Burow M E, Weldon C B, Tang Y, Navar G L, Krajewski S, Reed J C, Hammond T G, Clejan S, Beckman B S
Molecular and Cellular Biology Program, Tulane Cancer Center, Tulane University School of Medicine, New Orleans, Louisiana 70112, USA.
Cancer Res. 1998 Nov 1;58(21):4940-6.
Widespread use of MCF-7 human breast carcinoma cells as a model system for breast cancer has led to variations in these cells between different laboratories. Although several reports have addressed these differences in terms of proliferation and estrogenic response, variations in sensitivity to apoptosis have not yet been described. Tumor necrosis factor alpha (TNF-alpha) has been shown to both induce apoptosis and inhibit proliferation in MCF-7 cells. We observed that TNF-alpha inhibited proliferation in MCF-7 cell variants from three different laboratories (designated M, L, and N). MCF-7 M cells were resistant to TNF-alpha-induced apoptosis, whereas MCF-7 L cells were moderately resistant to the effect of TNF-alpha. A third variant, MCF-7 N, underwent apoptosis when exposed to TNF-alpha. Analysis of the p55 TNF-alpha receptor (TNFR) 1 expression revealed the greatest expression in MCF-7 N cells, whereas the MCF-7 L and M cells expressed 89 and 67% of MCF-7 N cell TNFR1 levels, respectively. Ceramide generation occurred in all three variants in response to TNF-alpha treatment, with MCF-7 N cells expressing the greatest increase. Cleavage of the CPP32/caspase 3 substrate poly(ADP-ribose) was observed in MCF-7 N and L cells as early as 3 and 6 h, respectively, but poly(ADP-ribose) cleavage was not observed in MCF-7 M cells. The delayed protease activation in the L variant may represent the mechanism by which these cells display delayed sensitivity to TNF-a-induced apoptosis. Expression of the Bcl-2, Mcl-1, Bcl-X, Bax, and Bak proteins was analyzed to determine whether the differences in MCF-7 cell sensitivity to apoptosis could be correlated to the differential expression of these proteins. Whereas Bak, Bcl-X, and Mcl-1 levels were identical between variants, the levels of Bcl-2 were 3.5-3.8-fold higher and the levels of Bax were 1.5-1.7-fold lower in the resistant variants (M and L) as compared with those of the sensitive variant (N). Taken together, these results suggest that differences in susceptibility to TNF-alpha-induced apoptosis among MCF-7 breast cancer cell variants may be explained by differences in TNFR expression, ceramide generation, differential expression of the Bcl-2 family of proteins, and protease activation.
MCF-7人乳腺癌细胞作为乳腺癌模型系统的广泛应用导致了不同实验室中这些细胞存在差异。尽管有几份报告从增殖和雌激素反应方面探讨了这些差异，但对凋亡敏感性的差异尚未见描述。肿瘤坏死因子α（TNF-α）已被证明在MCF-7细胞中既能诱导凋亡又能抑制增殖。我们观察到TNF-α抑制了来自三个不同实验室（分别命名为M、L和N）的MCF-7细胞变体的增殖。MCF-7 M细胞对TNF-α诱导的凋亡具有抗性，而MCF-7 L细胞对TNF-α的作用具有中等抗性。第三个变体MCF-7 N在暴露于TNF-α时会发生凋亡。对p55 TNF-α受体（TNFR）1表达的分析显示，MCF-7 N细胞中的表达量最高，而MCF-7 L和M细胞分别表达MCF-7 N细胞TNFR1水平的89%和67%。在所有三个变体中，TNF-α处理后都会产生神经酰胺，其中MCF-7 N细胞的增加最为明显。在MCF-7 N和L细胞中分别最早在3小时和6小时观察到CPP32/半胱天冬酶3底物聚（ADP-核糖）的切割，但在MCF-7 M细胞中未观察到聚（ADP-核糖）的切割。L变体中蛋白酶激活的延迟可能代表了这些细胞对TNF-α诱导的凋亡表现出延迟敏感性的机制。分析了Bcl-2、Mcl-1、Bcl-X、Bax和Bak蛋白的表达，以确定MCF-7细胞对凋亡敏感性的差异是否与这些蛋白的差异表达相关。与敏感变体（N）相比，抗性变体（M和L）中的Bak、Bcl-X和Mcl-1水平相同，但Bcl-2水平高3.5 - 3.8倍，Bax水平低1.5 - 1.7倍。综上所述，这些结果表明，MCF-7乳腺癌细胞变体对TNF-α诱导凋亡的敏感性差异可能由TNFR表达、神经酰胺生成、Bcl-2家族蛋白的差异表达以及蛋白酶激活的差异来解释。
