Overview of Project Tasks

As described in the introduction, the PBRC is requesting funding for an additional 5 years to conduct research concerning issues of nutrition relevant to the military. The research as outlined in the preproposal consists of the following:

• measuring energy and water needs of troops in the field,
• providing laboratory support for field studies,
• providing nutrition assessment support in field studies,
• enhancing the nutrition of military menus, and
• developing nutritional strategies for improved military performance under stressful conditions.

A total of nine specific projects were developed by the PBRC to meet the U.S. Army objectives. These specific tasks that the CMNR were asked to review include:

• Clinical Laboratory for Human and Food Samples (Task 1),
• Stable Isotope Laboratory (Task 2),
• Stress, Nutrition, and Mental Performance (Task 3),
• Stress, Sleep Deprivation, and Performance (Task 4),
• Stress, Nutrition, and Work Performance (Task 5),
• Menu Modification/Enhancing Military Diet Project (Task 6),
• Nutrient Database Integration Laboratory (Task 7),
• Stress, Nutrition, and Immune Function Laboratory (Task 8), and
• Metabolic Unit Projects.

Task 1: Clinical Laboratory for Human and Food Samples

Project Summary

This project is headed by Richard T. Tully, Ph.D., with major assistance from Jennifer C. Rood, Ph.D. The primary function of this laboratory under the U.S. Army grant is to provide nutrition laboratory research support to the military nutrition research program at USARIEM. This support includes performing biochemical assessment of nutritional status and performing analyses of nutrient and biochemical substances in foods used in military rations and clinical studies. This laboratory is the central laboratory responsible for analyzing blood, urine, stool, tissue, and food samples. In the future, the laboratory proposes in-field computerized data processing of samples to speed processing time, as well as establishing a database that would be accessible to many Army facilities.

General Comments

This review of the laboratory's operation reveals that Dr. Tully and his staff are well qualified and knowledgeable about the wide range of analytical methods required to support the research program. The procedures used for sample transportation, receipt, storage, preparation, and analysis are clearly defined. Sample analyses are accomplished in a timely manner to meet USARIEM's program needs. The laboratory has a sound quality assurance program and uses standard reference materials to check the adequacy of procedure performance. It is noted that the laboratory follows Good Laboratory Practices regulations and is accredited by the College of American Pathologists. It also is noted that additional method development is underway for tests needed for future research. For the size of its staff, the number of procedures currently being performed by the laboratory is very impressive.

The progress that has been made in rendering this laboratory fully operational is excellent. The laboratory houses state-of-the-art equipment and a well-trained staff. It is evident by the level of clinical laboratory support provided to USARIEM in conducting military nutrition research that this laboratory is vital to the DoD nutrition research program.

Specific Comments, Concerns, and Questions

The staff of the Clinical Research Laboratory has been consulting with the U.S. Department of Agriculture Food Composition Laboratory to obtain expertise in the accomplishment of food analysis. The committee agrees that this relationship is important and that such consultation should continue.

Recommendations

The committee suggests that the laboratory should facilitate additional contacts, such as with the FDA's Regional Laboratory in Atlanta, for nutrient analysis. In the area of clinical methods, it may be useful to establish a relationship with the laboratories at the Centers for Disease Control and Prevention in Atlanta. Participation in interlaboratory collaborative studies to verify new methods under the guidelines of the Association of Official Analytical Chemists is also recommended. These laboratories will provide guidance in the development and implementation of new laboratory tests that will best support the current clinical laboratory projects as well as in the development of the methodology required and proposed for food composition and analysis. As the Menu Modification/Enhancing Military Diets Project develops and broadens in scope, maintaining expertise in methodologic issues will be key.

The committee recommends that the additional expenditure of resources be permitted for the development and implementation of methods necessary for the assessment of immune function (in support of Task 8). The development of these methods will be coordinated by the immunologist who is directing the project on Stress and Immune Function (Task 8).

The committee recommends that efforts associated with expanding the capability of chemical analysis of food composition be restricted to obtaining data that are not available from other reliable sources and which directly support metabolic unit studies.

