The impacts of different dietary restriction regimens on aging and longevity: from yeast to humans

Calorie restriction

Caloric restriction (CR) is a reduction in energy intake below the level of calorie consumption under ad libitum conditions without causing malnutrition. The concept of caloric restriction has been recognized for centuries, with early philosophical and medical texts acknowledging the potential benefits of reduced food intake [2]. Scientific exploration of CR began in the early twentieth century, notably with studies on rats that demonstrated a significant extension of both mean and maximal lifespan through calorie reduction [3]. McCay’s groundbreaking research in the 1930s provided some of the first experimental evidence linking CR to longevity.

Subsequent studies expanded across various model organisms, consistently reaffirming the longevity benefits of CR across multiple species, including yeast, worms, flies, rodents, and primates [4–7]. In yeast, CR typically involves lowering glucose concentrations in the growth medium from 2% to 0.5% or less. This reduction significantly extends both the replicative lifespan (RLS; the number of daughter cells that a yeast mother cell can produce) [8] and the chronological lifespan (CLS; how long non-dividing cells remain viable) [9]. In nematode Caenorhabditis elegans, CR extends lifespan by up to 50%, depending on the method used. Approaches include bacterial dilution (reducing food availability) [10, 11] and genetic manipulation (such as eat-2 mutants with reduced feeding rate) [12]. CR also improves proteostasis and reduces the accumulation of age-related protein aggregates, contributing to improved cellular function and organismal health in worms [13]. In fruit flies, Drosophila, studies have observed CR-induced lifespan extension by 30–50%, particularly when CR is initiated early in life [14, 15]. CR has also been shown to preserve physical activity and delay the onset of age-related decline in reproductive functions in flies [16, 17].

In rodents, the impact of CR is typically investigated by reducing calorie intake to around 60–70% of the ad libitum consumption. Reductions of calorie intake at this level extend both mean and maximum lifespan across various rodent species, including mice and rats [3, 18–20]. Rodent studies have also been instrumental in understanding the long-term effects of CR on disease onset. In studies where rodents are subjected to CR, the onset of age-related diseases, such as cancer, cardiovascular disease, and neurodegeneration, is delayed or attenuated [18, 21, 22]. For example, CR has been shown to reduce the incidence and multiplicity of spontaneous and chemically induced tumors in mice and rats, particularly in tissues such as the liver, lung, and mammary glands [23]. Moreover, CR markedly lowers the development of atherosclerosis [24], one major contributor to cardiovascular morbidity in aging populations. In terms of neurodegeneration, CR has been linked to enhanced synaptic plasticity and decreased accumulation of amyloid-beta, a hallmark of Alzheimer’s disease pathology, in mouse models [25–27].

Research on non-human primates, such as rhesus monkeys (Macaca mulatta) and grey mouse lemurs (Microcebus murinus), has significantly contributed to the understanding of the potential effects of CR on aging and healthspan in humans, as these primates share many physiological and genetic similarities with humans. Results from these studies have demonstrated that CR improved overall health and longevity while delaying the onset of age-related diseases in these primates [28–31]. For example, CR has been linked to a reduced incidence of cancer, cardiovascular diseases, and metabolic disorders [28, 29]. Additionally, it helps preserve brain volume and enhances cognitive function in aged monkeys [28, 32], suggesting its neuroprotective effects.

Research on CR in humans faces challenges due to difficulties in conducting long-term controlled studies. However, evidence gathered from clinical trials, epidemiological research, and observational studies strongly indicates that reducing calorie intake provides significant health advantages [33, 34]. One of the most well-known human studies is the CALERIE (Comprehensive Assessment of Long-term Effects of Reducing Intake of Energy), a 2-year, randomized controlled trial for non-obese individuals [35]. The results showed that moderate caloric restriction (around 25% reduction in daily caloric intake) led to significant improvements in several key biomarkers of aging, such as insulin sensitivity, blood pressure, inflammatory markers, and lipid profiles [36, 37]. Participants in the CALERIE trial also showed improvements in muscle function and immune system performance [38, 39], indicating that CR can have broad, positive effects on tissue maintenance during aging. (All the human clinical studies mentioned here and in the later sections are summarized in Table 2.)

