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J Gerontol A Biol Sci Med Sci. Jan 2011; 66A(1): 97–108.
Published online Oct 5, 2010. doi:  10.1093/gerona/glq168
PMCID: PMC3032519

Design and Conduct of the CALERIE Study: Comprehensive Assessment of the Long-term Effects of Reducing Intake of Energy

Abstract

Background.

In a robust and consistent manner, sustained caloric restriction (CR) has been shown to retard the aging process in a variety of animal species. Nonhuman primate studies suggest that CR may have similar effects in longer-lived species. The CALERIE (Comprehensive Assessment of the Long-term Effects of Reducing Intake of Energy) research program is the first systematic investigation of CR in nonobese human beings. In the phase 2 study, it is hypothesized that 2 years of sustained CR, involving a 25% reduction of ad libitum energy intake, results in beneficial effects similar to those observed in animal studies. This article presents the design and implementation of this study.

Methods.

The study is a multicenter, parallel-group, randomized controlled trial. A sample of 225 participants (22.0 ≤ body mass index [BMI] < 28.0 kg/m2) is being enrolled with 2:1 allocation to CR.

Results.

An intensive dietary and behavioral intervention was developed to achieve 25% CR and sustain it over the 2 years. Adherence is monitored using a doubly labeled water technique. Primary outcomes are resting metabolic rate and core temperature, and are assessed at baseline and at 6-month intervals. Secondary outcomes address oxyradical formation, cardiovascular risk markers, insulin sensitivity and secretion, immune function, neuroendocrine function, quality of life and cognitive function. Biologic materials are stored in a central repository.

Conclusions.

An intricate protocol has been developed to conduct this study. Procedures have been implemented to safeguard the integrity of the data and the conclusions drawn. The results will provide insight into the detrimental changes associated with the human aging process and how CR mitigates these effects.

Keywords: Aging, Caloric restriction, Resting metabolic rate, Markers of inflammation, Randomized controlled trial

PRIMARY aging is the inevitable deterioration of cells and tissues independent of illness and environmental factors. Secondary aging is the decline resulting from external influences such as disease and detrimental lifestyle. Both are important codeterminants of health (1). The attenuation of primary aging increases maximal life span, whereas delays in secondary aging mostly affect average life span. Caloric restriction (CR) is the only known intervention that slows both primary and secondary aging in laboratory animals.

Since the original discovery by McCay and colleagues (2) in the 1930s, CR has consistently been shown to extend life spans across a range of animal species including yeast, nematodes, flies, fish, and rodents (3,4). Although findings on lifespan in longer-lived species including nonhuman primates are as yet inconclusive (5), CR monkeys display a substantially reduced age-related morbidity (68). In humans, information is lacking from controlled trials of the effects of CR on surrogate markers believed to be indicative of primary aging (9). The only evidence comes from observational studies in longer-lived humans (10) and individuals who self-impose CR (11,12).

Although the exact mechanisms are not fully understood (4,13), CR has been shown in animal studies to reduce metabolic rate and oxidative damage and improve markers of age-related diseases such as insulin resistance for diabetes (14). Studies in rhesus monkeys suggest that prolonged CR opposes many age-associated pathophysiological changes (6,15,16). Because the benefits associated with prolonged CR are important to the health and survival of humans, it has become an important research objective to assess the feasibility, safety, and health-related effects of prolonged CR in well-controlled human trials.

CALERIE (Comprehensive Assessment of the Long-term Effects of Reducing Intake of Energy) is a research program performing the first clinical trials evaluating the effects of CR on the biomarkers of aging and longevity in nonobese human beings. Phase 1 of the program was conceived as site-specific, short-term pilot studies. The designs of these studies have been published elsewhere (9,17,18), and although they applied different eligibility criteria, different interventions (some involving exercise components), and focused on different outcome measures, they established the feasibility of a carefully controlled CR intervention in this population. Publications from these studies have detailed the short-term effects of CR on resting metabolic rate (RMR) (9), cardiovascular (19) and diabetes (2022) risk factors, body composition (18,23), and cognitive function (24).

The following lessons were learned from these studies. First, an intricate and detailed screening process is required to screen out volunteers unlikely to persevere with the rigors of the CR intervention over the full 24 months. A carefully constructed CR intervention is essential but must be flexible enough to allow different approaches to achieve CR. Intensive contact at the beginning is critical when new habits are being formed and recidivism is likely. Equally, feedback must be provided to participants throughout the intervention so that corrections can be made while problems are small. A detailed quality control process must be applied to ensure that all study procedures are conducted consistently and reliably across sites and over time. Finally, although no safety issues emerged in these studies, a proactive safety surveillance protocol is no less important in a longer-term study of this intervention.

