Vuk-Pavlović S, Kovach J S
Division of Developmental Oncology Research, Mayo Foundation, Rochester, Minnesota 55905.
FASEB J. 1989 Dec;3(14):2633-40. doi: 10.1096/fasebj.3.14.2556313.
Kinetics of regulation of membrane receptors for tumor necrosis factor-alpha (TNF) in human breast adenocarcinoma MCF-7 cells was investigated. When MCF-7 cells were incubated with radioiodinated human recombinant TNF, they bound TNF specifically and accumulated it intracellularly. Preincubation of cells with native TNF up to 1 x 10(-9) M for 12 h stimulated specific binding of TNF, indicating that concentrations of membrane receptors for TNF were regulated by the ligand. Accumulation of radioactivity in cells incubated with [125I]TNF proceeded at a constant rate for up to 24 h. Kinetics of binding and internalization of TNF were similar in the presence and absence of protein synthesis for at least 1 h, suggesting that the TNF receptor was either replenished from an intracellular pool of receptors or was recycled (reutilized) during the course of TNF internalization. Data were analyzed kinetically by fitting equations of compartmental models of ligand-cell interactions with and without the term for receptor recycling. Fits were obtained only for the model with receptor recycling; absence of the term for receptor recycling resulted in physically impossible best-fit parameter values. Analysis of the best-fit parameters indicated that both internalization and recycling of the receptor were stimulated by the ligand.
研究了人乳腺腺癌MCF - 7细胞中肿瘤坏死因子-α（TNF）膜受体的调节动力学。当MCF - 7细胞与放射性碘化的人重组TNF孵育时，它们特异性结合TNF并在细胞内积累。用天然TNF将细胞预孵育至1×10⁻⁹ M，持续12小时，可刺激TNF的特异性结合，表明TNF膜受体的浓度受配体调节。用[¹²⁵I]TNF孵育的细胞中放射性的积累在长达24小时内以恒定速率进行。在有和没有蛋白质合成的情况下，TNF的结合和内化动力学至少在1小时内相似，这表明TNF受体要么从细胞内受体池中补充，要么在TNF内化过程中被循环利用（再利用）。通过将配体 - 细胞相互作用的房室模型方程与有和没有受体循环项进行拟合，对数据进行动力学分析。仅对具有受体循环的模型获得了拟合；没有受体循环项会导致物理上不可能的最佳拟合参数值。对最佳拟合参数的分析表明，受体的内化和循环均受配体刺激。
