Ruzicka F J, Schmid S M, Groveman D S, Cummings K B, Borden E C
Cancer Res. 1987 Sep 1;47(17):4582-9.
Studies of various established human bladder and renal carcinoma cell lines cultured in vitro demonstrated the presence of specific, saturable, high affinity binding sites for 125I-labeled human interferon Beta ser IFN-beta ser). This recombinant produced interferon labeled with approximately one atom of 125I/molecule of IFN expressed minimal or no loss of antiviral activity. A single class of binding sites (1000-2000/cell) with an affinity constant of 10(10)-10(11) L/M was measured at 4 degrees C for cells exhibiting widely different sensitivity to the antiproliferative effect of IFN-beta ser. Major fluctuations in the binding of 125I-labeled IFN-beta ser to cellular receptors were observed during in vitro proliferation of four of five cell lines examined. A significant decrease (P less than 0.001) in specific binding was observed 48 h after cultures were established. Cell cycle analysis suggested that within the first 24 h and in the very late log and stationary phase of growth of ACHN (human renal carcinoma) cells, variations in the binding of 125I-labeled IFN-beta ser were partially attributable to binding fluctuations during the mitotic cycle. The 2- to 3-fold decline 24 h following plating of ACHN cells corresponded to a 70% decrease in the number of cells in G0-G1. T24 (human transitional cell carcinoma) and ACHN cells, synchronized by serum starvation, demonstrated increased binding of 125I-labeled IFN-beta ser 4-16 h following serum replenishment. This increase in receptor binding occurred prior to the onset of DNA and protein synthesis and was followed by a decline immediately prior to cell division. Binding site analysis indicated that the increased binding prior to DNA synthesis was due to a 5- to 6-fold increase in receptor affinity for the radiolabeled ligand. After an initial 40% decline in receptors per cell following serum stimulation, receptor concentration remained essentially unchanged. Induction of 2',5'-oligoadenylate synthetase in ACHN cells and antiproliferative activity in RT112, RT4, T24 (human transitional cell carcinoma), and ACHN cells by IFN-beta ser decreased significantly 48 h following plating. These changes in the biological activity of this interferon corresponded to growth related fluctuations in the IFN-beta ser binding.
对多种在体外培养的已建立的人膀胱和肾癌细胞系进行的研究表明，存在针对125I标记的人β-干扰素（ser IFN-β ser）的特异性、可饱和、高亲和力结合位点。这种重组产生的干扰素每分子IFN用约一个125I原子标记，其抗病毒活性极少丧失或无丧失。在4℃下，对于对ser IFN-β的抗增殖作用表现出广泛不同敏感性的细胞，测量到一类结合位点（1000 - 2000个/细胞），其亲和常数为10(10)-10(11) L/M。在所检测的五个细胞系中的四个细胞系的体外增殖过程中，观察到125I标记的ser IFN-β与细胞受体的结合有主要波动。在培养建立48小时后，观察到特异性结合显著下降（P小于0.001）。细胞周期分析表明，在ACHN（人肾癌）细胞生长的最初24小时内以及对数生长期后期和静止期，125I标记的ser IFN-β结合的变化部分归因于有丝分裂周期中的结合波动。ACHN细胞接种24小时后结合下降2至3倍，对应于G0 - G1期细胞数量减少70%。通过血清饥饿同步化的T24（人移行细胞癌）和ACHN细胞，在补充血清后4 - 16小时显示125I标记的ser IFN-β结合增加。这种受体结合的增加发生在DNA和蛋白质合成开始之前，紧接着在细胞分裂之前下降。结合位点分析表明，DNA合成之前结合增加是由于受体对放射性标记配体的亲和力增加5至6倍。血清刺激后，每个细胞的受体最初下降40%后，受体浓度基本保持不变。ACHN细胞中2',5'-寡腺苷酸合成酶的诱导以及RT112、RT4、T24（人移行细胞癌）和ACHN细胞中的抗增殖活性在接种48小时后显著下降。这种干扰素生物活性的这些变化与ser IFN-β结合的生长相关波动相对应。