Building on these lessons, the phase 2 study is conceived as a single-protocol, multicenter, randomized controlled trial (RCT) investigating the effects of sustained CR over a 2-year interval. The purpose of this article was to report its main design features. We describe the study population and eligibility criteria, provide an overview of the CR intervention, summarize the outcome evaluations and the frequency with which they are carried out, and describe monitoring procedures to safeguard participant safety.

SPECIFIC AIMS AND HYPOTHESES

The overall aim of the CALERIE phase 2 study is to test the hypothesis that 2 years of sustained CR, involving a 25% reduction of ad libitum (AL) energy intake (EI), in healthy men aged 21–50 years (inclusive) and healthy women aged 21–47 years (inclusive) results in the same beneficial effects as observed in animals subjected to similar levels of CR. The primary hypotheses is that CR results in metabolic adaptation as defined by (a) a reduction in RMR adjusted for changes in body composition and (b) a reduction in core body temperature. A reduction in metabolic rate has been proposed as one of the mechanisms by which CR slows aging, possibly by reducing oxidative damage associated with EI in excess of body energy needs. Secondary aims are to test whether CR reduces serum triiodothyronine concentrations and reduces inflammation as determined by plasma tumor necrosis factor-α concentration. Triiodothyronine is a potential mediator of the predicted metabolic adaptation and provides insight into the mechanism of this hypothesized primary adaptation to CR. Reduction of inflammation is one of the adaptive responses suggested to mediate the salutary effects of CR on the aging process in rodents. The study is also investigating the safety implications of sustained CR in humans. A number of exploratory aims address the mechanisms by which CR mediates its biologic effects at cellular and subcellular levels. They are summarized in Table 1. Biologic samples (plasma, biopsy samples, circulating cells, and urine) are being stored in a repository for future mechanistic studies.

Table 1.
Exploratory Aims in CALERIE (Comprehensive Assessment of the Long-term Effects of Reducing Intake of Energy)

METHODS

Key Design Considerations

Almost all of the data in animal studies were obtained when CR had been sustained for long durations of time (relative to the life span of the organism), with body weight relatively stable for much of the CR period (2527). Outcomes observed during the initial weight loss phase in this study will likely parallel results commonly reported in short-term weight loss studies in overweight individuals (including the CALERIE phase 1 studies) and are therefore unlikely to produce new information. Sustained CR beyond the point when weight stability is firmly established, however, provides key data on whether CR effects become stable or whether early effects are transitory. From the phase 1 results, even with successful adherence, it may take a year (or longer) for the initial weight loss to transition to weight stability. Thus, a 2-year intervention period was selected to provide a significant period of weight stability.

Based upon findings from animal studies, a higher level of CR might be expected to result in greater biologic effects. However, CALERIE participants are not obese, and some are not even overweight, so that the health-related motivation to adhere to the intervention may be weaker for these participants (28). Thus, 25% CR was selected as optimal because it is likely to be sustained at a constant level over the entire 2 years yet at the same time produce measurable physiological effects.

The CALERIE phase 2 study is therefore structured as an intensive dietary and behavioral intervention focused on achieving and maintaining 25% CR throughout the 2-year interval. The intervention staff includes psychologists and registered dietitians, and their role is to deliver the CR intervention in a structured and consistent manner across the sites and over time. The intensity of the behavioral components of CALERIE exceeds that in most other CR studies due to the duration of the study and the importance of achieving the target CR level in the first year. Thus, frequent participant contact is provided throughout the intervention. Our approach is outlined in the following and described in detail in a separate manuscript.

Study Organization

The study is funded by the National Institute on Aging of the U.S. National Institutes of Health, and the full organizational structure is depicted in Figure 1. There are three clinical sites located at the Pennington Biomedical Research Center, Washington University Medical Center, and Tufts University. The Duke Clinical Research Institute serves as the coordinating center (CC). A steering committee is the main governing body and consists of the principal investigators of the clinical sites and the coordinating center as well as the National Institute on Aging project scientist. Other committees include a planning committee, quality control committee, intervention design and delivery committee, and so on. A variety of specialized procedures are conducted with study participants and interpreted at central laboratories and reading centers. A full description of all CALERIE constituents is found in Appendix 1.

Figure 1.
Organizational structure for the CALERIE (Comprehensive Assessment of the Long-term Effects of Reducing Intake of Energy) research group. Note: Biochem Lab = biochemistry ...