Finally, the committee recommends continued financial support of the Clinical Research Laboratory, as this laboratory provides services to the Stable Isotope Laboratory, Menu Modification/Enhancing Military Diets Project, and the Nutrient Database Integration Laboratory (see Tasks 1, 2, 6, and 7). Financial support should be maintained at a level consistent with USARIEM needs.

Task 2: Stable Isotope Laboratory

Project Summary

This project is directed by James P. DeLany, Ph.D., and involves the use of the doubly labeled water technique to determine energy expenditure in the field and total body water measurements to monitor changes in body composition. The work of this laboratory represents one of the major service functions of the PBRC for the Army. In the past funding period, the laboratory has been involved in 17 completed studies in collaboration with USARIEM, accomplishing 169 doubly labeled water, 222 water turnover, and 616 total body water determinations. At present, the laboratory is involved in analyses of samples from four additional studies conducted in conjunction with USARIEM.

General Comments

Although no specific new work is proposed in the preproposal presented to the CMNR, it is assumed that this laboratory will continue to be available to the Army on an "as needed" basis to complete similar kinds of measurements in studies yet to be specified. Dr. DeLany has been funded by the Defense Women's Health Research Program to evaluate total daily energy requirements and activity patterns in servicewomen performing a wide range of tasks.

Dr. DeLany indicated that new equipment had been ordered for the laboratory (and is due for delivery) to allow for an increased volume and variety of analyses to be conducted. This equipment will expand the capability of the laboratory to assist in the conduct of studies on various metabolic pathways, such as protein turnover, muscle protein synthesis, and metabolic fuel use during exercise, which could be done in conjunction with other studies proposed to meet the needs of the military (see Tasks 5 and 8).

Specific Comments and Concerns

The lack of integration between this laboratory and others that was suggested by the preproposal is disturbing to the CMNR. Given the directions proposed by other investigators in that preproposal, there are several opportunities for collaboration. These would include investigations such as the ones proposed under Tasks 5 and 8, to determine protein requirements and turnover as well as fuel use and the optimal type of fuel under conditions of intense physical stress that impact on immune function.

Recommendations

The committee recommends that the Stable Isotope Laboratory continue in its service function to assist the Army in its on-going research program. In addition, Dr. DeLany and coworkers should continue to assist the Army with developing experimental designs that ensure the optimal execution of studies using doubly labeled water as well as other stable isotopes. The committee believes that to date this facility has been underutilized, and ample opportunity exists for greater collaboration within the PBRC. The funding of this laboratory should be consistent with the level required to support the Army's nutrition research program.

Task 3: Stress, Nutrition, and Mental Performance

Project Summary

The basic project, headed by Ruth B. S. Harris, Ph.D., involves studies of the impact of stress on neurochemical and behavioral indices of performance using a rodent model. This line of investigation has developed from an interest in ascertaining how stress modifies brain chemistry and function, with the goal of designing appropriate nutritional and pharmacologic strategies to ameliorate the adverse effects of stress. A number of behavioral, nutritional, sensory, and neurochemical findings have been documented, some of which are being prepared for publication and many of which suggest productive future avenues to pursue.

Rodent models have been developed to evaluate the impact of nutritional interventions for their potential to moderate or prevent stress-induced neurochemical changes and their subsequent behavioral deficits. Many models have been evaluated, including a model of acute stress; a model of chronic stress induced by sleep deprivation or by restraint; chronic mild stress induced by exposure to a random mild stressor for several weeks; and an alternate model to study stress and retention utilizing a fixed-ratio training schedule in operant chambers. The stress models currently in use by the Nutritional Neuroscience Laboratory include a chronic stress and an acute stress model. The chronic stress model utilizes 4 days of Rapid Eye Movement sleep deprivation (REMd) to induce a state of stress. The acute stress model measures the effect of either 3 hours of restraint in a small cylinder or immobilization restraint for 30 minutes to 3 hours. This apparatus is a cylindrical enclosure attached to one side of an open field. A rat is placed inside the chamber, and the time to leave the chamber, number of reentries, total time in the chamber, and locomotor activity in the open field are recorded.

