Peter H. Duffy

Effect of chronic caloric restriction on physiological variables related to energy metabolism in the male Fischer 344 rat

In the present study, a number of physiological and behavioral measures that are related to metabolism were continuously monitored in 19-month-old male Fischer 344 rats that were fed ad libitum or fed a caloric restricted diet. Caloric restricted rats ate fewer meals but consumed more food during each meal and spent more time eating per meal than did rats fed ad libitum. Therefore, the timing and duration of meals as well as the total number of calories consumed may be associated with life extension. Average body temperature per day was significantly lower in restricted rats but body temperature range per day and motor activity were higher in restricted rats. Dramatic changes in respiratory quotient, indicating rapid changes in metabolic pathway and lower temperature, occurred in caloric restricted rats when carbohydrate reserves were depleted. Lower body temperature and metabolism during this time interval may result in less DNA damage, thereby increasing the survival potential of restricted rats. Nighttime feeding was found to synchronize physiological performance between ad libitum and caloric restricted rats better than daytime feeding, thereby allowing investigators to distinguish the effects of caloric restriction from those related solely to the time-of-day of feeding.

Effect of chronic caloric restriction on hepatic enzymes of intermediary metabolism in the male Fischer 344 rat

It is well established that caloric restriction extends life span and significantly retards the rate of occurrence of most age-associated degenerative disease processes. A paucity of data exists relative to the mechanisms by which caloric restriction accomplishes these events. We have examined the effect of caloric restriction in rats on several hepatic enzymes of intermediary metabolism. The activities of glycolytic and supporting enzymes including lactate dehydrogenase, pyruvate kinase, sorbitol dehydrogenase, and alcohol dehydrogenase were all decreased in response to caloric restriction. Fructose 1-phosphate aldolase and creatine phosphokinase were not altered. Likewise, enzymes associated with lipid metabolism (malic enzyme and glycerokinase) were reduced (fatty acid synthetase was reduced, but not to a statistically significant degree). Activities of enzymes supporting gluconeogenesis (glutamate oxaloacetate transaminase, tyrosine aminotransferase, glutamate pyruvate transaminase, glutamate dehydrogenase, amino acid oxidase, malate dehydrogenase, and glucose 6-phosphatase) were either unchanged or increased significantly by caloric restriction. Glucagon levels were decreased. Comparisons between young ad libitum fed and older calorically restricted rats revealed similar but not identical metabolic activity. These results suggest that caloric restriction produces an effect on intermediary metabolism, favoring the role of glucagon and glucose synthesis; but limiting the role of insulin and glucose catabolism in the liver. The former observation provides for the efficient support of peripheral tissues and the latter a level of energy production necessary only for self maintenance. Limited lipid metabolism suggests decreased potential for fatty acid epoxide formation and free radical damage to cellular macromolecules. Additionally, caloric restriction may delay the progressive age associated changes in the activities of some of the enzymes investigated.

Effects of aging and caloric restriction on hepatic drug metabolizing enzymes in the Fischer 344 rat. I: The cytochrome P-450 dependent monooxygenase system

The effects of long-term caloric restriction on the hepatic cytochrome P-450 dependent monooxygenase system were investigated in the 22-month-old Fischer 344 rat. Caloric restriction decreased the age-related changes in hepatic testosterone metabolism, which are associated with demasculinization of the liver. Caloric restriction also increased hepatic microsomal testosterone 6 beta-hydroxylase, lauric acid 12-hydroxylase and 4-nitrophenol hydroxylase activities over corresponding values in both ad libitum fed 22-month and 60-day-old control male rats. This suggests that cytochrome P-450 isozymes, P-450 pcn1&2, P-452 and P450j may be induced by caloric restriction. Such changes in cytochrome P-450 isozyme profiles could result in altered carcinogen activation, radical formation or drug detoxication in the calorically restricted rat.

