Cellular growth response to epidermal growth factor in colon carcinoma cells with an amplified epidermal growth factor receptor derived from a familial adenomatous polyposis patient.
Cellular growth response to epidermal growth factor in colon carcinoma cells with an amplified epidermal growth factor receptor derived from a familial adenomatous polyposis patient.
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发表时间：
1991-03
期刊：
影响因子：
11.2
通讯作者：
M. E. Gross;M. Zorbas;Y. J. Danels;R. Garcia;G. Gallick;Matilde Olive;M. Brattain;Bruce M. Boman;L. Yeoman
中科院分区：
文献类型：
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作者：
M. E. Gross;M. Zorbas;Y. J. Danels;R. Garcia;G. Gallick;Matilde Olive;M. Brattain;Bruce M. Boman;L. Yeoman
使用从家族性腺瘤性息肉病患者建立的DiFi细胞系研究了对外源性表皮生长因子（EGF）的受体结合和细胞生长反应。通过竞争性放射性配体结合试验和125 I-EGF结合等温线的Scatchard分析，计算出DiFi细胞上细胞膜EGF受体的数量为4.8 x 10（6）个受体/细胞。在配体结合试验之前进行的酸预洗步骤未显示额外的受体数量。一个单一的，低亲和力的受体群体，通过Scatchard分析确定，具有4.6 nM的表观Kd。该结果通过在存在和不存在受体拮抗剂单克隆抗体528 IgG的情况下进行的放射性配体结合研究得到证实，该抗体主要与EGF受体的低亲和力形式结合。当暴露于50 nM外源性EGF时，50-60%汇合的DiFi细胞表现出其细胞膜相关受体的快速但部分（30%）减少，这是隔离的特征。DiFi细胞暴露于50 nM EGF较长时间（4小时）不会导致EGF受体数量的任何进一步减少。在单层培养物中以及在软琼脂糖测定中研究DiFi细胞对外源性EGF的细胞生长反应。在大于1.7 nM的外源EGF浓度下观察到软琼脂集落形成的抑制，并且在大于1 nM的EGF浓度下发生单层生长的抑制。在免疫复合物激酶测定中，DiFi受体显示出与充分表征的A431细胞系相似的特异性活性。此外，酪氨酸上受体的磷酸化在性质上与A431细胞的类似，进一步表明通过EGF结合研究鉴定的DiFi受体具有生物学功能。
The receptor binding and cellular growth responses to exogenous epidermal growth factor (EGF) were studied using the DiFi cell line established from a familial adenomatous polyposis patient. The number of cell membrane EGF receptors on DiFi cells, as measured by competitive radioligand binding assays and Scatchard analysis of 125I-EGF binding isotherms, was calculated to be 4.8 x 10(6) receptors/cell. An acid prewash step performed prior to ligand binding assays did not reveal additional receptor numbers. A single, low-affinity receptor population was identified by Scatchard analysis, with an apparent Kd of 4.6 nM. This result was confirmed by radioligand binding studies performed in the presence and absence of the receptor-antagonist monoclonal antibody 528 IgG that binds predominantly to the low-affinity form of the EGF receptor. DiFi cells at 50-60% confluence, when exposed to 50 nM exogenous EGF, exhibited a rapid but partial (30%) reduction in their cell membrane-associated receptor, characteristic of sequestration. Exposure of DiFi cells to 50 nM EGF for longer periods of time (4 h) did not result in any further reduction in EGF-receptor number. The cellular growth response of DiFi cells to exogenous EGF was studied in monolayer cultures as well as in a soft agarose assay. Inhibition of soft agar colony formation was observed at exogenous EGF concentrations greater than 1.7 nM, and inhibition of monolayer growth occurred at EGF concentrations greater than 1 nM. In immune complex kinase assays, the DiFi receptor showed similar specific activity to that from the well-characterized A431 cell line. Additionally, phosphorylation of the receptor on tyrosine was qualitatively similar to that of A431 cells, further suggesting that the DiFi receptors identified by EGF-binding studies were biologically functional.