The studies proposed for the renewal project period continue several of the earlier lines of investigation into the mechanisms and effects of stress, in addition to a new project entitled "Genetic Markers for Stress Susceptibility" to investigate the interaction of stress and genetic background in the rodent model. A variety of strategies will be used to identify potential candidate genes.

General Comments

This laboratory has had changes in leadership since it was first established 6 years ago. Under the current leadership of Ruth B. S. Harris, Ph.D., significant progress has been made toward the aims proposed in the last project submission (see Appendix III).

While some of the laboratory's approaches will no doubt yield interesting outcomes, the committee feels that some focus is needed owing to the complexity of stress as a physiologic phenomenon and to the various techniques that have been used to evaluate the stress response experimentally.

Specific Comments, Concerns, and Questions

Specifically, the committee believes that a re-evaluation is needed of the models of stress being examined, so as to focus on one or two models that have the greatest potential relevance to the military mission. Once identified, these models should be examined behaviorally, neurochemically, pharmacologically, and nutritionally to identify vulnerable brain functions and potential countermeasures. Unless such an approach is taken, there is the possibility that this important area of investigation will remain unfocused, and thus unlikely to lead to the important insights regarding stress and mental function that are necessary to improve human performance under stressful military conditions.

Recommendations

The committee recommends that the investigators seek expert help in stress, pharmacology, and formulation of research diets to assist in the identification of appropriate pharmacologic and nutritional models and interventions. This may be accomplished in conjunction with the PBRC's 1997–2002 research goals as presented at the site visit, one of which is to bring in visiting professors; it may also be accomplished by holding a 1-day meeting specifically dedicated to examining animal models of stress, that have been used successfully in measuring stress. Speakers would evaluate models of stress with which they are familiar in the context of the stresses experienced in the military.

The committee strongly believes that the PBRC investigators must seek a model which more appropriately fits the human experience of stress; with the use of this model, important insights may be gained regarding the manner by which stress compromises brain function, an issue of particular importance to the military mission. Such insights may ultimately lead to effective nutritional and pharmacological measures to combat the negative effects of stress in a military context. In addition, the committee believes that investigations attempting to identify the genes responsible for controlling the response to stress are premature as experienced investigators and facilities are lacking at this time.

Task 4: Stress, Sleep Deprivation, and Performance

Project Summary

This clinical project, as presented by George A. Bray, M.D.; Richard A. Magill, Ph.D.; and William F. Waters, Ph.D., involves studies in sleep deprived human subjects to ascertain whether chemical and nutritional interventions can minimize the cognitive deficits associated with sleep loss. The investigators have pursued these studies for several years, examining such agents as tyrosine, amphetamine, caffeine, and phentermine. The preliminary presentation of results during the site visit indicates that amphetamine clearly improved several of the cognitive deficits produced by sleep deprivation. Smaller effects were noted for phentermine and caffeine, while unremarkable effects were observed for tyrosine. As presented, the results are consistent with earlier findings for each agent and suggest that weak pharmacologic agents are not particularly potent in improving performance deficits associated with sleep loss.

A proposal has been made to continue this line of investigation and to examine the effect on performance of administering tyrosine alone or in combination with caffeine during a longer period of sleep deprivation. Another proposed study would compare the effects of tryptophan alone with that of tryptophan in combination with melatonin to evaluate whether these agents can improve sleep efficacy during short naps provided intermittently during the sleep deprivation period.

General Comments

At the time of this review, while a large number of studies were reported to have been completed, the data had not yet been thoroughly analyzed statistically.

Specific Comments, Concerns, and Questions

Overall, it would appear that this project has succeeded in fulfilling its mission: it has shown that caffeine may be useful as a nonprescription agent for enhancing performance during sleep deprivation, but that tyrosine is not.

Based on the data collected and analyzed to date, it appears that additional studies are unlikely to lead to new and potent intervention strategies using these agents. Hence, the need to continue such studies is not evident. The committee is aware of other, well-established neuroscience sleep laboratories within the military system that currently are working on similar projects.

Recommendations

The CMNR feels that it would not be in the Army's best interest to continue to provide support for the conduct of the sleep-deprivation studies within the Sleep Laboratory.

