Sci. Aging Knowl. Environ., 8 March 2006
Vol. 2006, Issue 6, p. nf7
[DOI: 10.1126/sageke.2006.6.nf7]


Craving an Answer

After 70 years, researchers might be closing in on how calorie restriction extends life

Mitch Leslie

Physiologist Roger McCarter says his colleagues at Pennsylvania State University, University Park, rib him about why he's still working on calorie restriction (CR), the draconian diet that extends life in many organisms. Their attitude is, "You guys should have found out the answer by now," he says.

McCarter and other researchers have been trying to "find out the answer" to how dramatically reducing food consumption increases life span for more than 70 years. Why they persist is easy to grasp: A simple change in diet can prolong life by 30% or more. CR buys extra time for organisms as different as mice and yeast, and preliminary studies have identified possible payoffs for humans. How CR bestows these benefits remains uncertain partly because, like aging itself, the body's response to food scarcity is dauntingly complex. "There's so much going on during CR, it's extremely difficult to find out what's important and what isn't," says physiologist Arlan Richardson of the University of Texas Health Sciences Center in San Antonio. However, some scientists contend that in the last few years, we've taken some big steps toward understanding CR. They argue that although many mysteries persist, we are getting our first taste of the molecular interactions that underlie increased longevity. One of those researchers, molecular geneticist David Sinclair of Harvard Medical School in Boston, predicts that within 10 years, patients will be benefiting from drugs that mimic the effects of CR. Other CR watchers are less optimistic that we're on the verge of cracking this long-lasting puzzle.

The Upside of Hunger . . .

Since scientists first documented CR's life-stretching power in 1935 (see Masoro Review), they've catalogued myriad physiological and molecular benefits from dainty dinners. Sticking to the diet sends quantities of glucose and insulin in the blood plummeting and spurs tissues to become more sensitive to the hormone. All three variables can move in the opposite direction as we age or if we develop diabetes. Under CR, proteins and other cell components incur unusually small amounts of damage from reactive oxygen species (ROS), deleterious byproducts of metabolism that might help induce aging (see "The Two Faces of Oxygen" and Dugan and Quick Perspective). Calorie cutting curbs injury from another type of molecular renegade that might promote senescence: advanced glycation end products (AGEs), sticky derivatives of glucose that can foul up proteins and DNA (see Monnier and Obrenovich Perspective). Deprivation adjusts the pace of apoptosis, or cell suicide, although this process's relevance to aging remains controversial (see "More Than a Sum of Our Cells"). Apoptosis slows in organs such as the brain (1) where cells rarely divide but cranks up in the liver and small intestine. The increase might enable CR animals, which are less cancer-prone, to dispose of damaged cells before they spawn tumors.

Eating less spurs animals to shed fat and, counterintuitively, to become more active. Putting couch-bound humans to shame, elderly rodents on the diet can scamper 4 to 5 kilometers a day in a running wheel, notes McCarter. The regimen fends off cancer and diabetes and shields animals from some neurodegenerative damage. It diminishes buildup of plaques that might spur Alzheimer's disease, for instance (see "Eat Less, Nurture Neurons"). Which of these adjustments prolongs life is a matter of debate.

Researchers are also slugging it out over just how organisms earn their extra time (2). Meticulous demographic studies suggest that at least for some strains of rodents, CR slows the rate of aging, says Richardson. However, some Drosophila devotees and mouse mavens have presented evidence that CR works by procrastination, delaying the onset of aging.

. . . and the Downside

The benefits of CR aren't available to all, however. Some organisms seem to be impervious to the demanding diet. For instance, biochemist Rajindar Sohal of the University of Southern California in Los Angeles and colleagues have shown that CR doesn't add time for houseflies or a common mouse strain (3, 4). Demographer James Carey's group at the University of California, Davis, netted similar results for the pesky Medfly (see "Not in Medflies").

Other studies suggest that older animals are beyond CR's reach. Sohal's group demonstrated that eating less killed elderly mice instead of prolonging their golden months (4). And research also has shown that weight loss in elderly people increases the risk of dying (see "Weighty Decisions"). Those conclusions remain controversial. Recent work records gains for graying mice (see "Never Too Old?") and geezer flies (see "Second Chance").

Even in species in which CR is effective, life for a rationed rat or famished fly isn't necessarily happier, even if it is longer. Extra time requires sacrifices (see "Dietary Drawbacks"). Animals living la vida low-cal often reduce or cease reproduction--a disaster from an evolutionary perspective. They can suffer other setbacks that might prove fatal outside of cushy lab digs. Losing fat sounds great if you're trying to squeeze into last year's jeans, but drastic shrinkage--combined with a reduced body temperature--increases vulnerability to the cold. CR animals can perish from temperatures that don't faze normal rodents, says Richardson. The small size of many hungry animals can also be a handicap because bigger eaters might bully them in the scramble for food or mates. Extreme dieting also slows wound healing and might subvert the immune system. People who've tried CR report other irksome side effects. A handy accessory for CR aficionados is a dietary scale to weigh portions. But CR how-tos, found in places such as the Web site of the Calorie Restriction Society, recommend toting around pillows as well, because the loss of body fat can make sitting on a hard surface uncomfortable.

