Coutts A, Murphy L J, Murphy L C
Department of Biochemistry, Faculty of Medicine, University of Manitoba, Winnipeg, Canada.
Breast Cancer Res Treat. 1994;32(2):153-64. doi: 10.1007/BF00665766.
We have used ligand blotting and Northern blotting techniques to examine the effects of progestins and antiestrogens on expression of insulin-like growth factor binding proteins (IGFBPs) by T-47D human breast cancer cells under conditions where these agents are growth inhibitory. Under basal conditions, conditioned medium from T-47D cells was found to contain IGFBPs of 39, 33, and 27 kDa. Northern blot and/or Western blot analysis have identified these as IGFBP 2, 5, and 4, respectively. Medroxyprogesterone acetate (MPA) treatment resulted in a time- and dose-dependent decrease in IGFBP 4 and 5 mRNA abundance and secretion of these proteins, while little if any effect was observed on IGFBP 2 expression. A decrease in the steady state mRNA levels for IGFBP 4 and 5 was observed with as little as 0.1 nM MPA. Using 10 nM MPA a maximum decrease in IGFBP 4 and 5 mRNA levels was observed between 12 and 24 hours. While RU 486 alone had little or no effect on IGFBP 4 expression, it inhibited the effect of MPA. However, in the same samples, IGFBP 5 expression was inhibited by RU 486, and RU 486 was unable to reverse the effects of progestins on the expression of IGFBP 5. Furthermore, another synthetic progestin, Org 2058, but not dexamethasone, inhibited IGFBP 4 and IGFBP 5 expression. The antiestrogen ICI 164384 also transiently decreased the steady state mRNA levels of both IGFBP 4 and IGFBP 5. Regulation of expression of the IGFBPs by these agents suggests a potential role for the IGFBPs in the growth response of T-47D cells to these agents.
我们运用配体印迹法和Northern印迹技术，在孕激素和抗雌激素具有生长抑制作用的条件下，研究它们对T-47D人乳腺癌细胞胰岛素样生长因子结合蛋白（IGFBPs）表达的影响。在基础条件下，发现T-47D细胞的条件培养基中含有分子量分别为39、33和27 kDa的IGFBPs。Northern印迹和/或Western印迹分析分别将这些蛋白鉴定为IGFBP 2、5和4。醋酸甲羟孕酮（MPA）处理导致IGFBP 4和5的mRNA丰度以及这些蛋白的分泌呈时间和剂量依赖性降低，而对IGFBP 2的表达几乎没有影响。低至0.1 nM的MPA就能使IGFBP 4和5的稳态mRNA水平下降。使用10 nM的MPA，在12至24小时之间观察到IGFBP 4和5的mRNA水平最大程度降低。虽然单独使用RU 486对IGFBP 4的表达几乎没有影响，但它抑制了MPA的作用。然而，在相同样本中，RU 486抑制了IGFBP 5的表达，并且RU 486无法逆转孕激素对IGFBP 5表达的影响。此外，另一种合成孕激素Org 2058而非地塞米松抑制了IGFBP 4和IGFBP 5的表达。抗雌激素ICI 164384也短暂降低了IGFBP 4和IGFBP 5的稳态mRNA水平。这些药物对IGFBPs表达的调节表明IGFBPs在T-47D细胞对这些药物的生长反应中可能发挥作用。