Task 6: Menu Modification/Enhancing Military Diets project and Task 7: Nutrient Database Integration Laboratory

Project Summary

In its preproposal, the PBRC has divided the original Menu Modification Project into two projects. The newly named project, Enhancing Military Diets Project, was initiated during September-October 1995 and will continue the recipe development activities using testing procedures similar to those developed for the previous project under the direction of Catherine Champagne, Ph.D. Chef Kelly Patrick has developed new recipes and has begun bench-top testing. The PBRC currently is assembling an acceptability panel that will consist of 50 to 100 panelists who fit the profile of U.S. military personnel to evaluate the recipes. Acceptability and consumption studies will follow at the Army Quartermaster Cooks School at Fort Lee, Virginia.

Past activities in the Menu Modification Project focused on the development of recipes to achieve the stated reduction in fat and cholesterol necessary to achieve the desired goals of no more than 30 percent of total calories from fat, and approximately 300 mg/d of cholesterol, for meals served in the Army garrison dining halls (A Rations). Forty-seven recipes have been tested for acceptability by panels at the PBRC and at Louisiana Tech. Similar methods of acceptability testing done at Fort Polk gave results comparable to those at Louisiana Tech, supporting the use of such nonmilitary panels to predict acceptability in an actual Army feeding situation. The lack of such comparable data was a concern expressed at the last CMNR site visit (see Appendix IV).

A 2-week study also was conducted at Fort Polk in which the new recipes were incorporated into dining hall menus during the second week. Data collected on food consumption in the dining hall plus food records for food consumed off base showed a significant reduction in fat intake (from 34.5 percent to 31.8 percent) with the modified menu compared to the regular menu for food eaten in the dining hall. It was noted, however, that approximately 50 percent of the food intake of personnel in Army facilities is from food consumed off base (outside of military dining facilities). The total fat consumed from these meals averaged 35.4 percent during each period of the study.

Based on interviews with Army personnel, PBRC investigators have proposed a list of modified recipes for testing, with an emphasis on ethnic foods and new breakfast items.

The new task proposed for the Nutrient Database Integration Laboratory involves the development and expansion of activities to assist the Army in estimation of the nutrient content of recipes and menus, as well as in the evaluation of dietary intake records from field studies conducted by USARIEM. Since 1999, the PBRC has participated in a support role in a number of studies. The Nutrient Data Systems Section at USARIEM has been involved in active data collection since 1993. In assessing military diets, a modified visual estimation method as well as food records have been utilized. In January 1995, the PBRC was approached by the Army sponsor to facilitate a more efficient form of collecting and disseminating dietary intake data from garrison and field studies. One such collaborative study which has been undertaken is the Savannah study conducted in July and August 1996. The leader of the data collection team for this project was Dr. Champagne, and the co-leader responsible for data entry was Ray Allen, Ph.D. A database system called MiDAS (Military Data Acquisition System) has been developed for use in the assessment of food intake in military settings. The PBRC staff will integrate all Armed Forces recipes, special formulations (Meals, Ready-to-Eat), and other food formulations into one centralized database system as part of this task.

General Comments for Tasks 6 and 7

In the letter report dated May 12, 1992, the CMNR noted several shortcomings in the Menu Modification Project. These included: (1) a lack of sensitivity to the needs of the military garrison feeding program, (2) inadequate evaluation procedures of modified menus, and (3) lack of interaction between the menu developers and the military menu system (see Appendix IV). The CMNR is pleased to note that PBRC investigators have overcome many of these shortcomings and have established working relationships with Army dietitians at USARIEM and with Army facilities such as Fort Polk and Fort Lee. Future activities with USARIEM include a visit of the PBRC team to the Army Quartermasters Cooks School at Fort Lee, Virginia to observe current training of Army cooks and a meeting with personnel responsible for determining the Army Master menu, as well as meeting with personnel responsible for food purchasing.

The proposed new leader of this project, Alana Cline, Ph.D., has extensive knowledge of military feeding systems and will facilitate further strengthening of the interactions of the PBRC with USARIEM. The past and proposed activities of this project are viewed as highly valuable by the Army and will make possible the accomplishment of tasks they currently are unable to do.