Figure 1
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Longevity by the ounce. A kitchen scale is an essential tool for calorie restricters. [Credit: Joy Skipper/StockFood/Getty Images]

We humans are more pampered--and more adaptable--than lab rodents, so we can dodge some of these side effects. Extreme dieters who get cold can throw on a sweater or turn up the thermostat; if they are worried about reproductive repercussions, they can have children before starting CR. But problems such as reduced immunity aren't as easy to overcome.

CR in Context

How CR extends existence has confounded scientists since they started studying it. In a review published last year, Sinclair (5) counted 10 hypotheses. Some of these proposals were plain wrong. For instance, researchers initially asserted that CR worked its magic by delaying development. That argument collapsed after experiments showed that animals could survive longer even if they didn't start dieting until adulthood. Other hypotheses, such as the idea that CR stretches life span by reducing ROS, might be partly right, Sinclair notes, but they don't account for the range of metabolic changes that animals undergo.

Figure 3
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Hypotheses to explain CR

Even without a comprehensive explanation of how the body translates less food into more time, scientists might be getting a better idea of what type of response the ascetic diet induces. According to a hypothesis that's gaining support, the reaction to CR qualifies as a form of hormesis, the phenomenon in which mild stresses promote longevity (see "Stress for Success"). Moderate doses of radiation or toxins spur organisms to deploy defenses such as heat shock proteins and antioxidant enzymes that make them tougher--and lengthen life. Likewise, food scarcity stresses creatures and triggers a host of protective pathways, says physiologist Edward Masoro, a professor emeritus at the University of Texas Health Sciences Center in San Antonio who now lives in Charleston, South Carolina. The explanation clarifies why a strategy that involves slowing or curtailing reproduction could evolve. Facing an emergency, organisms temporarily shift their resources into self-preservation. Unlike previous theories for CR, such as the notion that reduced food intake drives down metabolic rate, the hypothesis ascribes creatures' responses to active, not passive, changes, says Sinclair. Linking CR with hormesis pushes the field forward by helping scientists define what type of phenomenon they are seeing, he says: "We can now view the physiological changes [of CR] as being part of a survival response."

Recent findings encourage some scientists that they are closing in on at least one of the control pathways that trigger this survival response. "I think we are close to having a mechanism" for CR, says molecular biologist Leonard Guarente of the Massachusetts Institute of Technology in Cambridge. The linchpins of the pathway, he says, are the sirtuin proteins, which promote longevity in a number of species. "It's not everything, but it is one thing," says Guarente, acknowledging that factors in addition to sirtuins might underlie CR.

Not everyone agrees that we are approaching enlightenment. "The field is still muddled," says Masoro. Researchers have focused too much on single genes and pathways, and as a result they underestimate CR's complexity, he says. It likely spurs multiple responses, and different ones will predominate in different species, he says.

Counting Calories or Measuring Nutrients?

The term "calorie restriction" implies that the diet's energy content matters most. However, some work suggests that organisms take their cue from specific nutrients. A series of studies that began in the early 1990s reveals that mice that nibble a diet low in the amino acid methionine live longer than usual, although critics say they might be eating less, too (see "Defining the Diet"). Evolutionary biologist Linda Partridge of University College London and colleagues reported similar results after serving fruit flies cuisine with varying amounts of sugar and yeast. Eating less of the fungi, which provided the flies' only source of protein, resulted in a bigger life-span boost than did cutting back on sweets (6). Changing how often animals eat can elicit the same effect as CR. Alternating periods of deprivation and plenty succeed where normal calorie reductions don't--they extend longevity in Medflies (see "Natural Foods"). Similarly, mice of a strain that responds to CR live longer if they dine every other day, even though they take in a normal amount of calories. Neuroscientist Mark Mattson of the National Institute on Aging in Baltimore, Maryland, and colleagues showed that this regimen increased brain cells' resistance to a nerve-damaging compound (7).

These findings have inspired some researchers to advocate jettisoning the term "calorie restriction" in favor of "food restriction" or "dietary restriction." Masoro counters that for rats at least, his group's studies in the 1980s confirmed that calories are what counts. Hope for unity remains, he says, thanks to the hormesis hypothesis. Whether they stint on calories or nutrients such as methionine, these inadequate meals qualify as stresses, says Masoro.