Basic Study Design

The CALERIE phase 2 study is designed as a multicenter, parallel-group, randomized controlled trial. A sample of 225 participants is being enrolled and assigned at random to the CR intervention or the AL control group. A 2:1 allocation ratio in favor of CR is being applied to maximize the number of individuals receiving the intervention of greater scientific interest. There is no gradual ramping of CR, and the 25% energy reduction applies for the entire 2-year period. CR participants are permitted to vary the composition of their diets as needed or desired provided that nutritional adequacy is maintained. A comprehensive set of outcome evaluations is carried out before randomization, with follow-up evaluations at months 1, 3, 6, 9, 12, 18, and 24 after starting the intervention. It is expected that 10% of study participants will drop out or be withdrawn from the intervention in each of the 2 follow-up years, so that a sample of approximately 180 participants is expected to complete the study.

Study Population and Eligibility Criteria

Healthy individuals from both genders and all races are eligible to participate. Men must be between 21 and 50 years of age, whereas women must be between 21 and 47 years of age (to avoid menopause in most women), and all participants must have a BMI in the range 22.0 ≤ BMI < 28.0 kg/m2.

The lower age limit was chosen because adult height in normal healthy individuals is attained by age 18 and peak bone mineral accrual usually occurs by age 21 (29,30). The upper limit was chosen because in animal studies, data from CR initiated after 50% of average life span (the human equivalent of 38 years) are sparse and conflicting. However, to ensure an adequate recruitment base, the upper limit is restricted to the ages shown previously. Consistent with animal studies (31), the upper BMI limit was selected to exclude obese individuals. The lower BMI limit was selected primarily for safety reasons. That is, the commonly used standard of underweight is BMI less than or equal to 18.5 kg/m2, and the lower limit provides an adequate buffer from this threshold as participants lose weight over the 2-year intervention.

Exclusion criteria are detailed in Table 2. In general, volunteers are ineligible if they have significant medical conditions (eg, history or clinical manifestation of cardiovascular disease and diabetes), abnormal laboratory markers (eg, elevated potassium, or hemoglobin and hematocrit levels below their respective lower limits of normal), psychiatric or behavioral problems (eg, history or clinical manifestation of any eating disorders, Beck Depression Inventory [BDI] (32) score ≥20), and regular use of medications except oral contraceptives. Women must practice an acceptable form of contraception; breast-feeding and pregnant women are excluded.

Table 2.
Exclusion Criteria

Individuals engaged in heavy physical activity (eg, jogging, running, or bicycling) for 30 minutes or more, five or more times per week, are also excluded to avoid the confounding effects of an increase in energy expenditure. Specifically, from the phase 1 studies, we anticipate that 25% CR might lead to an intentional decrease in regular physical activity, so that individuals training for competitive sporting events are excluded. The resulting change in activity energy expenditure would complicate and confound the determination of EI (and study adherence) using the doubly labeled water (DLW) method over the course of the study. Volunteers partaking at the recommended level of physical activity according to the Physical Activity Guidelines for Americans (150 minutes of moderate-intensity exercise or 75 minutes of vigorous-intensity exercise per week) would not meet this standard and are not excluded from the study.

Recruitment and Screening Strategies

Recruitment is continuous, and includes media advertising, direct mail, health promotion events, databases, Internet Web sites, and referral sources. An initial telephone screening requests the volunteer’s age, height, and weight (from which BMI is calculated), as well as basic medical information. Volunteers who are clearly ineligible are screened out at this point. Then, a staged screening process consisting of a variety of biologic and behavioral assessments is undertaken over a series of three clinic visits as shown in Table 3. The behavioral screening focuses on factors related to safety and the potential for CR adherence and retention over the 2 years. Food allergies and other special diet concerns that could affect participation in the study (eg, alcohol consumption) are evaluated by a nutritionist. A 14-day food record is completed to assess the participant’s ability to provide complete and detailed information over a significant interval of time. At the end of the screening process, a multidisciplinary team of behavioral experts, nutritionists, and other clinical staff scrutinizes the candidate’s entire safety and behavioral profile and determines if he or she is suitable for inclusion in CALERIE.

Table 3.
Procedures Carried Out During the Staged Screening Process

Randomization and Blinding

Participants are assigned at random to the AL or CR treatment arms. Randomization is stratified by sex and BMI within each clinical site, with BMI dichotomized into normal weight, that is, 22.0 ≤ BMI < 25.0 kg/m2, versus overweight, that is, 25.0 ≤ BMI < 28.0 kg/m2. Within each stratum, individuals are allocated in a 2:1 ratio in favor of the 25% CR intervention. Randomization sequences within each stratum were generated a priori by the coordinating center using a permuted block randomization technique (38). Actual treatment assignment is carried out centrally using a telephone-based, interactive voice-response system (39). Given the nature of the CR intervention and control, it is not possible to blind participants or staff to the treatment assignments. Nevertheless, intervention staff have been distinguished from evaluation staff, and cross-communication between the two is circumscribed.

