Development of a bioassay to screen for chemicals mimicking the anti-aging effects of calorie restriction

Abstract

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 DFCR-RE sequences bound to hepatocyte nuclear factor-4 (HNF-4), 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 and its regulator peroxisome proliferator-activated receptor (PPAR) coactivator-1 (PGC-1), 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.

Pathobiology of Aging Mice and GEM

“…These and other small but long-lived animals have led to exploration of the roles of somatotropic hormones, insulin-like growth factor 1 (IGF-1), and insulin signaling in life span and development of agerelated morbidities. 22,23,52,104,214 Inability to consume sufficient calories due to dental or other physical problems, as well as ''voluntary diet restriction'' due to diets or additives that are not palatable, should be considered as potential contributors to small size and increased life span in relatively undersized mice and is a concern in studies that aim to study or model longevity by mechanisms other than reduced consumption. 69,89,195 Resveratrol and rapamycin are examples of compounds that may extend life span at least in part by mechanisms that mimic caloric restriction.…”

Mechanisms of lifespan extension and preventive effects of calorie restriction on tumor development: Possible link between central neuroendocrine system and peripheral metabolic adaptation

“…HNF-4α is an important transcription factor in glucose and lipid metabolism in the liver and forms a complex with PGC-1α 31,32) . Levels of the HNF-4α/PGC-1α complex are increased in the liver of CR animals, and putative HNF-4RE-containing genes are up-regulated in pro-longevity contexts 4) .…”

“…We previously reported that the putative HNF-4α response element (HNF-4RE) could play a role in the beneficial effects of CR 4) . HNF-4α is an important transcription factor in glucose and lipid metabolism in the liver and forms a complex with PGC-1α 31,32) .…”

“…Therefore, lower glucose levels are likely to be achieved under lower calorie intake conditions. To avoid development of hypoglycemic disorders, the HNF-4α/ PGC-1α pathway must be activated under CR conditions, which would then lead to activation of other target genes involved in stress resistance, and xenobiotic and other detoxification mechanisms 4) . Hence, activation of gluconeogenic pathways in CR animals could induce various metabolic and protective effects that could contribute to increased lifespan and suppression of age-related disorders.…”

“…However, imposing CR on humans may be difficult to achieve, because they would have to experience unbearable hunger. Hence, many researchers are now investigating CR mimetics (CRM), which are compounds that mimic the beneficial effects of CR without actual food restriction [4][5][6][7] . Development of CR mimetics (CRM) could be a magic bullet not only for age-associated disorders, but also for various rare disorders, such as mitochondrial diseases.…”