Finding the Right Path(way)

A fight has broken out among CR researchers about which molecular pathway detects food scarcity and alerts the organism to take action. The battle pits Guarente and Sinclair against Brian Kennedy and Matt Kaeberlein of the University of Washington, Seattle.

Guarente's work suggests that the job of gauging nutrient supplies falls to an enzyme called Sir2p and its relatives, the sirtuins. Sir2p helps shut down genes by pruning acetyl groups from histones, the proteins that DNA coils around in chromosomes. The enzyme draws out a yeast's replicative life span, or how many times it can reproduce, by inhibiting the formation of ribosomal DNA circles that build up and eventually kill the cell. Although the ribosomal DNA loops don't determine longevity in other species, Sir2p's kin do. Its worm counterpart prolongs survival, and researchers are evaluating the mammalian version, SIRT1.

Six years ago, Guarente and colleagues discovered that cutting rations for yeast cells that lacked Sir2p didn't increase life span, implying that the enzyme was essential for CR's powers. They also showed that a compound called NAD, which cells accrue when they break down food, rouses Sir2p (8). Work from Sinclair's lab indicates that Sir2p in a yeast cell responds not to NAD but to nicotinamide, an NAD precursor that shackles Sir2p (see "Hungering for Simplicity" and "UnSIRtainty Principle"). In either case, Guarente says, the findings suggest that the sirtuins assess food availability and unleash protective pathways when times get tough.

What piqued researchers' interest was the possibility of harnessing sirtuins to deliver CR's benefits without its pain. Guarente and Sinclair have each helped found companies (see "'Gero-Tech' Sprouts, But Will It Bloom?") that are trying to identify and commercialize sirtuin stimulators. Sinclair's group at Harvard has already fingered one such compound, an ingredient of red wine called resveratrol, and demonstrated that it lengthens life in yeast, worms, and flies (see "Raise a Glass to Long Life" and "Resveratrol to the Rescue"). Moreover, the researchers found that depriving flies that slurped resveratrol didn't extend life further, indicating that the compound and CR exert their effects through the same pathway.

However, recent findings from Kennedy, Kaeberlein, and colleagues relegate Sir2p to the role of bystander (see "Calorie Restriction Un-SIR-tainty"). The team studied a battery of genetically altered yeast, each of which lacked a different gene. Cells without Sir2p gained more from CR than did cells with the gene, the researchers discovered.

In a follow-up study of the mutant yeast (9), the scientists pinpointed a dozen genes that extended life span. Many of the defects squelched the TOR pathway. Researchers have previously nominated this pathway as a promising candidate for integrating life span and nutrition (see Kapahi and Zid Perspective). It manages mechanisms that affect survival, including the recycling of cellular contents and protein production. Adding further support, Kaeberlein and co-workers found that CR doesn't promote persistence in fungi lacking one of the TOR genes. The team has recently shown that squelching TOR extends the other main measure of yeast longevity, chronological life span, or how long the cell lasts (10). The data don't exclude Sir2p, Kaeberlein says, but "my opinion is that at least in yeast, there's no role for Sir2 in CR."

To those fighting words, Guarente retorts that although TOR might play a part in CR, "overwhelming evidence" supports Sir2p's importance in yeast. Sir2p's dependence on NAD wouldn't make sense if the enzyme weren't responding to nutritional status, he says. He adds that other organisms such as flies (11) require sirtuins to earn CR's advantages. The clashing results might boil down to differences in growing conditions, Sinclair says. He and Guarente put their yeast on a diet of 0.5% glucose, whereas Kaeberlein's fungi had to make do with one-tenth as much sugar. Sir2p and its relatives might run the show when CR is mild, Sinclair says, and TOR intervenes when starvation looms. Another possible explanation is that the groups used different strains of yeast for some of the experiments.

Fat and Mitochondria

Sir2p and TOR are just two of the potential CR contributors that have captured researchers' curiosity. In the CR hierarchy, they would be generals. But which underlings respond to their orders and tweak metabolism and longevity is unknown. Some experts have thrown their weight behind fat. Enthusiasm for the proposal has waxed and waned over the last couple of decades, says Richardson, and currently it's fashionable again. Researchers such as Sohal even contend that the reduced flab might account for all of CR's payoffs, particularly in lab rodents, which often balloon on an all-you-can-gnaw diet. CR works, according to this view, because it slashes the animals' rations to about what they would eat in the wild. Evolutionary biologist Steven Austad of the University of Texas Health Sciences Center in San Antonio disputes the conclusion. CR is so severe that rodents stop reproducing, which doesn't usually happen in nature (see Austad Perspective). That observation suggests that CR is not just mimicking what happens outside the lab door.