CR Intervention

A complete description of the CR intervention, with specific details concerning the participant materials, subject monitoring, and staff interventions to maintain adherence, is provided in a separate manuscript, and a summary is provided here.

The intervention is a 25% caloric reduction from baseline AL EI. As described in the following, total energy expenditure (TEE) is determined by DLW to assess baseline EI (40). This is carried out specifically for each participant, and the 25% energy reduction prescription for that participant is based on this calculation. Moreover, CALERIE is not prescribing a specific, “one-size-fits-all” dietary composition. Instead, participants make dietary selections (with the advice of intervention staff) that best allow them achieve the CR goal. Participants are allowed to vary their dietary choices as needed or desired over the course of the intervention provided that nutritional adequacy is maintained.

Thus, the overall intervention strategy uses an intensive behavioral approach coupled with dietary modifications anticipated to enhance adherence to CR. Intervention staff include psychologists and nutritionists (hereafter referred to as “counselors”) who supervise the delivery of the CR intervention in a structured and consistent manner. Behavioral strategies known to be effective in long-term weight loss studies (41,42) in tandem with dietary composition changes known to enhance satiety and reduce hunger (43) are applied.

Individual counseling is the primary mode for delivering the intervention; group counseling provides social support and an important secondary source of education. During individual sessions, the counselor provides customized information to assist with reaching and sustaining the CR goal. Group sessions are provided using an “open enrollment” process, which allows participants to begin with modules that cover information appropriate for their length of study participation. Written manuals were created specifically for CALERIE to guide participants and their counselors.

A number of intervention enhancements are provided. First, for the first 28 days of the intervention, complete CR meals are provided accompanied by detailed daily menus. Thus, by direct example, participants receive guidance on achieving their individualized CR goal (44,45). During this period, they also receive detailed training on food portion size estimation, learn to keep a food diary, and are provided a personal data assistant device with diet analysis software, all designed to prepare them for self-selecting their own diets for the duration of the study. Meals provided during the first 28 days are fully adequate in all essential nutrients. When CR diets are self-selected, participants are guided to consume macronutrients in the Acceptable Macronutrient Distribution Ranges of the Dietary Reference Intakes set by the Institute of Medicine’s Food and Nutrition Board (ie, 45%–65%, 20%–35%, and 10%–35% for carbohydrate, fat, and protein, respectively). Nutrient intakes are continually monitored by registered dietitians throughout the trial via daily self-monitoring reports and weighed 6-day food records completed every 6 months. Moderate alcohol consumption is permitted; however, participants are advised of the poor nutritional value of these calories.

As participants transition to self-selected CR, a proactive and comprehensive plan provides an array of supporting services using a “toolbox” approach. Toolbox options focus on psychological, nutritional, and behavioral strategies to improve CR adherence when it begins to falter. The assignment of toolbox options is guided by an algorithmic approach that is iterative and continually updated until adherence is restored.

A Web-based computer tracking system was developed at the coordinating center to provide a standardized approach to implementing the adherence algorithms. It is accessed by the counselors, and tracks the assignment and implementation of toolbox options. It includes expected weight loss trajectories to track the participant's weight against his/her individualized goal. Ratings of hunger, satiety, and desire to eat, as well as self-reported calories from the food diaries are entered providing direct feedback to the participant and counselor. The computer tracking system prompts the counselor when a priori criteria for adherence are not being met, whereupon preprogrammed algorithms suggest toolbox strategies to promote better adherence.

Longer duration of contact with participants has been found to promote health behavior change and favor adherence to diet interventions (46). Intensive participant contact is maintained throughout the study using both individual and group counseling sessions. Moreover, in weekly meetings of each site’s intervention team, computer tracking system reports for each participant are reviewed. Adherence problems and potential barriers are assessed, and toolbox options are selected according to algorithm guidelines.

AL Control Group

Participants assigned to the control group continue on their current diets on a completely AL basis and receive no specific dietary intervention or counseling. They have quarterly contact with study investigators and undertake the outcome assessments at roughly the same schedule as CR participants. Thus, the control group represents the natural history of this participant population as it ages over the 2 years.

