Development of a bioassay to screen for chemicals mimicking the anti-aging effects of calorie restriction
Development of a bioassay to screen for chemicals mimicking the anti-aging effects of calorie restriction
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DOI：
10.1016/j.bbrc.2010.09.032
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发表时间：
2010-10-15
影响因子：
3.1
通讯作者：
Shimokawa, Isao
中科院分区：
文献类型：
--
作者：
Chiba, Takuya;Tsuchiya, Tomoshi;Shimokawa, Isao
在艾姆斯侏儒（DF）小鼠中抑制生长激素/胰岛素样生长因子-I通路，在正常小鼠中限制热量摄入（CR）可延长寿命并延迟年龄相关疾病的发作。结合起来，这些干预措施对艾姆斯DF小鼠的寿命具有累加效应。因此，DF和CR调节的共同信号通路可能对寿命具有累加效应。在这项研究中，我们试图识别信号机制，并开发一个系统来评估细胞和小鼠的促长寿状态。我们先前通过DNA微阵列分析确定了DF和CR小鼠肝脏中上调的基因。这些基因的上游序列的基序分析揭示了四个主要的共识序列基序，已被命名为侏儒症和热量限制反应元件（DFCR-RE）。与肝细胞核因子-4 α（HNF-4 α）结合的合成序列之一，HNF-4 α是参与肝脏代谢的重要转录因子。此外，利用这些序列信息，我们开发了一种高灵敏度的生物测定法来鉴定模拟CR抗衰老作用的化学物质。当含有分泌型碱性磷酸酶（SEAP）基因上游元件的报告基因构建体与HNF-4 α及其调节因子过氧化物酶体增殖物激活受体（PPAR）γ共激活因子-1 α（PGC-1 α）共转染时。与未转染的对照相比，SEAP活性增加。此外，使用该构建体建立的瞬时转基因小鼠在CR小鼠中显示出比自由进食小鼠增加的SEAP活性。这些数据表明，由于其快速性，易用性和特异性，我们的生物测定将比目前用于筛选CR模拟物的系统更有用，该系统模拟CR的有益作用。我们的系统将是特别有用的高通量筛选天然和合成的候选分子。(C)2010年爱思唯尔公司All rights reserved.
Suppression of the growth hormone/insulin-like growth factor-I pathway in Ames dwarf (DF) mice, and caloric restriction (CR) in normal mice extends lifespan and delays the onset of age-related disorders. In combination, these interventions have an additive effect on lifespan in Ames DF mice. Therefore, common signaling pathways regulated by DF and CR could have additive effects on longevity. In this study, we tried to identity the signaling mechanism and develop a system to assess pro-longevity status in cells and mice. We previously identified genes up-regulated in the liver of DF and CR mice by DNA microarray analysis. Motif analysis of the upstream sequences of those genes revealed four major consensus sequence motifs, which have been named dwarfism and calorie restriction-responsive elements (DFCR-REs). One of the synthesized sequences bound to hepatocyte nuclear factor-4 alpha (HNF-4 alpha), an important transcription factor involved in liver metabolism. Furthermore, using this sequence information, we developed a highly sensitive bioassay to identify chemicals mimicking the anti-aging effects of CR. When the reporter construct, containing an element upstream of a secreted alkaline phosphatase (SEAP) gene, was co-transfected with HNF-4 alpha and its regulator peroxisome proliferator-activated receptor (PPAR) gamma coactivator-1 alpha (PGC-1 alpha). SEAP activity was increased compared with untransfected controls. Moreover, transient transgenic mice established using this construct showed increased SEAP activity in CR mice compared with ad libitum-fed mice. These data suggest that because of its rapidity, ease of use, and specificity, our bioassay will be more useful than the systems currently employed to screen for CR mimetics, which mimic the beneficial effects of CR. Our system will be particularly useful for high-throughput screening of natural and synthetic candidate molecules. (C) 2010 Elsevier Inc. All rights reserved.