Protein/amino acid restriction

Protein restriction (PR) or amino acid restriction (AAR) refers to dietary interventions that lower total protein intake or limit specific amino acids, respectively, while maintaining overall nutritional adequacy. Common PR strategies include lowering daily protein intake, which also leads to reduced caloric intake, or substituting protein content in the diet with carbohydrates without altering the total caloric intake. The longevity effects of PR have been well-documented in several model organisms, including yeast, fruit flies, and rodents [40]. On the other hand, targeting specific amino acids such as methionine, tryptophan, or branched-chain amino acids (BCAAs) like leucine, isoleucine, and valine has also been demonstrated to increase longevity.

In yeast, limiting the availability of all nonessential amino acids [41] or specific amino acids such as methionine, asparagine, or glutamate extends RLS or CLS independently of calorie intake [42–46]. Due to the lack of a precisely defined food source for cultivating C. elegans in the laboratory, direct implementation of protein or amino acid restriction has not been feasible. However, indirect evidence from worms cultured on metformin-treated bacteria suggests that methionine restriction may be responsible for the observed longevity phenotype in metformin-treated worms [47]. In Drosophila, both protein and amino acid restriction have been shown to extend lifespan, particularly when methionine or BCAA is restricted [48–52]. Grandison et al. demonstrated that reducing dietary protein, while supplementing only methionine, reversed the lifespan-extending effect, highlighting methionine’s central role in aging regulation [53]. In addition to methionine restriction, recent studies have suggested that BCAA restriction may also increase survival in flies [49].

In rodents, reducing dietary protein by 40–83% typically leads to a 10–20% increase in lifespan [54, 55], whereas methionine restriction alone has been shown to extend lifespan by 30–40%, independent of total caloric intake [56–58]. Besides lifespan extension, methionine restriction has been shown to reduce oxidative stress and promote leanness in rodents [59, 60]. Similarly, emerging evidence suggests that restricting BCAAs may promote longevity and improve metabolic profiles in mice [61, 62]. Notably, both protein restriction and BCAA restriction exhibit sex-dependent effects in mice [62, 63]. Finally, dietary restrictions of tryptophan have been reported to extend the lifespan of mice and rats and delay the onset of age-associated diseases in various studies dating back to the 1970s and 1980s [64–66]. Interestingly, a recent study suggests that tryptophan restriction may be an evolutionarily conserved intervention for promoting longevity. The administration of ibuprofen, an inhibitor of tryptophan uptake, extends the lifespan of yeast, worms, and flies [67]. However, the underlying mechanism of how tryptophan restriction may exhibit its longevity benefits remains largely unknown.

Direct evidence linking PR to lifespan extension remains limited in humans. However, accumulating data suggest that reducing total protein intake, particularly from animal sources, and specific amino acids may yield metabolic benefits and promote healthy aging. Epidemiological studies, such as analyses of data from the National Health and Nutrition Examination Survey (NHANES), have shown that adults aged 50–65 consuming high levels of protein (≥ 20% of daily calories) had a 75% increase in overall mortality and a fourfold increase in cancer mortality, compared to those with moderate protein intake [68]. Interestingly, these associations were not observed in individuals over 65, highlighting the importance of age-specific recommendations. Importantly, the source of protein appears to matter; plant-based proteins were associated with reduced mortality risk, while animal-based protein was linked to increased risk [69]. Short-term intervention trials in humans further support the health benefits of protein restriction. For instance, randomized controlled studies have shown that reducing protein intake improves glucose homeostasis [70]. A recent study by Nicolaisen et al. demonstrates that when lean and healthy men followed a low-protein diet (meeting minimum requirements) for 5 weeks, they tended to consume more calories, either from fats or carbohydrates, to maintain their body weight. Intriguingly, the protein-restricted diet improved whole-body insulin sensitivity when proteins were replaced by carbohydrates, which may involve FGF21 (fibroblast growth factor 21) [71].

Intermittent fasting

Intermittent fasting (IF) has gained considerable attention in recent years, not only as a weight management strategy but also for its potential effects on longevity and healthspan. Intermittent fasting refers to eating patterns that cycle between periods of eating and fasting. Common approaches include alternate-day fasting (ADF) and periodic fasting (PF). The emerging time-restricted fasting (TRF) may also be considered as an alternative form of IF. TRF limits daily food consumption to a specific time window (e.g., 8 h) and fasting for the rest of the day. ADF alternates between regular eating days and fasting days with little or no caloric intake. PF involves restricting calories or complete fasting for longer periods (e.g., 2–5 days) at regular intervals.