Outcome Determinations

Consistent with the overall aims of the study, a comprehensive array of outcome assessments is carried out on CALERIE participants, and a schematic of the schedule of evaluations is provided in Table 4. Some evaluations are only carried out on CR participants, for example, DLW at months 6 and 18. This provides more complete information on energy expenditure so that adherence in the CR group can be determined more precisely during the intervening months. Some procedures, for example, RMR, dual-energy x-ray absorptiometry, nutrient intake, and the Stanford 7-day Physical Activity Recall (33), are carried out concurrently with the DLW studies so that results are coincident with the DLW results.

Table 4.
Schedule of Outcome Evaluations

Primary outcomes are RMR measured via indirect calorimetry over a period of 30 minutes using Vista-MX metabolic measurement system (VacuMed, Ventura, CA) and core temperature recorded every minute over a 24-hour interval. Participants swallow a 8.7 × 23–mm Jonah radiocapsule (Mini Mitter Co., Inc., Bend, OR), and are fitted with the VitalSense Monitor to record the minute-by-minute temperature readings. Temperatures are averaged over specific intervals of interest, for example, 24 hours, daytime, nighttime, and so on. The secondary outcomes, thyroid-stimulating hormone and serum triiodothyronine, are measured using chemiluminescent immunoassays (ADVIA Centaur; Siemens Healthcare Diagnostics, Tarrytown, NY). Tumor necrosis factor-α is measured by multiplex immunoassay (Human Serum Adipokine Panel B; Millipore, Billerica, MA). These assays are carried out at the central biochemistry laboratory. Supplementary data provides a complete summary of all scientific methodologies for the outcome measures in CALERIE.

TEE Measured by DLW

DLW is used to estimate TEE in CALERIE participants. It is noninvasive and nonrestrictive, and has been shown to provide a valid measure of actual TEE under free-living conditions (4750). The DLW method involves enrichment of body water with the stable (natural and nonradioactive) isotopes, deuterium (2H) and oxygen-18 (18O), followed by the determination of their monoexponential washout kinetics in urine. It is based on the principle that the disappearance rate of 2H reflects water turnover rate, whereas the disappearance rate of 18O reflects both water and CO2 turnover rates. The difference between the two disappearance rates, therefore, represents the rate of CO2 production. Knowing the respiratory quotient or food quotient, TEE can be calculated from the CO2 production rate. Validation studies (5153) using calorimetry have shown that DLW method provides an accurate assessment of the CO2 production rate and hence TEE.

Two consecutive 14-day DLW studies are conducted with each participant at baseline with the average used to determine AL TEE; from this, the 25% CR prescription for that participant is derived. Urine samples are collected before dosing as well as on days 0, 7, and 14 following dosing, and shipped on dry ice to the central DLW laboratory. There, the samples are prepared for hydrogen and oxygen isotope enrichments measured by gas isotope ratio mass spectrometry using validated procedures (54,55).

Participant Rights and Safety

CALERIE is being conducted according to the ethical principles in the Declaration of Helsinki. The study is registered on ClinicalTrials.gov Website (identifier: NCT00427193). All participants review and sign the informed consent document and Health Insurance Portability and Accountability Act authorization before undertaking any study procedures. Oversight is provided by a data and safety monitoring board. The study protocol (including any revisions) is reviewed and approved by the data and safety monitoring board and institutional review boards at the participating institutions. Periodic reports summarizing participant safety are presented to the data and safety monitoring board for review. Remedial action including modifying study procedures is taken as appropriate.

Protection of participants from risks related to the intervention is of paramount concern to the CALERIE investigators. Meals and meal plans were carefully designed to maintain an adequate and balanced diet in terms of micro- and macronutrient content. Macronutrient recommendations, within ranges of intakes from the National Academy of Sciences Dietary Reference Intake guidelines (56), are followed. A complete daily vitamin and mineral supplement is provided to ensure that participants in both treatment arms meet the current recommendations for these nutrients. A daily calcium supplement of 1,000 mg is also provided. All CALERIE participants are advised of current health recommendations from the surgeon general (Centers for Disease Control and Prevention) for a minimum of 30 min/d of moderate physical activity for a minimum of 5 d/wk.

Clinical laboratory tests including hematology, serum chemistry, and urinalysis are carried out at screening and baseline, and at months 1, 3, 6, 9, 12, and 24. Vital signs are also recorded. A complete physical examination is carried out at screening, baseline, and at months 12 and 24. A heightened surveillance protocol for CR participants is applied for the following medical conditions: anemia, elevations in low-density lipoprotein cholesterol, decreases in bone mineral density, electrocardiogram abnormalities, depression and other mental health disorders, excessive weight loss, and nutritional inadequacy/eating disorders. Detailed and specific definitions for each condition are provided, and criteria for temporarily discontinuing the CR intervention, resuming it, or permanently discontinuing it are described in the CALERIE protocol.