The Nutrient Database Integration Laboratory project will take advantage of the excellent PBRC computer facilities and expertise in dietary intake methodology to provide USARIEM with much-needed assistance in obtaining dietary data faster and more efficiently. Nutrient databases that have been incorporated into the PBRC database laboratory include the Bogalusa Heart Study and USDA Handbook 8 databases, as well as the Army's CAN (Computerized Analysis of Nutrients) database system. The PBRC nutrient database has been validated against the University of Minnesota Nutrition Coding Center database, and also has been utilized in two large, NIH-sponsored multicenter clinical trials.

Specific Comments, Concerns, and Questions for Tasks 6 and 7

Since it was demonstrated that approximately 50 percent of the food intake of personnel in Army facilities represents food consumed outside of military dining halls, it is questionable whether the development of low fat menus alone will achieve the goals of reducing total fat intake. However, any measurable reduction should be considered constructive, and those few individuals selecting all their meals in the military dining halls will benefit substantially.

There is additional concern regarding the micronutrient content of the modified menus. In particular, it is felt that additional attention must be focused on the levels of iron, calcium, zinc, folate, and vitamin B₆ (that is, micronutrients likely to be limiting when the use of ingredients from animal sources is curtailed in favor of lowering fat) provided by these dishes, in addition to the concern about their fat content.

Recommendations for Tasks 6 and 7

The CMNR recommends that estimation of plate waste be included in future field acceptability studies, such as the upcoming study at Fort Bliss, to provide for a more quantitative and qualitative assessment of dietary intakes.

In addition to menu modification, the committee strongly recommends to the PBRC and to the Army the use of nutritional education approaches such as those in the Army's Performance Power nutrition education program in order to achieve dietary intakes that meet military dietary goals.

The CMNR believes the Menu Modification/Enhancing Military Diets Project is relevant to the Army mission and provides valuable support to USARIEM. Since there are a number of nutrient databases in existence, it will be important to integrate these to avoid unnecessary duplication and to assure the integrity of this database. The proposed new task to develop the Nutrient Database Integration Laboratory is the next logical step in the development of a unified nutrition program that may facilitate the evaluation and comparison of dietary intake data with comparable data from the civilian sector at a later date.

Task 5: Stress, Nutrition, and Work Performance and Task 8: Stress, Nutrition, and Immune Function Laboratory

Project Summary

As described by Jeffery J. Zachwieja, Ph.D., Task 5, entitled ''Stress, Nutrition, and Work Performance,'' proposes to use outpatient volunteers to develop repeatable and reliable models to assess effects of carbohydrate delivery, amino acids, and caffeine on work performance and tolerance.

In addition, Dr. Zachwieja proposes to conduct studies using a rat model of stress. These rat studies, as described, will be closely integrated with Task 3 (Stress, Nutrition, and Mental Performance) of Ruth B. S. Harris, Ph.D., of the Nutritional Neuroscience Laboratory and with Task 8 of David W. Horohov, Ph.D., of the Stress, Nutrition, and Immune Function Laboratory. Descriptions of Dr. Horohov's task indicate that many, if not all, of his initial studies will be done in a rat model of stress in an attempt to develop a reproducible and predictable rat model for characterizing the mechanics of stress and immune dysfunction.

General Comments for Tasks 5 and 8

The committee believes that the integration of Tasks 5 and 8 is highly desirable but suggests that the objectives of both tasks would best be accomplished in human subjects. The superb PBRC Metabolic Unit would easily allow these performance studies to be conducted so that studies of balance, body composition, protein turnover, energy metabolism (these last three using stable isotopes), and immune function could all be combined and data obtained under closely controlled conditions. The independent variables would be physical activity and nutritional alterations (including modest-to-severe protein/energy deficiency as experienced in Ranger training). The integration of Tasks 5 and 8 will support a key directive from the Military Nutrition Division at USARIEM to test and evaluate nutrients and pharmacologic agents that may enhance performance. Such studies could build upon previous data obtained in Ranger trainees, as highlighted in previous CMNR publications (IOM, 1992, 1993). These studies documented the effects of demanding physical activity combined with limited food intake and limited sleep, which resulted in a 12 to 15 percent average weight loss over a period of about 9 weeks that was coupled with episodic increases in accidents and infections (IOM, 1993). This situation of low body fat, undernutrition, altered endocrine patterns, and perturbations in immune function suggested the need to develop important and practical nutritional countermeasures. Two important testable issues are whether improving the protein-to-energy ratio, under conditions of modest-to-severe energy deficiency, would improve immune function and physical performance. The carbohydrate delivery studies proposed by Dr. Zachwieja also could be important in the search for effective countermeasures.