Compelling evidence for the longevity-promoting effects of IF comes from research in various model organisms. In organisms such as worms [72] and flies [73, 74], intermittent fasting has been shown to extend lifespan by 20–40%. A variety of IF regimens, including ADF, PF, and TRF, have been tested in mice and rats, yielding robust evidence that IF can enhance longevity and delay the onset of age-related pathologies [75–78]. One of the earliest studies by Goodrick et al. demonstrated that ADF significantly extended both median and maximum lifespan in several strains of mice. These benefits were observed even when fasting was initiated in mid-life, suggesting that IF can be beneficial even if not practiced lifelong. Subsequent studies confirmed that rodents subjected to IF exhibit increased lifespan comparable to those under caloric restriction (CR), although the degree of extension varies with strain, sex, and fasting protocol [79, 80]. IF in rodents consistently results in improved insulin sensitivity, lower fasting glucose and insulin levels, and protection against high-fat diet-induced obesity [81–84]. Rodent studies have also shown that IF reduces the incidence and progression of age-related cancers and neurodegenerative diseases [27, 85–87]. Notably, several studies also indicate that the beneficial effects of IF are not attributable to reduced calorie intake [81, 84, 88]. It is also worth noting that the CR regimens implemented in early rodent studies were often associated with TRF, as the animals were fed once daily and consumed most of their meal within a short timeframe.

Although conclusive evidence that intermittent fasting (IF) extends human lifespan remains absent, numerous surrogate endpoints and health-related outcomes suggest that IF may provide longevity benefits [89–91]. IF elicits significant metabolic improvements. A controlled trial conducted by Sutton et al. in prediabetic men found that limiting food intake to a 6-h window improved insulin sensitivity, even in the absence of weight loss [92]. Complementary findings from Wilkinson et al. showed that 10-h TRF reduced fasting glucose levels and lowered HbA1c (Hemoglobin A1c) in individuals with metabolic syndrome [93]. In the same study, TRF also improved cardiovascular outcomes, such as blood pressure and atherogenic lipid levels, in patients. Recent findings by Kapogiannis et al. indicate that IF enhances executive function and memory while also attenuating the rate of brain aging in older adults [94].

Metabolic reprogramming

At the cellular level, DR promotes a transition from glucose-dependent metabolism toward alternative fuel utilization pathways. Different DR regimens, including CR, PR, and IF, trigger overlapping yet distinct metabolic adaptations. Previous studies in C. elegans and Drosophila demonstrate that CR leads to a metabolic shift toward fatty acid metabolism [95, 96], enhancing the organism’s ability to utilize stored lipids for energy production. Similarly, elevated lipogenesis and lipolysis are observed in mice subjected to CR or IF [97, 98], indicating that dynamic remodeling of lipid metabolism is needed to maintain metabolic homeostasis during periods of nutrient scarcity. It is worth noting that PR also influences lipid metabolism. In flies fed a high-fat diet, PR has been shown to alter lipid metabolic pathways [99]. Dietary methionine restriction persistently increases total energy expenditure and enhances metabolic flexibility while increasing uncoupled respiration in both fed and fasted states [100]. Enhanced fatty acid oxidation has long been linked to various longevity pathways [101, 102]. Research in C. elegans and Drosophila has also demonstrated that genetic upregulation of lipolytic pathways and fatty acid β-oxidation contributes to lifespan extension [103, 104], suggesting that a metabolic shift toward more efficient energy utilization may be critical for the longevity benefits.

IF, which involves cycling between periods of feeding and fasting, also leads to elevated ketogenesis, which is not found in CR or PR. Growing evidence supports the health benefits of ketone bodies in animal models [105, 106]. However, Tomita et al. found that ketone body supplementation may produce opposing effects, depending on the timing of administration. Ketone body supplementation prolongs lifespan when given to aged mice but increases mortality when administered early in life [107]. These findings suggest that the broader metabolic reprogramming caused by IF may have a greater impact on promoting longevity than the elevation of ketone bodies alone.

Nutrient-sensing signaling networks

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Competing interests

The authors declare no competing interests.