For example, depression is monitored using the BDI (32). A score of 30 or greater (suggestive of a severe depression) at any point during the intervention triggers an interview with the site psychologist to determine whether the CR intervention needs to be permanently discontinued. If 20 ≤ BDI < 30 (suggestive of moderate depression), the BDI is readministered within 1 week. If this score is still suggestive of a moderate depression, the CR intervention is temporarily discontinued and the participant is advised to seek medical help. The CR intervention is only restarted if the BDI decreases to below 20 or a qualified mental health professional indicates that it is safe to resume. If BDI score is still 20 or greater after 1 month of treatment, the CR intervention is permanently discontinued.

Power Calculations

Based on our experience in the phase 1 studies, it was considered feasible with the resources at the three clinical sites to enroll 225 participants. A dropout rate of around 10% per year was observed in the phase 1 studies, so that a sample of approximately 180 participants is expected to complete the study. Standard deviations for the primary and secondary outcome variables were derived from the phase 1 studies. Between-group differences anticipated at the end of the study were informed by the phase 1 studies, the literature and expert opinion. Standard sample size procedures (57,58) were applied with the treatment allocation set to 2:1 in favor of the CR intervention and the type I error rate (two-tailed) set to α = .05.

The results are summarized in Table 5 and indicate that the study enjoys power in excess of 90% for both primary outcomes. Similar calculations were carried out for the secondary outcomes and a number of the exploratory outcomes (not shown). Power in excess of 80% is anticipated for many (but not all) of the these variables including serum triiodothyronine, high-density lipoprotein cholesterol, insulin and glucose, and fat mass and fat-free mass. We note that the main analytic strategy is a repeated measures analysis. It includes all the intermediate time points as well as partial information provided by participants before they drop out. All things being equal, there should be greater power from this analysis, so that these calculations are somewhat conservative.

Table 5.
Power to Detect Anticipated Differences in the Primary Outcomes With a Sample of 180 Completers

DISCUSSION

The first studies of the effect of energy restriction in humans were conducted in lean men by Keys and colleagues (59) in the 1950s. In these classic experiments, lean volunteers received 50% of their habitual calorie intake. Decreased RMR when adjusted for body surface area (−31%), body weight (−20%), and cell mass (−16%) was observed. However, there were indications of malnutrition with deficiencies in many micronutrients, and for this reason, a comprehensive safety surveillance protocol is being applied in CALERIE.

Most of the other studies of the effect of CR on energy metabolism have been carried out in obese populations. In several studies, a very low calorie diet resulted in a decrease in RMR, which remained significant when normalized to body weight or fat-free mass (6062). A meta-analysis of studies in postobese patients found a lower RMR, even after adjustment for body size and body composition (63). Comparable results in the change in RMR and sedentary 24-hour energy expenditure were also obtained among Biospherians subjected to CR for 2 years (64). Part of the adaptation seen in these different studies may be related to the cost of physical activity as elegantly shown by Weigle and Brunzell (65); however, adaptations unaccounted for by changes in body composition or physical activity are also suggested.

Thus, there is evidence for changes in energy expenditure with CR that include decreases associated with loss of weight and lean body tissue, as well as decreases that appear to be independent of body composition change, which are indicative of a metabolic adaptation to CR. Metabolic adaptation provides greater efficiency of energy utilization and is likely to have both a hormonal basis and to be linked to physical manifestations of low energy expenditure such as reduced core body temperature. Core body temperature is known to vary with metabolic rate (66) and is considered a biomarker of longevity in rodents and nonhuman primates (67,68). Preliminary data from CALERIE phase 1 indicated a significant reduction in core body temperature in the treatment groups that included CR (9).

One possible mechanism of the antiaging effects of CR is through its effects on mitochondrial reactive oxygen species production leading to a reduction in oxidative tissue damage. This is consistent with some experimental evidence in laboratory animals. If mitochondrial oxyradical production is a primary factor in aging-associated changes, it is reasonable to hypothesize that CR’s effects on oxyradical damage in mitochondrial DNA, proteins, or lipids, or all, may be more pronounced than its effects on other cellular components. In fact, effects of CR on oxidative damage to mitochondrial DNA have been reported (69). Thus, it seems reasonable to focus attention and resources on oxyradical production and mitochondrial molecular damage as important outcome measures in CALERIE. This also provides the rationale for storing skeletal muscle and adipose fat tissue in a repository for future mechanistic studies.

