have a 2‐5‐fold increase in colony formation in comparison to the B82L‐wt cells. The cells possessing laminin attachment ability also exhibit laminin‐induced motility, and this movement is directional (chemotaxis) rather than random (chemokinesis), indicating functional laminin receptors and signaling pathways. To examine the specific laminin receptors involved in these effects, the influence of anti‐integrin subunit antibodies on cell adhesion and migration was evaluated. These studies showed that an anti‐α6integrin antibody can completely inhibit the clonal cells' attachment and migration to laminin, and anti‐α6immunoblots revealed that only the clones express measurable levels of α6. These data indicate that α6‐containing integrins contribute to the lamininmediated attachment and motility of these clones and that this system may also influence the morphology and anchorage‐independent growth of these fibroblasts. In addition, these cells provide a unique system for examining the interaction between EGF and laminin receptor action. © 1995 Wiley‐Liss, Inc.