Specific Comments, Concerns, and Questions for Tasks 5 and 8

The CMNR feels strongly that the advantages of using the human model for immunological studies far outweigh any advantages of using rat models. As noted earlier in this report, the rat models currently being developed for use in Tasks 3, 5, and 8 leave much to be desired in terms of their direct applicability to military situations involving stress. The use of a human model permits longer-term measurements of antibody response to test antigens as well as delayed dermal hypersensitivity responses. Also, immunological studies of rats can be impaired by lack of species-specific reagents. Both Drs. Zachwieja and Horohov have had extensive experience in conducting studies in human subjects. Combining Tasks 5 and 8 as human studies, and reducing the reliance on rodent studies, may have some economic advantages in the long run.

Recommendations for Tasks 5 and 8

The CMNR believes that the proposed new tasks, once the work being conducted within individual laboratory units becomes integrated and adapted to clinical and metabolic unit studies, will have high degrees of military relevance and should be supported as new areas of research.

Metabolic Unit Projects

Project Summary

An overview of the Metabolic Unit Projects was presented by Donna H. Ryan, M.D.; Jeffery J. Zachwieja, Ph.D.; and Steven R. Smith, M.D. During the past funding period, with input from the Army two metabolic studies were conducted. The first study, "Assessment of Intra-and Inter-Individual Metabolic Variation in Special Operations Forces Soldiers," sought to determine the possible need for individually-designed diets to optimize performance in Special Operations Forces soldiers, who are highly trained and severely stressed. The outcome of the study suggests that the inter-individual variation in response to changes in dietary carbohydrate is small and does not warrant individualization of military diets.

The second study, "Effects of Prolonged Inactivity on Musculoskeletal and Cardiovascular Systems with Evaluation of a Potential Countermeasure," reported by Dr. Smith, was done in conjunction with NASA and USARIEM. It sought to establish a rapidly developing model of microgravity that could then be used to test possible interventions for use by the space program. A model has been developed that involves administering low doses of tri-iodothyronine (T3) in conjunction with bed rest to mimic the effects of inactivity. After 4 weeks, changes in bone resorption markers are similar to those seen in more conventional human models after 4 months.

General Comments

The Metabolic Unit facility, in conjunction with the Stable Isotope Laboratory and Clinical Research Laboratory, provides an outstanding opportunity to control dietary interventions and physical conditions and to perform analyses of related metabolic parameters so that specific questions of interest to the military can be studied effectively. Future plans include testing interventions using testosterone and exercise. The CMNR is concerned with the use of female subjects in further investigations of the musculoskeletal response to microgravity. Until demonstrable countermeasures to maintain bone and muscle mass can be developed in males, female subjects should not be used because of their increased risk related to bone density.

The design of the physical facility and the equipment available for metabolic studies at the Pennington Biomedical Research Center are outstanding, and the available staff are well trained and experienced in the conduct of metabolic studies.

Specific Comments, Concerns, and Questions

No further use of the Metabolic Unit was proposed in the preproposal presented for this funding period. The rationale presented was that use of an animal model would be less expensive than experiments involving the Metabolic Unit. However, in listening to the proposals to evaluate work performance, diet, and immune function, the CMNR believes that much of the proposed work could be conducted better and with more direct application to the needs of the military using human subjects (see Tasks 5 and 8).

Recommendations

The CMNR recommends the use of human subjects in metabolic studies as these studies will be more inclusive and better directed to the needs of the military.

The CMNR recommends seeking the advice of additional experts in the field of bone mineral metabolism in the design and implementation of studies on microgravity, to maximize the efficacy of research while minimizing the risk to subjects. This area of research appears to be of greater importance for NASA than for the Army, and any further development and support should be provided by NASA.