The CALERIE phase 2 study is the first comprehensive evaluation of the effects of CR in nonobese humans. It assesses the broad spectrum of physiological effects of CR in multiple physiological domains related to primary and secondary aging. It is informed by animals studies and our phase 1 studies, and addresses whether effects similar to those in animal models are realized in human beings. It is sufficiently large and powered to detect CR effects in multiple important physiological outcome domains.

The clear challenge in CALERIE is to obtain and maintain adherence to the 25% CR goal, and a number of processes are being implemented to meet this challenge. First, TEE derived from DLW is being used to provide an estimate of level of CR required for any participant. Second, the CR intervention is based on the well-established understanding that the decrease in total calories, rather than changes in the relative proportion of micronutrients, accounts for the effects of CR (4). Moreover, a variety of tools are provided to monitor adherence and to intervene proactively when adherence begins to falter. Intensive participant contact is maintained throughout the study using both individual and group counseling sessions.

It would certainly be desirable to have a longer-term follow-up comprising a longer portion of the human life span than the 2 years allotted to CALERIE. This would permit a more meaningful comparison between the human and nonhuman primate studies, for example. Similarly, it would be desirable to perform a long-term observational study on participants after they complete the 2-year intervention. However, CALERIE is labor intensive and entails significant expense and resources. It is also burdensome to study participants calling for a complete lifestyle change that runs counter to the usual lifestyle of community-dwelling Americans. Given these factors, the study is limited to 225 participants. However, we believe that the effort and expense will be rewarded by unprecedented insight into the effects of CR in normal and mildly overweight young- to middle-aged human subjects.

FUNDING

National Institute on Aging, National Institutes of Health (U01AG022132, U01AG020478, U01AG020487, and U01AG020480).

SUPPLEMENTARY MATERIAL

Supplementary material can be found at: http://biomed.gerontologyjournals.org/

Supplementary Data:

Appendix 1. Investigators and Staff Participating in CALERIE (Comprehensive Assessment of the Long-term Effects of Reducing Intake of Energy)

The following is a list of the principal investigators (PIs), Coinvestigators (CIs), site intervention leaders (SILs), intervention counselors (ICs), study managers (SMs), project leaders (PLs), study coordinators (SCs), and other staff (OS) participating in the CALERIE study.

Pennington Biomedical Research Center (clinical site)—PI: Eric Ravussin, PhD; CI: Catherine Champagne, RD, PhD; Alok Gupta, MD; Corby Martin, PhD; Leanne Redman, PhD; Steven Smith, MD; Donald Williamson, PhD; SIL: Corby Martin, PhD; IC: Michelle Begnaud, RD; Barbara Cerniauskas, RD; Allison Davis, MS; Jeanne Gabrielle, PhD; Heather Walden, MS; SM: Natalie Currier, RD; Mandy Shipp, RD; SC: Sarah Masters; Melody McNicoll; OS: Shelly Prince, MS, RD; Courtney Brock, RD; Renee Puyau, RD; Conrad Earnest, PhD; Jennifer Rood, PhD; Tiffany Stewart, PhD; Lillian Levitan, PhD; Crystal Traylor, WHNP; Susan Thomas, WHNP; Valerie Toups, LPN; Karen Jones, RN; Stephanie Tatum, RN; Celeste Waguespack, RN; Kimberly Crotwell, LPN; Lisa Dalfrey, LPN; Amy Braymer, LPN; Rhonda Hilliard, LPN; Onolee Thomas, RN; Jennifer Arceneaux, RN; Stacie LaPrarie, RN; Allison Strate, RN; Jana Ihrig, RN; Susan Mancuso, RN; Christy Beard, RN; Alicia Hymel; Desti Shepard; John Correa; Denise Jarreau; Brenda Dahmer; Grace Bella; Elizabeth Soroe; Bridget Conner; Paige McCown; Stephanie Anaya; Melissa Lupo.