Effects of aging and caloric restriction on hepatic drug metabolizing enzymes in the fischer 344 rat. II: Effects on conjugating enzymes

The effects of long-term caloric restriction on the hepatic phase II drug metabolizing enzymes were investigated in the male Fischer 344 rat. Rats that had been restricted to 60% of their pair-fed control consumption from 14 weeks post-partum exhibited altered conjugating enzyme activities at 22 months. Caloric restriction significantly reduced the age-related decrease in glutathione-S-transferase activity towards 1,2-dichloro-4-nitrobenzene, but did not significantly alter the age-related changes in UDP-glucuronyltransferase or sulfotransferase activities towards hydroxysteroids. Caloric restriction appeared to increase hepatic microsomal UDP-glucuronyltransferase activity toward bilirubin and gamma-glutamyltranspeptidase activities. These observations suggest that caloric restriction has multiple effects on the hepatic phase II drug metabolizing enzymes in the rat. Such effects may alter hepatic metabolism and activation or detoxification of drugs and carcinogens.

Chronic caloric restriction induces stress proteins in the hypothalamus of rats

The induction of stress proteins (sps) in the hypothalamus of female Fischer 344 rats in response to caloric restriction (CR) and to heat stress was investigated. Caloric restriction was found to elicit sps 27, 34, 70, and 90 in the hypothalamus of both young and old rats while none was found in the hypothalamus of ad libitum (AL) fed controls. Heat stress initiated heat shock proteins (hsps/sps) 27, 70, and 90 in the hypothalamus of the young (AL) fed animals, the same proteins evoked by feeding stress. The same sps were induced in the old (AL) rats although the expression showed substantial decline with age. This reduction was less marked, however, with the old CR rats. Stress protein 34, an infrequently reported protein, was related to feeding and was not induced by heat shock. Recent reports point to the important role sps play in the cellular reaction to stress, as well as their involvement in the higher functions. The findings reported here suggest that sps are involved in the regulatory mechanisms allowing CR animals to tolerate stress related to metabolic substrate deprivation.

The effects of different levels of dietary restriction on aging and survival in the Sprague-Dawley rat: Implications for chronic studies

A study was undertaken to determine the effects of incremental levels of dietary restriction (DR) in rats. Survival, growth, reproductive, and dietary intake (DI) variables were monitored in a chronic study in which male Sprague Dawley (SD) rats (NCTR colony) were fed their ration ad libitum (AL), or DR. The main objectives were to determine if low levels of DR could be used to increase the survival rate of SD rats in the chronic bioassay, and to identify the survival characteristics of a long-lived SD rat strain (NCTR colony). The average life span of AL rats was 115 months. At 104 weeks on study (110 weeks of age), the survival rate for the AL and 10%, 25%, and 40% DR groups was 63.4, 87.5, 87.5, and 97.5%, respectively. The largest increase in survival (24.1%) occurred between AL and 10% DR, indicating that very low levels of DR have a significant effect on survival. Whole-body, liver, prostate, and epididymis weights and body length were decreased by DR, whereas brain weight, testicular weight, and skull length were not altered by DR. Rats from the NCTR colony were found to be ideal for chronic studies because they are much longer-lived than other SD stocks. Although the 104-week survival rate for these SD, non-obese AL rats exceeds the FDA’s “Redbook” survival guideline (> 50%) for chronic bioassays, the use of DR is advocated because it reduces individual variability in body weight.

Role of Glucocorticoids and “Caloric Stress” in Modulating the Effects of Caloric Restriction in Rodentsa

Caloric restriction (i.e., the balanced reduction of the protein, carbohydrate and fat content of the diet without reduction of its micronutrient content) has been shown, in rodents, to be an extremely powerful modulator of a broad spectrum of age-associated degenerative diseases as well as life span.’-5 Caloric restriction has been found to delay the occurrence of many age-associated neoplastic diseases or to slow their progression, often to such an extent that the clinical expression of the diseases is eliminated.* Since reducing individual dietary components such as protein or fat without reducing the overall caloric intake is less effective in increasing longevity or suppressing neoplasia than caloric restriction by itself, it is reasonable to assume that the observed effects of caloric restriction are primarily dependent upon a specific reduction in calories and not specific dietary compon e n t ~ . ~ ~ ’ The multiple actions of caloric restriction suggest that it may influence primary aging processes themselves; however, recent examination of pathological data suggests that caloric restriction delays different age-dependent pathological conditions at different rates.8 This observation would argue against the modulation by caloric restriction of a simple fundamental process of cellular aging, but rather, it implies that caloric restriction modifies the organism’s homeostatic set-point to one that is less susceptible to some but not all pathologies. Unfortunately, such theories on the nature of the caloric restriction effect will remain merely speculatory until the precise molecular and biochemical mechanisms by which caloric restriction influences the processes of aging and disease are fully elucidated.

