Previous gene expression profiling studies from the lab of David Sinclair at Harvard Medical School, Boston, had shown that resveratrol opposes most transcriptional changes in the liver of mice fed a high-calorie diet (Baur and Sinclair, 2006). In mammals, both resveratrol and CR activate SIRT1, which regulates cell physiology in various contexts, including glucose metabolism, DNA repair, and apoptosis. SIRT1 has been shown to curb neurodegeneration in mouse models of Alzheimer disease (Kim et al., 2007), apparently by enhancing the non-amyloidogenic arm of amyloid precursor protein (APP) processing mediated by α-secretase cleavage (Qin et al., 2006 and ARF related news story). Having demonstrated resveratrol’s prowess at treating the effects of high-calorie diet in mice as well as its protective role in mouse models of AD, “the question was, are these effects related to the CR effect, or not?” Sinclair told ARF. He and Rafael de Cabo, at the National Institute on Aging in Baltimore, Maryland, are senior authors of a paper that addresses this question in the July issue of Cell Metabolism.

The new study—using freshly isolated RNA, as well as a different microarray platform and newer analysis software—confirms the 2006 liver data and extends it to three additional tissues. First author Kevin Pearson and colleagues studied the effects of resveratrol on mice fed, beginning at 12 months of age, the following diets: standard, every-other-day (EOD) feeding, and high-calorie. The researchers found directionally correlated changes of gene expression in 82 percent (liver), 76 percent (skeletal muscle), 96 percent (adipose), and 64 percent (heart) of functional pathways affected by the resveratrol and EOD regimens. Furthermore, the resveratrol-induced transcriptional changes correlated with functional benefits. “The effects we saw in our mice were just what we’d expect to see if you’re slowing aging—less heart disease, fewer cataracts, greater mobility,” Sinclair said. “These animals were on a healthy diet, but we made them even healthier with resveratrol.”

Working independently and publishing 4 June in PLoS ONE, researchers led by Tomas Prolla at the University of Wisconsin, Madison, report similar results in their microarray analysis comparing transcription profiles induced by CR and resveratrol. First author Jamie Barger and colleagues fed mice from middle age (14 months) to old age (30 months) a control diet, CR diet, or resveratrol-supplemented control diet. The researchers report a “striking transcriptional overlap” of CR and resveratrol (99.7 percent of gene expression changes correlating by direction) in heart, skeletal muscle, and brain (neocortex), and show that both regimens prevent age-related cardiac problems.

Leonard Guarante, an MIT scientist among those who originally proposed that SIRT1 would mediate the benefits of CR eight or nine years ago, is delighted by the new findings. “The notion that there will be mimetics of caloric restriction that people can take in pill form is becoming a reality,” he told ARF in a phone interview. “That’s a big deal. It will have implications for major diseases.”

One such disease—a cluster of aging-related conditions known as the metabolic syndrome, which has been linked to dementia (see ARF related news story), was the focus of a mouse study published 3 July in PNAS online by researchers led by Matthias Tschöp at the University of Cincinnati College of Medicine, Ohio, with collaborators at the Spanish National Cancer Research Center in Madrid. To test the effect of SIRT1 on metabolic damage resulting from a prolonged high-fat diet, lead author Paul Pfluger and colleagues engineered mice with an additional transgenic copy of the entire SIRT1 gene, expressed from its own promoter within its natural genomic context. In these mice, SIRT1 was overexpressed two- to fourfold in a variety of tissues including liver, brown fat, and muscle. This moderate overexpression of SIRT1 protected the mice from lipid-induced inflammation, glucose intolerance, and fatty liver (hepatic steatosis) inflicted by high-fat diet, the researchers report.

Guarente sees these findings as additional support for the notion that activating SIRT1 can mimic the effects of CR. “There are no drugs at all—just a SIRT1 transgenic mouse,” he said, noting “significant overlap in the phenotype of these animals and those with resveratrol treatment.”

Tschöp and colleagues’ data suggest that SIRT1 confers protection from metabolic harm by inducing expression of antioxidant enzymes manganese superoxide dismutase (MnSOD) and nuclear respiratory factor 1 (Nrf1) and reducing expression of proinflammatory cytokines tumor necrosis factor α and interleukin 6.

Valter Longo at the University of Southern California, Los Angeles, cautions that the observed protection may not reflect lasting benefit. “Just because you have more antioxidant enzymes in certain conditions doesn’t mean the mouse has adopted a long-term chronic protective mode,” he told ARF.

As an illustration of this principle, a paper in this month’s Cell Metabolism by Longo’s group and Canadian collaborators at the University of Ottawa, shows apparent contradictions in short-term, context-specific versus long-term, systemic effects of SIRT1 in neurons. Prior work in the Longo lab had shown that deletion of Sir2—the yeast homolog of mammalian SIRT1—increases stress resistance (Fabrizio et al., 2005). But not much was understood about the mechanisms behind these effects and, in particular, whether SIRT1 regulates oxidative stress in the brain, where SIRT1 expression is highest in humans (Michishita et al., 2005). To address these issues, first author Ying Li and colleagues subjected primary rat cortical neurons and other cultured cells to conditions that induce oxidative stress (treatment with hydrogen peroxide or menadione). The researchers found that preincubating the rat neurons with SIRT1 inhibitors (nicotinamide or sirtinol) significantly increased cell survival under both stress-inducing conditions in a dose-dependent manner. Further experiments revealed that the neuroprotective effects of SIRT1 inhibition are linked to increased acetylation and decreased phosphorylation of insulin receptor substrate (IRS)-2, and reduced activation of the Ras-ERK1/2 pathway.