Tufts University (clinical site)—PI: Susan B. Roberts, PhD; CI: Sai Krupa Das, PhD; Simin Meydani, PhD; Roger Fielding, MD; Isaac Greenberg, PhD; Anastassios Pittas, MD; Edward Saltzman, MD; Tammy Scott, PhD; SIL: Cheryl Gilhooly, RD, PhD; IC: Kimberly Gerber, PhD; Isaac Greenberg, PhD; Marjory Kaplan, PhD; Christy Karabetian, MA; Russell Kennedy, PhD; Lisa Robinson, RD; OS: Assefa, Senait; Verona Bembridge; Maria Berlis; Scarlett Buer; Robert Carabello; Cherie Campbell; Lauren Collins, RN; Marybeth Doherty, RN; Alicia Freed, RD; Chervonte Hernandez; Gyna Jean-Baptiste, RN; Mary Krasinski, RN; Marie Lim-Lucas, MPH, RD; Ekaterina Maslova; Barbara Maxwell, RN; Jean McShea, RN; Ann Muchowski, RN; Margaret Mulkerrin; Kerry Murphy; Carol Nelsen, RN; Megan O’Neill; Helen Rasmussen, RD, PhD; Brenda Roche; Eneida Roman; Gregory Sproull; Marie St. Victor, RN; Susan Storer, RN; Katherine Strissel, PhD; Stephanie Valliere; Margaret Vilme, RN; Justin Wheeler; Jill Wiley, RN; Fania Yangarber.

Washington University (clinical site)—PI: John O. Holloszy, MD; CI: Luigi Fontana, MD; Sam Klein, MD; Charles Lambert, PhD; B. Selma Mohammed, MD, PhD; Susan Racette, PhD; Dennis Villareal, MD; SIL: Rick Stein, PhD; IC: Karen Cotton, Psy D; Margaret Hof, MS, RD, LD; Cherie Massmann, MA, LPC, NCC; Kathleen Obert, MS, RD, LD; Marni Pearlman, MA, PLPC; Tina M Reising, Psy D; Laura Weber, MSEd, RD, LD; SM: Mary Uhrich, MS; SC: Morgan Schram, MS; OS: Mel Meyer, RN, BSN, CRC; Chelsea Carlen, BS; Lisa Kee, DTR; Barbara Larson, DTR; Mary McFerson, BS, DTR; Rebecca Sabatino, BS; Bridgett Toennies, RRT.

Duke Clinical Research Institute (coordinating center)—PI: James Rochon, PhD; CI: Connie W. Bales, PhD; Carl F. Pieper, DrPH; William Kraus, MD; PL: Katherine M. Galan, RN; OS: Richard Adrian, BS; Eleanor Law Allen, BA; William Blasko, BS; Manjushri Bhapkar, MS; Nikka Brown, BSN; Maria Butts, RN, BSN; Elaina K. Cossin, BS; Jennifer Curry, AAS; Jamie Daniel, BS, MS; Kathleen S. Diemer, RN; Lee Greiner, BS, MS; Darryl Johnson, BS; Cassandra Jones, BSEE; Lauren Lindblad, MS; Luanne McAdams, RN, MSN; Marty Mansfield, BA, PhD; Senthil Murugesan, MS; Lucy Piner, MS, ACSM CES; Christopher Plummer, BS; Mike Revoir, BS; Pamela Smith, RN, BSN; Monica Spaulding, MPH; James Topping, MS.

Baylor College of Medicine (doubly labeled water laboratory)—PI: William W. Wong, PhD; OS: Lucinda L. Clarke, AA; Chun W. Liu, BS; J. Kennard Fraley, MPH.

University of California at San Francisco (dual-energy x-ray absorptiometry reading center)—PI: Ann V. Schwartz, PhD; CI: John Shepherd, PhD; OS: Lisa Palermo, MS; Susan Ewing, MS; Michaela Rahorst; Caroline Navy.

University of Vermont (biochemistry laboratory)—PI: Michael Lewis, MD, MBA; CI: Russell P. Tracy, PhD; OS: Rebekah Boyle, BS, MS; Elaine Cornell, BS; Patrick Daunais, BS; Dean Draayer, PhD; Melissa Floersch, BS; Nicole Gagne, BA; Florence Keating, BS; Angela Patnoad, BS.

University of Cincinnati (nutrition reading center)—PI: Marcia Schmidt, MS, RD, LD; OS: Marcia Gavin BS, RD, LD; Frida Wiener MS, RD, LD; Ashley Hughes, DTR; Laura Benken.

University of Pittsburgh (intervention counseling curriculum)—PI: Amy Otto, PhD.

Data and safety monitoring board—Jeffrey Halter, MD (chair); David M. Buchner, MD, MPH; Patricia Elmer, PhD; Mark Espeland, PhD; Steven B. Heymsfield, MD; Xavier Pi-Sunyer, MD; Thomas Prohaska, MD; Sue Shapses, PhD; John Speakman, DSc; Richard Weindruch, PhD.

National Institute on Aging (primary funding agency)—Evan C. Hadley, MD; Judy Hannah, PhD; Sergei Romashkan, MD.

National Institute of Diabetes and Digestive and Kidney Diseases (cosponsor)—Mary Evans, PhD.

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