Backeljauw P F, Dai Z, Clemmons D R, D'Ercole A J
Department of Pediatrics, University of North Carolina, Chapel Hill 27599-7220.
Endocrinology. 1993 Apr;132(4):1677-81. doi: 10.1210/endo.132.4.7681763.
FRTL-5 cells, a diploid nontransformed line of rat thyroid follicular cells, exhibit a marked mitogenic response to insulin-like growth factors (IGFs) when they are exposed to TSH. Because IGF binding proteins (IGFBPs) are important modulators of IGF actions, we investigated the capacity of FRTL-5 cells to synthesize IGFBPs. We found that FRTL-5 cell conditioned media contained a single band of approximately 31 kilodaltons on ligand blot analysis. This band represents IGFBP-5 because: 1) it can be immunostained with a specific antibody raised against human IGFBP-5; 2) by Northern analysis, total RNA from FRTL-5 cells contains a major 6-kilobase transcript when hybridized with a cDNA for rat IGFBP-5; and 3) no transcripts were observed when Northern blots of FRTL-5 cells were hybridized with complementary DNAs for rat IGFBP-1, -2, -3, -4 or -6. Conditioned media IGFBP-5 increased in response to IGF-I, IGF-II, and insulin in a dose-dependent fashion, compared with unstimulated FRTL-5 cells. At maximally effective concentrations IGF-I was 3.5- and 6-fold more potent than IGF-II and insulin, respectively. The addition of a monoclonal antibody (Sm 1.2) to IGF-I completely inhibited IGF-I stimulation of the IGFBP-5 6-kilobase transcript and the appearance of IGFBP-5 in FRTL-5 conditioned media. Stimulation of IGFBP-5 synthesis by the IGFs and insulin appeared to be regulated at the messenger RNA (mRNA) level, because each stimulated similar increases in both IGFBP-5 mRNA and media protein at maximally effective concentrations. TSH, on the other hand, inhibited basal levels of IGFBP-5 mRNA and attenuated the increase in IGFBP-5 mRNA stimulated by IGF-I. FRTL-5 cells provide a relatively uncomplicated model to study the regulation and action of IGFBP-5 and the mechanisms by which IGFs interact with this binding protein.
FRTL-5细胞是大鼠甲状腺滤泡细胞的二倍体非转化细胞系，当暴露于促甲状腺激素（TSH）时，对胰岛素样生长因子（IGFs）表现出明显的促有丝分裂反应。由于IGF结合蛋白（IGFBPs）是IGF作用的重要调节因子，我们研究了FRTL-5细胞合成IGFBPs的能力。我们发现，在配体印迹分析中，FRTL-5细胞条件培养基含有一条约31千道尔顿的单带。这条带代表IGFBP-5，原因如下：1）它可以用针对人IGFBP-5产生的特异性抗体进行免疫染色；2）通过Northern分析，当与大鼠IGFBP-5的cDNA杂交时，FRTL-5细胞的总RNA含有一个主要的6千碱基转录本；3）当FRTL-5细胞的Northern印迹与大鼠IGFBP-1、-2、-3、-4或-6的互补DNA杂交时，未观察到转录本。与未刺激的FRTL-5细胞相比，条件培养基中的IGFBP-5以剂量依赖的方式对IGF-I、IGF-II和胰岛素作出反应而增加。在最大有效浓度下，IGF-I分别比IGF-II和胰岛素有效3.5倍和6倍。向IGF-I中加入单克隆抗体（Sm 1.2）完全抑制了IGF-I对IGFBP-5 6千碱基转录本的刺激以及IGFBP-5在FRTL-5条件培养基中的出现。IGFs和胰岛素对IGFBP-5合成的刺激似乎在信使RNA（mRNA）水平受到调节，因为在最大有效浓度下，它们各自刺激IGFBP-5 mRNA和培养基蛋白出现相似的增加。另一方面，TSH抑制IGFBP-5 mRNA的基础水平，并减弱IGF-I刺激的IGFBP-5 mRNA的增加。FRTL-5细胞为研究IGFBP-5的调节和作用以及IGFs与这种结合蛋白相互作用的机制提供了一个相对简单的模型。