To extend the neuroprotection results in vivo, the researchers examined brains of 18-month-old SIRT1 knockout mice for markers of oxidative damage—protein carbonyl content and lipid peroxidation. They found 17 and 20 percent decreases in each marker, respectively, in SIRT1 knockouts compared with wild-type mice. However, the SIRT1-deficient mice had reduced lifespan under both normal and calorie-restricted conditions. “We’re showing that when you reduce SIRT1 activity in neurons, you have protection, but if you take away all the SIRT1, then you have a negative effect,” Longo said, noting that SIRT1 knockout mice are smaller and have developmental defects in addition to a shorter lifespan. To further dissect SIRT1’s neuroprotective role from its broader systemic effects, he has generated brain-specific SIRT1 knockout mice. Longo told ARF that his lab is doing behavioral studies on these animals, and may cross them with Frank LaFerla's triple transgenic 3xTg-AD line (expressing mutated human APP, presenilin 1, and tau) to see if SIRT1 deficiency in the brain can rescue AD phenotypes.

A paper by Guarente and colleagues in the 1 July issue of Genes and Development adds to the idea that SIRT1-targeting strategies may not always produce the expected changes across different organs. By assaying levels of SIRT1 and its small-molecule regulators NAD and NADH, and assessing phenotypes of a liver-specific SIRT1 knockout mouse on various diets, first author Danica Chen and colleagues report this surprising finding: in the liver, SIRT1 is reduced by CR and activated by a high-calorie diet. The finding suggests that the effects of targeting SIRT1 may be more complex than previously thought.

The effects of targeting SIRT1 may also manifest themselves differently depending on when, during an animal’s lifespan, the treatment is started. In the study led by Sinclair and de Cabo, resveratrol and calorie restriction begun in mice at midlife (12 months) did not affect longevity. Sinclair told Alzforum that his group is beginning new studies in which the dietary regimens are begun at six weeks of age.

In the human testing arena, Sirtris Pharmaceuticals, Inc.—a company Sinclair cofounded in 2004 and that was acquired last month by GlaxoSmithKline—has developed a version of resveratrol with increased stability and bioavailability for treatment of aging-related diseases. This liquid compound is in Phase 2 clinical trials for type 2 diabetes. Other Sirtris small-molecule SIRT1 activators that are unrelated to resveratrol but have 1,000 times its potency (Milne et al., 2007) have recently entered the clinic in Phase 1 human safety trials. The longest any of these compounds have been in humans is three months, a senior director at Sirtris told ARF.

In other age-related news published 29 June in Nature Medicine online, researchers led by Carlos López-Otín at Universidad de Oviedo, Madrid, have extended longevity in a mouse model of premature aging using statins and aminobisphosphonates. Their strategy is intriguing in light of prior studies suggesting that statins—well-known as cholesterol-lowering agents—might help prevent AD by reducing synthesis of isoprenoids (see ARF related news story and ARF news story). In the new work, first author Ignacio Varela and colleagues show that combined treatment with statins and aminobisphosphonates improved age-related defects—including growth retarding, weight loss, and defects of fat metabolism, hair loss, and bone—in Zmpste24-/- mice with a premature aging phenotype. The drugs conferred these benefits presumably by blocking farnesylation and geranylgeranylation.—Esther Landhuis.

References:
Barger JL, Kayo T, Vann JM, Arias EB, Wang J, Hacker TA, Wang Y, Raederstorff D, Morrow JD, Leeuwenburgh C, Allison DB, Saupe KW, Cartee GD, Weindruch R, Prolla TA. A low dose of dietary resveratrol partially mimics caloric restriction and retards aging parameters in mice. PLoS ONE. 2008 Jun 4;3(6):e2264. Abstract

Pearson KJ, Baur JA, Lewis KN, Peshkin L, Price NL, Labinskyy N, Swindell WR, Kamara D, Minor RK, Perez E, Jamieson HA, Zhang Y, Dunn SR, Sharma K, Pleshko N, Woollett LA, Csiszar A, Ikeno Y, Le Couteur D, Elliott PJ, Becker KG, Navas P, Ingram DK, Wolf NS, Ungvari Z, Sinclair DA, de Cabo R. Resveratrol delays age-related deterioration and mimics transcriptional aspects of dietary restriction without extending life span. Cell Metab. 2008 Jul;8(1):1-12. Abstract

Pfluger PT, Herranz D, Velasco S, Serrano M, Tschöp MH. Sirt1 protects against high-fat diet-induced metabolic damage. PNAS Early Edition 2008 July 3. Abstract

Li Y, Xu W, McBurney MW, Longo VD. Sirt1 Inhibition reduces IGF-I/IRS-2/Ras/ERK1/2 signaling and protects neurons. Cell Metab. 2008 Jul;8(1):38-48. Abstract

Chen D, Bruno J, Easlon E, Lin SJ, Cheng HL, Alt FW, Guarente L. Tissue-specific regulation of SIRT1 by calorie restriction. Genes Dev. 2008 Jul 1;22(13):1753-7. Epub 2008 Jun 11. Abstract

Varela I, Pereira S, Ugalde AP, Navarro CL, Suárez MF, Cau P, Cadiñanos J, Osorio FG, Foray N, Cobo J, de Carlos F, Lévy N, Freije JM, López-Otín C. Combined treatment with statins and aminobisphosphonates extends longevity in a mouse model of human premature aging. Nat Med. 2008 Jun 29. [Epub ahead of print] Abstract