Effects of caloric restriction on rodent drug and carcinogen metabolizing enzymes: implications for mutagenesis and cancer

Caloric restriction in rodents results in increased longevity and a decreased rate of spontaneous and chemically induced neoplasia. The low rates of spontaneous neoplasia and other pathologies have made calorically restricted rodents attractive for use in chronic bioassays. However, caloric restriction also alters hepatic drug metabolizing enzyme (DME) expression and so may also alter the biotransformation rates of test chemicals. These alterations in DME expression may be divided into two types: (1) those that are the direct result of caloric restriction itself and are detectable from shortly after the restriction is initiated; (2) those which are the result of pathological conditions that are delayed by caloric restriction. These latter alterations do not usually become apparent until late in the life of the organism. In rats, the largest direct effect of caloric restriction on liver DMEs is an apparent de-differentiation of sex-specific enzyme expression. This includes a 40-70% decrease in cytochrome P450 2C11 (CYP2C11) expression in males and a 20-30% reduction of corticosterone sulfotransferase activity in females. Changes in DME activities that occur late in life in calorically restricted rats include a stimulation of CYP2E1-dependent 4-nitrophenol hydroxylase activity and a delay in the disappearance of male-specific enzyme activities in senescent males. It is probable that altered DME expression is associated with altered metabolic activation of chemical carcinogens. For example the relative expression of hepatic CYP2C11 in ad libitum-fed or calorically restricted rats of different ages is closely correlated with the amount of genetic damage in 2-acetylaminofluorene- or aflatoxin B1-pretreated hepatocytes isolated from rats of the same age and caloric intake. This suggests that altered hepatic drug and carcinogen metabolism in calorically restricted rats can influence the carcinogenicity of test chemicals.

The EFF the effect of dietary restriction of MYC protooncogene expression in mice: A preliminary study

Abstract The effect of chronic dietary restriction on the expression of the c -myc protooncogene was determined in the livers of a hybrid mouse strain (C57B16 × C3H F1 hybrid) at three time points during a 24-h period: 1 h after lights on (1 HALO), 5 h prior to feeding (12 HALO), and 2 h after feeding (19 HALO). In addition, in whole animals studies, changes in core body temperature were monitored. In mice which had been subjected to a chronic diet restriction (60% of the intake of ad libitum controls), c -myc expression was significantly reduced at 1 HALO and 19 HALO compared to corresponding ad libitum animals. In addition, significant differences in c -myc expression were found between time points, in both the ad libitum and restricted groups, suggesting that myc protooncogene expression in the liver may be regulated in a circadian fashion. C -myc expression may correlate with body temperature, suggestng a possible association with metabolic output.

Rationale for the Use of Dietary Control in Toxicity Studies—B6C3F1 Mouse:

Significant variability in critical study parameters such as tumor incidences and survival, increasing tumor incidence and decreasing survival in common toxicity test models, and agent-induced changes in body weight (BW) and BW distribution all generate concern about the reproducibility, consistency, and equity of chronic toxicity tests used in regulation. These concerns have led to suggestions to control BW in chronic tests by the modulation of dietary intake without inducing malnutrition [dietary control (DC)] thereby minimizing tumor and survival variability both between and within studies. Evaluating the reports of the best controlled set of chronic experiments, the National Toxicology Program bioassay series, from studies initiated from 1981 to 1990, there is an increase in tumor incidence, especially liver tumors, with a consistent increase in BW. The studies are classified as to whether normal or aberrant BW growth curves occur. When the studies with normal growth curves are considered, the variance in the BW at 12 mo on test (BW12) can account for over 50% of the variance in liver tumor incidence. Additional stratification by study type, which alter tumor prevalences, as well as appreciation of housing effects [group housing decreases survival (in male mice) and induces tumors in males and females when compared to individual housing], further increase the strength of the correlations, accounting for up to 90% of the variance seen in tumor incidences. These updated analyses further support the hypothesis that it is the BW variation that is resulting in much of the variability seen in tumor incidences and refine the suggestions for the BW curves used as the desired targets for DC.