Figure 2
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Thinking thin. Researchers have investigated a host of factors that might account for CR's benefits. The factors here are not all acting at the same level and might interact. Sir2p is a NAD-dependent deacetylase. TOR is a nutrient-responsive kinase. [Credit: (Clockwise from the top): Fred Hossler/Visuals Unlimited/Getty Images; Don Fawcett/Visuals Unlimited/Getty Images; David McCarthy/Photo Researchers Inc.]

Regardless of whether typical lab animals outeat their wild cousins, researchers theorize that fat causes problems because it is not inert; it leaks hormones such as leptin and adiponectin that adjust appetite and metabolism. Work by diabetes researcher C. Ronald Kahn of the Joslin Diabetes Center in Boston and co-workers supports the notion that trimming fat underlies at least some of CR's life-stretching effects. The researchers found that mice that lacked the insulin receptor in their fat cells gorged but remained sleek--by rodent standards, at least--and lived longer than normal (see "Lasting Without Fasting"). A study by Guarente and colleagues links SIRT1 to fat shedding. The work suggests that SIRT1 blocks a protein called PPAR{gamma}, stimulating cells to lose fat (see "Counterattack"). To peel back fat's functions, researchers need to determine whether its hormones alter longevity, Guarente notes.

CR researchers have also zoomed in on the mitochondria, the powerhouses that provide the cells' energy but also emit ROS. CR animals suffer less damage from ROS, but researchers don't know why. After an epic battle, they can say that the mechanism almost certainly doesn't involve a reduced metabolic rate, according to gerontologist Brian Merry of the University of Liverpool in the U.K. The question proved so tricky because CR and normal animals differ in a multitude of ways besides weight, including body composition and organ size. Although some holdouts remain, most researchers now agree that metabolism in CR mammals doesn't slow down, he says.

Merry and colleagues nabbed one possible answer to the question of why mitochondria from hungry animals spawn fewer ROS if their metabolic rates don't change. The organelles might spring leaks. Much as a dam generates power by releasing stored water, a mitochondrion harvests energy from a buildup of hydrogen ions, allowing them to trickle out through membrane proteins, the molecular equivalent of the dam's turbines. Merry and colleagues found that mitochondria in CR animals become leakier, and hydrogen ions escape without passing through the molecular turbines, a detour that might reduce ROS production. The leaks close, the researchers learned, if the animals get doses of insulin (12).

The Human Factor

The biggest question about CR--whether people can reap the benefits--remains unanswered. Preliminary evidence is promising. Putting our primate relatives on short rations elicits many of the same salubrious metabolic changes as seen in rodents (see "Monkey in the Middle"), although the longevity data for these animals, which can live more than 20 years, aren't in yet. Moreover, several papers report similar results for folks who aren't waiting for the final word from scientists and have decided to slash their food consumption. Work published in January reveals that the hearts of middle-aged people who restrict their calories appear younger--sporting stretchier walls, for example--than the hearts of their nondieting counterparts (13).

But a key limitation of the human studies is that participants were often CR true believers, known as CRonies, who voluntarily adopted the diet, says Eric Ravussin of the Pennington Biomedical Research Center in Baton Rouge, Louisiana. Even before trimming their food intake, these folks tend to be thinner than average, and their youthful physiology might indicate that they've taken good care of themselves. To eliminate the "self-selection" issue, Ravussin's group has teamed with researchers at Tufts University in Boston and Washington University in St. Louis, Missouri, to conduct the first randomized clinical trials of CR. The researchers have already completed a 1-year preliminary study. The results, some of which are in press, reveal that the dieters improved their sensitivity to insulin, had healthier blood lipid profiles, and showed less oxidative stress, says Ravussin. This fall, the team plans to launch a bigger, longer study, enlisting patients with body mass indexes between 23 and 28--from average weight to just shy of obese--and randomly assigning half of them to a diet with 25% fewer calories than usual. Over the next 2 years, the researchers will measure insulin quantities and sensitivity, blood lipids, and a host of other variables. The study won't last long enough to determine whether CR can extend participants' lives, but it will provide a better indication of whether CR helps humans, Ravussin says.

Plenty of other mysteries about CR linger. For example, researchers don't know why some creatures fail to benefit from reduced rations and whether their recalcitrance tells us anything about how CR works. An important question for the world's graying population is whether there's a cutoff age above which reducing calories becomes harmful. Solving all of CR's mysteries might require researchers to live a little longer.

March 8, 2006

A writer in Portland, Oregon, Mitch Leslie has restricted himself to two brownies a week.

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Citation: M. Leslie, Craving an Answer. Sci. Aging Knowl. Environ. 2006 (6), nf7 (2006).

Science of Aging Knowledge Environment. ISSN 1539-6150